Effect of curcumin analogs onα-synuclein aggregation and cytotoxicity

PubMed Central

Jha, Narendra Nath; Ghosh, Dhiman; Das, Subhadeep; Anoop, Arunagiri; Jacob, Reeba S.; Singh, Pradeep K.; Ayyagari, Narasimham; Namboothiri, Irishi N. N.; Maji, Samir K.

2016-01-01

Alpha-synuclein (α-Syn) aggregation into oligomers and fibrils is associated with dopaminergic neuron loss occurring in Parkinson’s disease (PD) pathogenesis. Compounds that modulate α-Syn aggregation and interact with preformed fibrils/oligomers and convert them to less toxic species could have promising applications in the drug development efforts against PD. Curcumin is one of the Asian food ingredient which showed promising role as therapeutic agent against many neurological disorders including PD. However, the instability and low solubility makes it less attractive for the drug development. In this work, we selected various curcumin analogs and studied their toxicity, stability and efficacy to interact with different α-Syn species and modulation of their toxicity. We found a subset of curcumin analogs with higher stability and showed that curcumin and its various analogs interact with preformed fibrils and oligomers and accelerate α-Syn aggregation to produce morphologically different amyloid fibrils in vitro. Furthermore, these curcumin analogs showed differential binding with the preformed α-Syn aggregates. The present data suggest the potential role of curcumin analogs in modulating α-Syn aggregation. PMID:27338805

The Synaptic Cell Adhesion Molecule, SynCAM1, Mediates Astrocyte-to-Astrocyte and Astrocyte-to-GnRH Neuron Adhesiveness in the Mouse Hypothalamus

PubMed Central

Sandau, Ursula S.; Mungenast, Alison E.; McCarthy, Jack; Biederer, Thomas; Corfas, Gabriel

2011-01-01

We previously identified synaptic cell adhesion molecule 1 (SynCAM1) as a component of a genetic network involved in the hypothalamic control of female puberty. Although it is well established that SynCAM1 is a synaptic adhesion molecule, its contribution to hypothalamic function is unknown. Here we show that, in addition to the expected neuronal localization illustrated by its presence in GnRH neurons, SynCAM1 is expressed in hypothalamic astrocytes. Cell adhesion assays indicated that SynCAM is recognized by both GnRH neurons and astrocytes as an adhesive partner and promotes cell-cell adhesiveness via homophilic, extracellular domain-mediated interactions. Alternative splicing of the SynCAM1 primary mRNA transcript yields four mRNAs encoding membrane-spanning SynCAM1 isoforms. Variants 1 and 4 are predicted to be both N and O glycosylated. Hypothalamic astrocytes and GnRH-producing GT1-7 cells express mainly isoform 4 mRNA, and sequential N- and O-deglycosylation of proteins extracted from these cells yields progressively smaller SynCAM1 species, indicating that isoform 4 is the predominant SynCAM1 variant expressed in astrocytes and GT1-7 cells. Neither cell type expresses the products of two other SynCAM genes (SynCAM2 and SynCAM3), suggesting that SynCAM-mediated astrocyte-astrocyte and astrocyte-GnRH neuron adhesiveness is mostly mediated by SynCAM1 homophilic interactions. When erbB4 receptor function is disrupted in astrocytes, via transgenic expression of a dominant-negative erbB4 receptor form, SynCAM1-mediated adhesiveness is severely compromised. Conversely, SynCAM1 adhesive behavior is rapidly, but transiently, enhanced in astrocytes by ligand-dependent activation of erbB4 receptors, suggesting that erbB4-mediated events affecting SynCAM1 function contribute to regulate astrocyte adhesive communication. PMID:21486931

Cell penetrating peptides fused to a thermally targeted biopolymer drug carrier improve the delivery and antitumor efficacy of an acid-sensitive doxorubicin derivative.

PubMed

Walker, Leslie; Perkins, Eddie; Kratz, Felix; Raucher, Drazen

2012-10-15

Elastin-like polypeptide (ELP) is a macromolecular carrier with thermally responsive properties that can passively accumulate in solid tumors and additionally aggregate in tumor tissue when exposed to hyperthermia. In this study, ELP was conjugated to the anticancer drug doxorubicin (DOXO) and three different cell penetrating peptides (CPP) in order to inhibit tumor growth in mice compared to free doxorubicin. Fluorescence microscopy studies in MCF-7 breast carcinoma cells demonstrated that the three different CPP-ELP-DOXO conjugates delivered doxorubicin to the cell nucleus. All CPP-ELP-DOXO conjugates showed cytotoxicity with IC(50) values in the range of 12-30 μM at 42 °C, but the ELP carrier with SynB1 as the cell penetrating peptide had the lowest intrinsic cytotoxicity. Therefore, the antitumor efficacy of SynB1-ELP-DOXO was compared to doxorubicin under hyperthermic conditions. C57BL/6 female mice bearing syngeneic E0771 murine breast tumors were treated with either free doxorubicin or the SynB1-ELP-DOXO conjugate with or without focused hyperthermia on the tumor. Under hyperthermic conditions, tumor inhibition with SynB1-ELP-DOXO was 2-fold higher than under therapy with free doxorubicin at the equivalent dose, and is thus a promising lead candidate for optimizing thermally responsive drug polymer conjugates. Copyright © 2012 Elsevier B.V. All rights reserved.

Yellow fever virus envelope protein expressed in insect cells is capable of syncytium formation in lepidopteran cells and could be used for immunodetection of YFV in human sera.

PubMed

Barros, Maria C E S; Galasso, Tatiane G C M; Chaib, Antônio J M; Degallier, Nicolas; Nagata, Tatsuya; Ribeiro, Bergmann M

2011-05-27

Yellow fever is an haemorrhagic disease caused by a virus that belongs to the genus Flavivirus (Flaviviridae family) and is transmitted by mosquitoes. Among the viral proteins, the envelope protein (E) is the most studied one, due to its high antigenic potencial. Baculovirus are one of the most popular and efficient eukaryotic expression system. In this study a recombinant baculovirus (vSynYFE) containing the envelope gene (env) of the 17D vaccine strain of yellow fever virus was constructed and the recombinant protein antigenicity was tested. Insect cells infected with vSynYFE showed syncytium formation, which is a cytopathic effect characteristic of flavivirus infection and expressed a polypeptide of around 54 kDa, which corresponds to the expected size of the recombinant E protein. Furthermore, the recombinant E protein expression was also confirmed by fluorescence microscopy of vSynYFE-infected insect cells. Total vSynYFE-infected insect extracts used as antigens detected the presence of antibodies for yellow fever virus in human sera derived from yellow fever-infected patients in an immunoassay and did not cross react with sera from dengue virus-infected patients. The E protein expressed by the recombinant baculovirus in insect cells is antigenically similar to the wild protein and it may be useful for different medical applications, from improved diagnosis of the disease to source of antigens for the development of a subunit vaccine.

Vitamins K interact with N-terminus α-synuclein and modulate the protein fibrillization in vitro. Exploring the interaction between quinones and α-synuclein.

PubMed

da Silva, Fernanda Luna; Coelho Cerqueira, Eduardo; de Freitas, Mônica Santos; Gonçalves, Daniela Leão; Costa, Lilian Terezinha; Follmer, Cristian

2013-01-01

In the last decades, a series of compounds, including quinones and polyphenols, has been described as having anti-fibrillogenic action on α-synuclein (α-syn) whose aggregation is associated to the pathogenesis of Parkinson's disease (PD). Most of these molecules act as promiscuous anti-amyloidogenic agents, interacting with the diverse amyloidogenic proteins (mostly unfolded) through non-specific hydrophobic interactions. Herein we investigated the effect of the vitamins K (phylloquinone, menaquinone and menadione), which are 1,4-naphthoquinone (1,4-NQ) derivatives, on α-syn aggregation, comparing them with other anti-fibrillogenic molecules such as quinones, polyphenols and lipophilic vitamins. Vitamins K delayed α-syn fibrillization in substoichiometric concentrations, leading to the formation of short, sheared fibrils and amorphous aggregates, which are less prone to produce leakage of synthetic vesicles. In seeding conditions, menadione and 1,4-NQ significantly inhibited fibrils elongation, which could be explained by their ability to destabilize preformed fibrils of α-syn. Bidimensional NMR experiments indicate that a specific site at the N-terminal α-syn (Gly31/Lys32) is involved in the interaction with vitamins K, which is corroborated by previous studies suggesting that Lys is a key residue in the interaction with quinones. Together, our data suggest that 1,4-NQ, recently showed up by our group as a potential scaffold for designing new monoamine oxidase inhibitors, is also capable to modulate α-syn fibrillization in vitro. Copyright © 2012 Elsevier Ltd. All rights reserved.

Reduced expression of the NMDA receptor-interacting protein SynGAP causes behavioral abnormalities that model symptoms of Schizophrenia.

PubMed

Guo, Xiaochuan; Hamilton, Peter J; Reish, Nicholas J; Sweatt, J David; Miller, Courtney A; Rumbaugh, Gavin

2009-06-01

Abnormal function of NMDA receptors is believed to be a contributing factor to the pathophysiology of schizophrenia. NMDAR subunits and postsynaptic-interacting proteins of these channels are abnormally expressed in some patients with this illness. In mice, reduced NMDAR expression leads to behaviors analogous to symptoms of schizophrenia, but reports of animals with mutations in core postsynaptic density proteins having similar a phenotype have yet to be reported. Here we show that reduced expression of the neuronal RasGAP and NMDAR-associated protein, SynGAP, results in abnormal behaviors strikingly similar to that reported in mice with reduced NMDAR function. SynGAP mutant mice exhibited nonhabituating and persistent hyperactivity that was ameliorated by the antipsychotic clozapine. An NMDAR antagonist, MK-801, induced hyperactivity in normal mice but SynGAP mutants were less responsive, suggesting that NMDAR hypofunction contributes to this behavioral abnormality. SynGAP mutants exhibited enhanced startle reactivity and impaired sensory-motor gating. These mice also displayed a complete lack of social memory and a propensity toward social isolation. Finally, SynGAP mutants had deficits in cued fear conditioning and working memory, indicating abnormal function of circuits that control emotion and choice. Our results demonstrate that SynGAP mutant mice have gross neurological deficits similar to other mouse models of schizophrenia. Because SynGAP interacts with NMDARs, and the signaling activity of this protein is regulated by these channels, our data in dicate that SynGAP lies downstream of NMDARs and is a required intermediate for normal neural circuit function and behavior. Taken together, these data support the idea that schizophrenia may arise from abnormal signaling pathways that are mediated by NMDA receptors.

High-density lipoprotein-like particle formation of Synuclein variants.

PubMed

Eichmann, Cédric; Kumari, Pratibha; Riek, Roland

2017-01-01

α-Synuclein (α-Syn) is an intrinsically disordered protein in solution whose fibrillar aggregates are the hallmark of Parkinson's disease (PD). Although the specific function of α-Syn is still unclear, its high structural plasticity is key for the interactions of α-Syn with biological membranes. Recently, it has been observed that α-Syn is able to form high-density lipoprotein-like (HDL-like) particles that are reminiscent of self-assembling phospholipid bilayer nanodiscs. Here, we extended our preparation method for the production of α-Syn lipoprotein particles to the β- and γ-Syn variants, and the PD-related familial α-Syn mutants. We show that all human Syns can form stable and homogeneous populations of HDL-like particles with distinct morphologies. Our results characterize the impact of the individual Syns on the formation capacity of these particles and indicate that Syn HDL-like particles are neither causing toxicity nor a toxicity-related loss of α-Syn in PD. © 2016 Federation of European Biochemical Societies.

α-synuclein and synapsin III cooperatively regulate synaptic function in dopamine neurons.

PubMed

Zaltieri, Michela; Grigoletto, Jessica; Longhena, Francesca; Navarria, Laura; Favero, Gaia; Castrezzati, Stefania; Colivicchi, Maria Alessandra; Della Corte, Laura; Rezzani, Rita; Pizzi, Marina; Benfenati, Fabio; Spillantini, Maria Grazia; Missale, Cristina; Spano, PierFranco; Bellucci, Arianna

2015-07-01

The main neuropathological features of Parkinson's disease are dopaminergic nigrostriatal neuron degeneration, and intraneuronal and intraneuritic proteinaceous inclusions named Lewy bodies and Lewy neurites, respectively, which mainly contain α-synuclein (α-syn, also known as SNCA). The neuronal phosphoprotein synapsin III (also known as SYN3), is a pivotal regulator of dopamine neuron synaptic function. Here, we show that α-syn interacts with and modulates synapsin III. The absence of α-syn causes a selective increase and redistribution of synapsin III, and changes the organization of synaptic vesicle pools in dopamine neurons. In α-syn-null mice, the alterations of synapsin III induce an increased locomotor response to the stimulation of synapsin-dependent dopamine overflow, despite this, these mice show decreased basal and depolarization-dependent striatal dopamine release. Of note, synapsin III seems to be involved in α-syn aggregation, which also coaxes its increase and redistribution. Furthermore, synapsin III accumulates in the caudate and putamen of individuals with Parkinson's disease. These findings support a reciprocal modulatory interaction of α-syn and synapsin III in the regulation of dopamine neuron synaptic function. © 2015. Published by The Company of Biologists Ltd.

Yellow fever virus envelope protein expressed in insect cells is capable of syncytium formation in lepidopteran cells and could be used for immunodetection of YFV in human sera

PubMed Central

2011-01-01

Background Yellow fever is an haemorrhagic disease caused by a virus that belongs to the genus Flavivirus (Flaviviridae family) and is transmitted by mosquitoes. Among the viral proteins, the envelope protein (E) is the most studied one, due to its high antigenic potencial. Baculovirus are one of the most popular and efficient eukaryotic expression system. In this study a recombinant baculovirus (vSynYFE) containing the envelope gene (env) of the 17D vaccine strain of yellow fever virus was constructed and the recombinant protein antigenicity was tested. Results Insect cells infected with vSynYFE showed syncytium formation, which is a cytopathic effect characteristic of flavivirus infection and expressed a polypeptide of around 54 kDa, which corresponds to the expected size of the recombinant E protein. Furthermore, the recombinant E protein expression was also confirmed by fluorescence microscopy of vSynYFE-infected insect cells. Total vSynYFE-infected insect extracts used as antigens detected the presence of antibodies for yellow fever virus in human sera derived from yellow fever-infected patients in an immunoassay and did not cross react with sera from dengue virus-infected patients. Conclusions The E protein expressed by the recombinant baculovirus in insect cells is antigenically similar to the wild protein and it may be useful for different medical applications, from improved diagnosis of the disease to source of antigens for the development of a subunit vaccine. PMID:21619598

Structure and properties of a complex of α-synuclein and a single-domain camelid antibody.

PubMed

De Genst, Erwin J; Guilliams, Tim; Wellens, Joke; O'Day, Elizabeth M; Waudby, Christopher A; Meehan, Sarah; Dumoulin, Mireille; Hsu, Shang-Te Danny; Cremades, Nunilo; Verschueren, Koen H G; Pardon, Els; Wyns, Lode; Steyaert, Jan; Christodoulou, John; Dobson, Christopher M

2010-09-17

The aggregation of the intrinsically disordered protein α-synuclein to form fibrillar amyloid structures is intimately associated with a variety of neurological disorders, most notably Parkinson's disease. The molecular mechanism of α-synuclein aggregation and toxicity is not yet understood in any detail, not least because of the paucity of structural probes through which to study the behavior of such a disordered system. Here, we describe an investigation involving a single-domain camelid antibody, NbSyn2, selected by phage display techniques to bind to α-synuclein, including the exploration of its effects on the in vitro aggregation of the protein under a variety of conditions. We show using isothermal calorimetric methods that NbSyn2 binds specifically to monomeric α-synuclein with nanomolar affinity and by means of NMR spectroscopy that it interacts with the four C-terminal residues of the protein. This latter finding is confirmed by the determination of a crystal structure of NbSyn2 bound to a peptide encompassing the nine C-terminal residues of α-synuclein. The NbSyn2:α-synuclein interaction is mediated mainly by side-chain interactions while water molecules cross-link the main-chain atoms of α-synuclein to atoms of NbSyn2, a feature we believe could be important in intrinsically disordered protein interactions more generally. The aggregation behavior of α-synuclein at physiological pH, including the morphology of the resulting fibrillar structures, is remarkably unaffected by the presence of NbSyn2 and indeed we show that NbSyn2 binds strongly to the aggregated as well as to the soluble forms of α-synuclein. These results give strong support to the conjecture that the C-terminal region of the protein is not directly involved in the mechanism of aggregation and suggest that binding of NbSyn2 could be a useful probe for the identification of α-synuclein aggregation in vitro and possibly in vivo. Copyright © 2010. Published by Elsevier Ltd.

Caffeine-water-polypeptide interaction in aqueous solution

NASA Astrophysics Data System (ADS)

Ghabi, Habib; Dhahbi, Mahmoud

1999-04-01

The interaction of caffeine monomer with the synthetic polypeptides polyasparagine (pAg) and polyaspartic acid (pAsp) was studied by UV spectrophotometry. The results show that different types of interactions are possible depending on the nature of polypeptide. The form of the complex was discussed.

Autism-related behavioral abnormalities in synapsin knockout mice.

PubMed

Greco, Barbara; Managò, Francesca; Tucci, Valter; Kao, Hung-Teh; Valtorta, Flavia; Benfenati, Fabio

2013-08-15

Several synaptic genes predisposing to autism-spectrum disorder (ASD) have been identified. Nonsense and missense mutations in the SYN1 gene encoding for Synapsin I have been identified in families segregating for idiopathic epilepsy and ASD and genetic mapping analyses have identified variations in the SYN2 gene as significantly contributing to epilepsy predisposition. Synapsins (Syn I/II/III) are a multigene family of synaptic vesicle-associated phosphoproteins playing multiple roles in synaptic development, transmission and plasticity. Lack of SynI and/or SynII triggers a strong epileptic phenotype in mice associated with mild cognitive impairments that are also present in the non-epileptic SynIII(-/-) mice. SynII(-/-) and SynIII(-/-) mice also display schizophrenia-like traits, suggesting that Syns could be involved in the regulation of social behavior. Here, we studied social interaction and novelty, social recognition and social dominance, social transmission of food preference and social memory in groups of male SynI(-/-), SynII(-/-) and SynIII(-/-) mice before and after the appearance of the epileptic phenotype and compared their performances with control mice. We found that deletion of Syn isoforms widely impairs social behaviors and repetitive behaviors, resulting in ASD-related phenotypes. SynI or SynIII deletion altered social behavior, whereas SynII deletion extensively impaired various aspects of social behavior and memory, altered exploration of a novel environment and increased self-grooming. Social impairments of SynI(-/-) and SynII(-/-) mice were evident also before the onset of seizures. The results demonstrate an involvement of Syns in generation of the behavioral traits of ASD and identify Syn knockout mice as a useful experimental model of ASD and epilepsy. Copyright © 2013 Elsevier B.V. All rights reserved.

Peptide Regulation of Cells Renewal Processes in Kidney Tissue Cultures from Young and Old Animals.

PubMed

Chalisova, N I; Lin'kova, N S; Nichik, T E; Ryzhak, A P; Dudkov, A V; Ryzhak, G A

2015-05-01

Polypeptide complex isolated from calf kidneys stimulates the processes of cell renewal in organotypic kidney tissue cultures from young and old rats. The polypeptide complex enhances expression of proliferation marker Ki-67 and reduces expression of proapoptotic peptide p53 in kidney explants obtained from young and old animals. Short peptides T-31 (AED) and T-35 (EDL) also stimulate proliferation and reduce apoptosis of the kidney cells, but to a lesser degree than the polypeptide complex. The results provide the basis for further investigation of the polypeptide complex as a preparation for the therapy of kidney diseases, including age-related pathologies.

Cross dimerization of amyloid-β and αsynuclein proteins in aqueous environment: a molecular dynamics simulations study.

PubMed

Jose, Jaya C; Chatterjee, Prathit; Sengupta, Neelanjana

2014-01-01

Self-assembly of the intrinsically unstructured proteins, amyloid beta (Aβ) and alpha synclein (αSyn), are associated with Alzheimer's Disease, and Parkinson's and Lewy Body Diseases, respectively. Importantly, pathological overlaps between these neurodegenerative diseases, and the possibilities of interactions between Aβ and αSyn in biological milieu emerge from several recent clinical reports and in vitro studies. Nevertheless, there are very few molecular level studies that have probed the nature of spontaneous interactions between these two sequentially dissimilar proteins and key characteristics of the resulting cross complexes. In this study, we have used atomistic molecular dynamics simulations to probe the possibility of cross dimerization between αSyn1-95 and Aβ1-42, and thereby gain insights into their plausible early assembly pathways in aqueous environment. Our analyses indicate a strong probability of association between the two sequences, with inter-protein attractive electrostatic interactions playing dominant roles. Principal component analysis revealed significant heterogeneity in the strength and nature of the associations in the key interaction modes. In most, the interactions of repeating Lys residues, mainly in the imperfect repeats 'KTKEGV' present in αSyn1-95 were found to be essential for cross interactions and formation of inter-protein salt bridges. Additionally, a hydrophobicity driven interaction mode devoid of salt bridges, where the non-amyloid component (NAC) region of αSyn1-95 came in contact with the hydrophobic core of Aβ1-42 was observed. The existence of such hetero complexes, and therefore hetero assembly pathways may lead to polymorphic aggregates with variations in pathological attributes. Our results provide a perspective on development of therapeutic strategies for preventing pathogenic interactions between these proteins.

Biophysics of α-Synuclein Membrane Interactions

PubMed Central

Pfefferkorn, Candace M.; Jiang, Zhiping; Lee, Jennifer C.

2011-01-01

Membrane proteins participate in nearly all cellular processes; however, because of experimental limitations, their characterization lags far behind that of soluble proteins. Peripheral membrane proteins are particularly challenging to study because of their inherent propensity to adopt multiple and/or transient conformations in solution and upon membrane association. In this review, we summarize useful biophysical techniques for the study of peripheral membrane proteins and their application in the characterization of the membrane interactions of the natively unfolded and Parkinson’s disease (PD) related protein, α-synuclein (α-syn). We give particular focus to studies that have led to the current understanding of membrane-bound α-syn structure and the elucidation of specific membrane properties that affect α-syn-membrane binding. Finally, we discuss biophysical evidence supporting a key role for membranes and α-syn in PD pathogenesis. PMID:21819966

α-Synuclein Aggregated with Tau and β-Amyloid in Human Platelets from Healthy Subjects: Correlation with Physical Exercise

PubMed Central

Daniele, Simona; Pietrobono, Deborah; Fusi, Jonathan; Lo Gerfo, Annalisa; Cerri, Eugenio; Chico, Lucia; Iofrida, Caterina; Petrozzi, Lucia; Baldacci, Filippo; Giacomelli, Chiara; Galetta, Fabio; Siciliano, Gabriele; Bonuccelli, Ubaldo; Trincavelli, Maria L.; Franzoni, Ferdinando; Martini, Claudia

2018-01-01

The loss of protein homeostasis that has been associated with aging leads to altered levels and conformational instability of proteins, which tend to form toxic aggregates. In particular, brain aging presents characteristic patterns of misfolded oligomers, primarily constituted of β-amyloid (Aβ), tau, and α-synuclein (α-syn), which can accumulate in neuronal membranes or extracellular compartments. Such aging-related proteins can also reach peripheral compartments, thus suggesting the possibility to monitor their accumulation in more accessible fluids. In this respect, we have demonstrated that α-syn forms detectable hetero-aggregates with Aβ or tau in red blood cells (RBCs) of healthy subjects. In particular, α-syn levels and its heteromeric interactions are modulated by plasma antioxidant capability (AOC), which increases in turn with physical activity. In order to understand if a specific distribution of misfolded proteins can occur in other blood cells, a cohort of human subjects was enrolled to establish a correlation among AOC, the level of physical exercise and the concentrations of aging-related proteins in platelets. The healthy subjects were divided depending on their level of physical exercise (i.e., athletes and sedentary subjects) and their age (young and older subjects). Herein, aging-related proteins (i.e., α-syn, tau and Aβ) were confirmed to be present in human platelets. Among such proteins, platelet tau concentration was demonstrated to decrease in athletes, while α-syn and Aβ did not correlate with physical exercise. For the first time, α-syn was shown to directly interact with Aβ and tau in platelets, forming detectable hetero-complexes. Interestingly, α-syn interaction with tau was inversely related to plasma AOC and to the level of physical activity. These results suggested that α-syn heterocomplexes, particularly with tau, could represent novel indicators to monitor aging-related proteins in platelets. PMID:29441013

Novel stereocontrolled approach to syn- and anti-oxepene-cyclogeranyl trans-fused polycyclic systems: asymmetric total synthesis of (-)-Aplysistatin, (+)-Palisadin A, (+)-Palisadin B, (+)-12-hydroxy-palisadin B, and the AB ring system of adociasulfate-2 and toxicol A.

PubMed

Couladouros, Elias A; Vidali, Veroniki P

2004-08-06

A new stereocontrolled method for the formation of trans-anti cyclogeranyl-oxepene systems is described. The demanding stereochemistry is secured by stereoselective coupling of a cyclogeranyl tertiary alcohol with a 1,2-unsymmetrically substituted epoxide, while the formation of the highly strained oxepene is achieved employing ring-closing metathesis. Since the stereochemistry of the trans-fused 6,7-ring system is determined by the epoxide, the method also allows the construction of trans-syn 6,7-ring systems. This approach leads to the synthesis of the AB fragment of Adociasulfate-2 and Toxicol A, for the first time. The flexibility and efficiency of the presented strategy is demonstrated by the total asymmetric synthesis of (-)-Aplysistatin, (+)-Palisadin A, (+)-12-hydroxy-Palisadin B, and (+)-Palisadin B, employing two similar key intermediates.

Biophysics of α-synuclein membrane interactions.

PubMed

Pfefferkorn, Candace M; Jiang, Zhiping; Lee, Jennifer C

2012-02-01

Membrane proteins participate in nearly all cellular processes; however, because of experimental limitations, their characterization lags far behind that of soluble proteins. Peripheral membrane proteins are particularly challenging to study because of their inherent propensity to adopt multiple and/or transient conformations in solution and upon membrane association. In this review, we summarize useful biophysical techniques for the study of peripheral membrane proteins and their application in the characterization of the membrane interactions of the natively unfolded and Parkinson's disease (PD) related protein, α-synuclein (α-syn). We give particular focus to studies that have led to the current understanding of membrane-bound α-syn structure and the elucidation of specific membrane properties that affect α-syn-membrane binding. Finally, we discuss biophysical evidence supporting a key role for membranes and α-syn in PD pathogenesis. This article is part of a Special Issue entitled: Membrane protein structure and function. Copyright © 2011. Published by Elsevier B.V.

Food-drug interactions: grapefruit juice.

PubMed

Diaconu, Camelia Harapu; Cuciureanu, Magdalena; Vlase, L; Cuciureanu, Rodica

2011-01-01

Food-drug interactions are increasingly recognized as important clinical events which may change significantly the bioavailability of oral administrated drugs. Grapefruit juice (GFJ) demonstrated multiple interactions with drugs leading to loss of the therapeutic effects or increased side-effects. GFJ decreases pre-systemic metabolism through a) competitive or mechanism-based inhibition of gut wall CYP3A4 isoenzymes and b) P-glycoprotein (P-gp), c) multidrug resistance protein-2 (MRP2) or d) organic anion-transporting polypeptide (OATP) inhibition. Although, GFJ presents high amounts of flavonoids (e.g. naringin, naringenin), furanocoumarins (e.g. 6',7'-dihydroxybergamottin, bergamottin) are the main chemicals involved in the pharmacokinetic interactions. As compounds of GFJ show additive or synergistic effects, all the major furanocoumarins are necessary for the maximal inhibitory effect. Also, related citrus fruits (sweeties, pummelo and sour orange) or various plants containing furanocoumarins may present pharmacological interactions, yet to be discovered.

Design of a single-chain polypeptide tetrahedron assembled from coiled-coil segments.

PubMed

Gradišar, Helena; Božič, Sabina; Doles, Tibor; Vengust, Damjan; Hafner-Bratkovič, Iva; Mertelj, Alenka; Webb, Ben; Šali, Andrej; Klavžar, Sandi; Jerala, Roman

2013-06-01

Protein structures evolved through a complex interplay of cooperative interactions, and it is still very challenging to design new protein folds de novo. Here we present a strategy to design self-assembling polypeptide nanostructured polyhedra based on modularization using orthogonal dimerizing segments. We designed and experimentally demonstrated the formation of the tetrahedron that self-assembles from a single polypeptide chain comprising 12 concatenated coiled coil-forming segments separated by flexible peptide hinges. The path of the polypeptide chain is guided by a defined order of segments that traverse each of the six edges of the tetrahedron exactly twice, forming coiled-coil dimers with their corresponding partners. The coincidence of the polypeptide termini in the same vertex is demonstrated by reconstituting a split fluorescent protein in the polypeptide with the correct tetrahedral topology. Polypeptides with a deleted or scrambled segment order fail to self-assemble correctly. This design platform provides a foundation for constructing new topological polypeptide folds based on the set of orthogonal interacting polypeptide segments.

Stacking interactions between nitrogen-containing six-membered heterocyclic aromatic rings and substituted benzene: studies in solution and in the solid state.

PubMed

Gung, Benjamin W; Wekesa, Francis; Barnes, Charles L

2008-03-07

The stacking interactions between an aromatic ring and a pyridine or a pyrimidine ring are studied by using a series of triptycene-derived scaffolds. The indicative ratios of the syn and anti conformers were determined by variable-temperature NMR spectroscopy. The syn conformer aligns the attached aromatic ring and the heterocycle in a parallel-displaced orientation while the anti conformer sets the two rings apart from each other. Comparing to the corresponding control compounds where a benzene ring is in the position of the heterocycle, higher attractive interactions are observed as indicated by the higher syn/anti ratios. In general, the attractive interactions are much less sensitive to the substituent effects than the corresponding nonheterocycles. The greatest attractive interactions were observed between a pyrimidine ring and a N,N-dimethylaminobenzene, consistent with a predominant donor-acceptor interaction. The interactions between a pyridine ring and a substituted benzene ring show that the pyridine is comparable to that of a NO2- or a CN-substituted benzene ring except for the unpredictable substituent effects.

Effect of acidic pH on the stability of α-synuclein dimers.

PubMed

Lv, Zhengjian; Krasnoslobodtsev, Alexey V; Zhang, Yuliang; Ysselstein, Daniel; Rochet, Jean Christophe; Blanchard, Scott C; Lyubchenko, Yuri L

2016-10-01

Environmental factors, such as acidic pH, facilitate the assembly of α-synuclein (α-Syn) in aggregates, but the impact of pH on the very first step of α-Syn aggregation remains elusive. Recently, we developed a single-molecule approach that enabled us to measure directly the stability of α-Syn dimers. Unlabeled α-Syn monomers were immobilized on a substrate, and fluorophore-labeled monomers were added to the solution to allow them to form dimers with immobilized α-Syn monomers. The dimer lifetimes were measured directly from the fluorescence bursts on the time trajectories. Herein, we applied the single-molecule tethered approach for probing of intermolecular interaction to characterize the effect of acidic pH on the lifetimes of α-Syn dimers. The experiments were performed at pH 5 and 7 for wild-type α-Syn and for two mutants containing familial type mutations E46K and A53T. We demonstrate that a decrease of pH resulted in more than threefold increase in the α-Syn dimers lifetimes with some variability between the α-Syn species. We hypothesize that the stabilization effect is explained by neutralization of residues 96-140 of α-Syn and this electrostatic effect facilitates the association of the two monomers. Given that dimerization is the first step of α-Syn aggregation, we posit that the electrostatic effect thereby contributes to accelerating α-Syn aggregation at acidic pH. © 2016 Wiley Periodicals, Inc. Biopolymers 105: 715-724, 2016. © 2016 Wiley Periodicals, Inc.

Association of vitamin D and vitamin D receptor gene polymorphisms with chronic inflammation, insulin resistance and metabolic syndrome components in type 2 diabetic Egyptian patients.

PubMed

Mackawy, Amal M H; Badawi, Mohammed E H

2014-12-01

To date the published data concerning the possible interplay between vitamin D (VitD) and Vit D receptor (VDR) gene polymorphism with the immune/inflammatory mediators in type 2 diabetes mellitus (DM) is insufficient. Some of the immune non-classical actions of vitamin D may point to its role in the pathogenesis of type 2 DM through down-regulation of cytokines (IL-6). Although there is evidence to support a relationship among vitamin D status, chronic inflammation and insulin resistance, the underlying mechanism requires further exploration. We aimed to investigate the role of vitamin D in chronic inflammation and insulin resistance in type 2 DM. Moreover, to examine the association of VDR gene polymorphisms [VDR 2228570 C > T (FokI); VDR 1544410 A > G (BsmI)] with the components of metabolic syndrome (MetSyn) in type 2 diabetic Egyptian patients . A total of 190 subjects were enrolled in this study, 60 controls and 130 type 2 diabetic patients (Group II). Group II was subdivided into 63 patients without MetSyn (subgroup IIa) and 67 patients with MetSyn (subgroup IIb). Genetic analysis for VDR gene polymorphisms was done in all subjects. VitD and IL-6 plasma levels were estimated. The TT genotype for the VDR FokI was significantly more frequent in subgroup IIb than in subgroup IIa and controls (X (2) = 6.83, P = 0.03 and X (2) = 16.592, P = 0.000) respectively. The T allele was more frequent in the MetSyn group as compared to diabetics without MetSyn (p = 0.001), odds ratio (OR) and 95% CI for the T allele of C > T (FokI) = 2.30 (1.37-3.86). We did not detect any significant difference in VDR BsmI genotypes between patients and control groups (P = 0.947). FokI VDR was significantly associated with the lipid profile parameters, VitD and IL-6 plasma levels in subgroup IIa and associated with HOMA-IR, insulin, VitD, IL-6 levels, waist circumference (WC) and body mass index (BMI) in subgroup IIb while BsmI VDR variant was associated only with VitD values in both subgroups. The present study suggests an interaction between VDR polymorphisms and important components of MetSyn, VitD and pro-inflammatory cytokines (IL-6). FokI VDR polymorphisms may be linked to mild inflammation and insulin resistance and might represent a genetic determinant for developing MetSyn in type 2 diabetic Egyptian patients. The challenge is determining the mechanisms of VitD action for recommendation of VitD supplementation that reduces the risks of MetSyn, insulin resistance and progression to type 2 diabetes.

Association of vitamin D and vitamin D receptor gene polymorphisms with chronic inflammation, insulin resistance and metabolic syndrome components in type 2 diabetic Egyptian patients

PubMed Central

Mackawy, Amal M.H.; Badawi, Mohammed E.H.

2014-01-01

Background To date the published data concerning the possible interplay between vitamin D (VitD) and Vit D receptor (VDR) gene polymorphism with the immune/inflammatory mediators in type 2 diabetes mellitus (DM) is insufficient. Some of the immune non-classical actions of vitamin D may point to its role in the pathogenesis of type 2 DM through down-regulation of cytokines (IL-6). Although there is evidence to support a relationship among vitamin D status, chronic inflammation and insulin resistance, the underlying mechanism requires further exploration. We aimed to investigate the role of vitamin D in chronic inflammation and insulin resistance in type 2 DM. Moreover, to examine the association of VDR gene polymorphisms [VDR 2228570 C > T (FokI); VDR 1544410 A > G (BsmI)] with the components of metabolic syndrome (MetSyn) in type 2 diabetic Egyptian patients . Subjects and methods A total of 190 subjects were enrolled in this study, 60 controls and 130 type 2 diabetic patients (Group II). Group II was subdivided into 63 patients without MetSyn (subgroup IIa) and 67 patients with MetSyn (subgroup IIb). Genetic analysis for VDR gene polymorphisms was done in all subjects. VitD and IL-6 plasma levels were estimated. Results The TT genotype for the VDR FokI was significantly more frequent in subgroup IIb than in subgroup IIa and controls (X2 = 6.83, P = 0.03 and X2 = 16.592, P = 0.000) respectively. The T allele was more frequent in the MetSyn group as compared to diabetics without MetSyn (p = 0.001), odds ratio (OR) and 95% CI for the T allele of C > T (FokI) = 2.30 (1.37–3.86). We did not detect any significant difference in VDR BsmI genotypes between patients and control groups (P = 0.947). FokI VDR was significantly associated with the lipid profile parameters, VitD and IL-6 plasma levels in subgroup IIa and associated with HOMA-IR, insulin, VitD, IL-6 levels, waist circumference (WC) and body mass index (BMI) in subgroup IIb while BsmI VDR variant was associated only with VitD values in both subgroups. Conclusion The present study suggests an interaction between VDR polymorphisms and important components of MetSyn, VitD and pro-inflammatory cytokines (IL-6). FokI VDR polymorphisms may be linked to mild inflammation and insulin resistance and might represent a genetic determinant for developing MetSyn in type 2 diabetic Egyptian patients. The challenge is determining the mechanisms of VitD action for recommendation of VitD supplementation that reduces the risks of MetSyn, insulin resistance and progression to type 2 diabetes. PMID:25606437

Preparation and Characterization of Stable α-Synuclein Lipoprotein Particles.

PubMed

Eichmann, Cédric; Campioni, Silvia; Kowal, Julia; Maslennikov, Innokentiy; Gerez, Juan; Liu, Xiaoxia; Verasdonck, Joeri; Nespovitaya, Nadezhda; Choe, Senyon; Meier, Beat H; Picotti, Paola; Rizo, Josep; Stahlberg, Henning; Riek, Roland

2016-04-15

Multiple neurodegenerative diseases are caused by the aggregation of the human α-Synuclein (α-Syn) protein. α-Syn possesses high structural plasticity and the capability of interacting with membranes. Both features are not only essential for its physiological function but also play a role in the aggregation process. Recently it has been proposed that α-Syn is able to form lipid-protein particles reminiscent of high-density lipoproteins. Here, we present a method to obtain a stable and homogeneous population of nanometer-sized particles composed of α-Syn and anionic phospholipids. These particles are called α-Syn lipoprotein (nano)particles to indicate their relationship to high-density lipoproteins formed by human apolipoproteins in vivo and of in vitro self-assembling phospholipid bilayer nanodiscs. Structural investigations of the α-Syn lipoprotein particles by circular dichroism (CD) and magic angle solid-state nuclear magnetic resonance (MAS SS-NMR) spectroscopy establish that α-Syn adopts a helical secondary structure within these particles. Based on cryo-electron microscopy (cryo-EM) and dynamic light scattering (DLS) α-Syn lipoprotein particles have a defined size with a diameter of ∼23 nm. Chemical cross-linking in combination with solution-state NMR and multiangle static light scattering (MALS) of α-Syn particles reveal a high-order protein-lipid entity composed of ∼8-10 α-Syn molecules. The close resemblance in size between cross-linked in vitro-derived α-Syn lipoprotein particles and a cross-linked species of endogenous α-Syn from SH-SY5Y human neuroblastoma cells indicates a potential functional relevance of α-Syn lipoprotein nanoparticles. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Syn-anti conformational switching in an ethane-bridged Co(II)bisporphyrin induced by external stimuli: effects of inter-macrocyclic interactions, axial ligation and chemical and electrochemical oxidations.

PubMed

Dey, Soumyajit; Rath, Sankar Prasad

2014-02-07

The syn-anti conformational switching has been demonstrated in the ethane-bridged dicobalt(II)bisporphyrin which is present in the syn-form only. The addition of either perylene or axial ligands to Co(II)(bisporphyrin) completely transforms the syn form into the anti because of strong π-π interaction and axial coordination, respectively. The complex undergoes four 1e-oxidations in CH2Cl2 which are indicative of strong through space interactions between the two cofacial Co-porphyrins at 295 K. The first oxidation is a metal centered one and occurs at a potential much lower than that of the monomeric analog. However, the second oxidation, which is again metal centered, was at a significantly higher potential. The large difference between the first two oxidations, as observed here, is due to much stronger inter-porphyrin interactions. The step-wise oxidations have been performed both chemically and electro-chemically while the progress of the reactions was monitored by UV-visible and (1)H NMR spectroscopy. After 1e-oxidation, a very broad (1)H NMR signal results with increased difference between two meso resonances, which indicates that the two macrocycles are in the syn-form with lesser interplanar separation as also observed by DFT. However, 2e-oxidation results in the stabilization of the anti form. The addition of axial ligands to Co(II)(bisporphyrin) also completely transforms the syn form into the anti form. While additions of THF and I2/I(-) both result in the formation of five-coordinate complexes, Co(II) is oxidized to Co(III) in the case of the latter. However, additions of 1-methylimidazole, pyridine and pyrazine as axial ligands result in the formation of a six-coordinate complex in which Co(II) is spontaneously oxidized to Co(III) in air.

Role of α-synuclein in inducing innate and adaptive immunity in Parkinson disease

PubMed Central

Allen Reish, Heather E.; Standaert, David G.

2015-01-01

Alpha-synuclein (α-syn) is central to the pathogenesis of Parkinson disease (PD). Gene duplications, triplications and point mutations in SNCA1, the gene encoding α-syn, cause autosomal dominant forms of PD. Aggregated and post-translationally modified forms of α-syn are present in Lewy bodies and Lewy neurites in both sporadic and familial PD, and recent work has emphasized the prion-like ability of aggregated α-syn to produce spreading pathology. Accumulation of abnormal forms of α-syn is a trigger for PD, but recent evidence suggests that much of the downstream neurodegeneration may result from inflammatory responses. Components of both the innate and adaptive immune systems are activated in PD, and influencing interactions between innate and adaptive immune components has been shown to modify the pathological process in animal models of PD. Understanding the relationship between α-syn and subsequent inflammation may reveal novel targets for neuroprotective interventions. In this review, we examine the role of α-syn and modified forms of this protein in the initiation of innate and adaptive immune responses. PMID:25588354

SYN-PEDS: SYNtactical Pediatric Evaluation and Diagnostic System

PubMed Central

Witten, Matthew; Maloney, David

1980-01-01

SYN-PEDS is a multimodular system which is designed to be an inhome interactive access to a neonatal and pediatric diagnostic information database. This system is designed to assist a parent in assessing his child's condition, as well as in determining whether or not the child needs immediate medical attention. This system is not designed to replace the pediatrician but rather, it is designed as a preventative and health maintenance information system which has the unusually nice side benefit if helping to reduce medical system costs by cutting down on the number of unnecessary visits to private and local clinics as well as private physicians. The current version of SYN-PEDS is composed of of four operative modules: CRITICAL, TREAT, CLINFO, and DIAGNOSE/SYMPTM. These four modules allow the parent/user to interact with the SYN-PEDS system in various modes. As an example, CLINFO is the module which provides clinical information on a variety of subjects. This module is for a parent who wishes information on a particular subject of interest.

SynBioSS-aided design of synthetic biological constructs.

PubMed

Kaznessis, Yiannis N

2011-01-01

We present walkthrough examples of using SynBioSS to design, model, and simulate synthetic gene regulatory networks. SynBioSS stands for Synthetic Biology Software Suite, a platform that is publicly available with Open Licenses at www.synbioss.org. An important aim of computational synthetic biology is the development of a mathematical modeling formalism that is applicable to a wide variety of simple synthetic biological constructs. SynBioSS-based modeling of biomolecular ensembles that interact away from the thermodynamic limit and not necessarily at steady state affords for a theoretical framework that is generally applicable to known synthetic biological systems, such as bistable switches, AND gates, and oscillators. Here, we discuss how SynBioSS creates links between DNA sequences and targeted dynamic phenotypes of these simple systems. Copyright © 2011 Elsevier Inc. All rights reserved.

Dissecting the Molecular Pathway Involved in PLK2 Kinase-mediated α-Synuclein-selective Autophagic Degradation.

PubMed

Dahmene, Manel; Bérard, Morgan; Oueslati, Abid

2017-03-03

Increasing lines of evidence support the causal link between α-synuclein (α-syn) accumulation in the brain and Parkinson's disease (PD) pathogenesis. Therefore, lowering α-syn protein levels may represent a viable therapeutic strategy for the treatment of PD and related disorders. We recently described a novel selective α-syn degradation pathway, catalyzed by the activity of the Polo-like kinase 2 (PLK2), capable of reducing α-syn protein expression and suppressing its toxicity in vivo However, the exact molecular mechanisms underlying this degradation route remain elusive. In the present study we report that among PLK family members, PLK3 is also able to catalyze α-syn phosphorylation and degradation in living cells. Using pharmacological and genetic approaches, we confirmed the implication of the macroautophagy on PLK2-mediated α-syn turnover, and our observations suggest a concomitant co-degradation of these two proteins. Moreover, we showed that the N-terminal region of α-syn is important for PLK2-mediated α-syn phosphorylation and degradation and is implicated in the physical interaction between the two proteins. We also demonstrated that PLK2 polyubiquitination is important for PLK2·α-syn protein complex degradation, and we hypothesize that this post-translational modification may act as a signal for the selective recognition by the macroautophagy machinery. Finally, we observed that the PD-linked mutation E46K enhances PLK2-mediated α-syn degradation, suggesting that this mutated form is a bona fide substrate of this degradation pathway. In conclusion, our study provides a detailed description of the new degradation route of α-syn and offers new opportunities for the development of therapeutic strategies aiming to reduce α-syn protein accumulation and toxicity. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Recombinant α- β- and γ-Synucleins Stimulate Protein Phosphatase 2A Catalytic Subunit Activity in Cell Free Assays

PubMed Central

Lek, Sovanarak; Vargas-Medrano, Javier; Villanueva, Ernesto; Marcus, Brian; Godfrey, Wesley; Perez, Ruth G.

2017-01-01

α-Synuclein (aSyn), β-Synuclein (bSyn), and γ-Synuclein (gSyn) are members of a conserved family of chaperone-like proteins that are highly expressed in vertebrate neuronal tissues. Of the three synucleins, only aSyn has been strongly implicated in neurodegenerative disorders such as Parkinson's disease, Dementia with Lewy Bodies, and Multiple System Atrophy. In studying normal aSyn function, data indicate that aSyn stimulates the activity of the catalytic subunit of an abundantly expressed dephosphorylating enzyme, PP2Ac in vitro and in vivo. Prior data show that aSyn aggregation in human brain reduces PP2Ac activity in regions with Lewy body pathology, where soluble aSyn has become insoluble. However, because all three synucleins have considerable homology in the amino acid sequences, experiments were designed to test if all can modulate PP2Ac activity. Using recombinant synucleins and recombinant PP2Ac protein, activity was assessed by malachite green colorimetric assay. Data revealed that all three recombinant synucleins stimulated PP2Ac activity in cell-free assays, raising the possibility that the conserved homology between synucleins may endow all three homologs with the ability to bind to and activate the PP2Ac. Co-immunoprecipitation data, however, suggest that PP2Ac modulation likely occurs through endogenous interactions between aSyn and PP2Ac in vivo. PMID:28829427

Novel protein-protein interaction between spermidine synthase and S-adenosylmethionine decarboxylase from Leishmania donovani.

PubMed

Mishra, Arjun K; Agnihotri, Pragati; Srivastava, Vijay Kumar; Pratap, J Venkatesh

2015-01-09

Polyamine biosynthesis pathway has long been considered an essential drug target for trypanosomatids including Leishmania. S-adenosylmethionine decarboxylase (AdoMetDc) and spermidine synthase (SpdSyn) are enzymes of this pathway that catalyze successive steps, with the product of the former, decarboxylated S-adenosylmethionine (dcSAM), acting as an aminopropyl donor for the latter enzyme. Here we have explored the possibility of and identified the protein-protein interaction between SpdSyn and AdoMetDc. The protein-protein interaction has been identified using GST pull down assay. Isothermal titration calorimetry reveals that the interaction is thermodynamically favorable. Fluorescence spectroscopy studies also confirms the interaction, with SpdSyn exhibiting a change in tertiary structure with increasing concentrations of AdoMetDc. Size exclusion chromatography suggests the presence of the complex as a hetero-oligomer. Taken together, these results suggest that the enzymes indeed form a heteromer. Computational analyses suggest that this complex differs significantly from the corresponding human complex, implying that this complex could be a better therapeutic target than the individual enzymes. Copyright © 2014 Elsevier Inc. All rights reserved.

Alpha-Synuclein Produces Early Behavioral Alterations via Striatal Cholinergic Synaptic Dysfunction by Interacting With GluN2D N-Methyl-D-Aspartate Receptor Subunit.

PubMed

Tozzi, Alessandro; de Iure, Antonio; Bagetta, Vincenza; Tantucci, Michela; Durante, Valentina; Quiroga-Varela, Ana; Costa, Cinzia; Di Filippo, Massimiliano; Ghiglieri, Veronica; Latagliata, Emanuele Claudio; Wegrzynowicz, Michal; Decressac, Mickael; Giampà, Carmela; Dalley, Jeffrey W; Xia, Jing; Gardoni, Fabrizio; Mellone, Manuela; El-Agnaf, Omar Mukhtar; Ardah, Mustafa Taleb; Puglisi-Allegra, Stefano; Björklund, Anders; Spillantini, Maria Grazia; Picconi, Barbara; Calabresi, Paolo

2016-03-01

Advanced Parkinson's disease (PD) is characterized by massive degeneration of nigral dopaminergic neurons, dramatic motor and cognitive alterations, and presence of nigral Lewy bodies, whose main constituent is α-synuclein (α-syn). However, the synaptic mechanisms underlying behavioral and motor effects induced by early selective overexpression of nigral α-syn are still a matter of debate. We performed behavioral, molecular, and immunohistochemical analyses in two transgenic models of PD, mice transgenic for truncated human α-synuclein 1-120 and rats injected with the adeno-associated viral vector carrying wild-type human α-synuclein. We also investigated striatal synaptic plasticity by electrophysiological recordings from spiny projection neurons and cholinergic interneurons. We found that overexpression of truncated or wild-type human α-syn causes partial reduction of striatal dopamine levels and selectively blocks the induction of long-term potentiation in striatal cholinergic interneurons, producing early memory and motor alterations. These effects were dependent on α-syn modulation of the GluN2D-expressing N-methyl-D-aspartate receptors in cholinergic interneurons. Acute in vitro application of human α-syn oligomers mimicked the synaptic effects observed ex vivo in PD models. We suggest that striatal cholinergic dysfunction, induced by a direct interaction between α-syn and GluN2D-expressing N-methyl-D-aspartate receptors, represents a precocious biological marker of the disease. Copyright © 2016 Society of Biological Psychiatry. Published by Elsevier Inc. All rights reserved.

Molecular Details of α-Synuclein Membrane Association Revealed by Neutrons and Photons

PubMed Central

Jiang, Zhiping; Hess, Sara K.; Heinrich, Frank; Lee, Jennifer C.

2015-01-01

α-Synuclein (α-syn) is an abundant neuronal protein associated with Parkinson’s disease that is disordered in solution, but exists in equilibrium between a bent- and an elongated-helix on negatively charged membranes. Here, neutron reflectometry (NR) and fluorescence spectroscopy were employed to uncover molecular details of the interaction between α-syn and two anionic lipids, phosphatidic acid (PA) and phosphatidylserine (PS). Both NR and site-specific Trp measurements indicate that penetration depth of α-syn is similar for either PA- or PS-containing membranes (~9–11 Å from bilayer center) even though there is a preference for α-syn binding to PA. However, closer examination of the individual Trp quenching profiles by brominated lipids reveal differences into local membrane interactions especially at position 39 where conformational heterogeneity was observed. The data also indicate that while W94 penetrates the bilayer as deeply as W4, W94 resides in a more polar surrounding. Taken together, we suggest the N- and C-terminal regions near positions 4 and 94 are anchored to the membrane, while the putative linker spanning residue 39 samples multiple conformations, which are sensitive to the chemical nature of the membrane surface. This flexibility may enable α-syn to bind diverse biomembranes in vivo. PMID:25790164

Multiple levels of redundant processes inhibit Caenorhabditis elegans vulval cell fates.

PubMed

Andersen, Erik C; Saffer, Adam M; Horvitz, H Robert

2008-08-01

Many mutations cause obvious abnormalities only when combined with other mutations. Such synthetic interactions can be the result of redundant gene functions. In Caenorhabditis elegans, the synthetic multivulva (synMuv) genes have been grouped into multiple classes that redundantly inhibit vulval cell fates. Animals with one or more mutations of the same class undergo wild-type vulval development, whereas animals with mutations of any two classes have a multivulva phenotype. By varying temperature and genetic background, we determined that mutations in most synMuv genes within a single synMuv class enhance each other. However, in a few cases no enhancement was observed. For example, mutations that affect an Mi2 homolog and a histone methyltransferase are of the same class and do not show enhancement. We suggest that such sets of genes function together in vivo and in at least some cases encode proteins that interact physically. The approach of genetic enhancement can be applied more broadly to identify potential protein complexes as well as redundant processes or pathways. Many synMuv genes are evolutionarily conserved, and the genetic relationships we have identified might define the functions not only of synMuv genes in C. elegans but also of their homologs in other organisms.

Multiple Levels of Redundant Processes Inhibit Caenorhabditis elegans Vulval Cell Fates

PubMed Central

Andersen, Erik C.; Saffer, Adam M.; Horvitz, H. Robert

2008-01-01

Many mutations cause obvious abnormalities only when combined with other mutations. Such synthetic interactions can be the result of redundant gene functions. In Caenorhabditis elegans, the synthetic multivulva (synMuv) genes have been grouped into multiple classes that redundantly inhibit vulval cell fates. Animals with one or more mutations of the same class undergo wild-type vulval development, whereas animals with mutations of any two classes have a multivulva phenotype. By varying temperature and genetic background, we determined that mutations in most synMuv genes within a single synMuv class enhance each other. However, in a few cases no enhancement was observed. For example, mutations that affect an Mi2 homolog and a histone methyltransferase are of the same class and do not show enhancement. We suggest that such sets of genes function together in vivo and in at least some cases encode proteins that interact physically. The approach of genetic enhancement can be applied more broadly to identify potential protein complexes as well as redundant processes or pathways. Many synMuv genes are evolutionarily conserved, and the genetic relationships we have identified might define the functions not only of synMuv genes in C. elegans but also of their homologs in other organisms. PMID:18689876

Mechanisms of α-Synuclein Induced Synaptopathy in Parkinson's Disease

PubMed Central

Bridi, Jessika C.; Hirth, Frank

2018-01-01

Parkinson's disease (PD) is characterized by intracellular inclusions of aggregated and misfolded α-Synuclein (α-Syn), and the loss of dopaminergic (DA) neurons in the brain. The resulting motor abnormalities mark the progression of PD, while non-motor symptoms can already be identified during early, prodromal stages of disease. Recent studies provide evidence that during this early prodromal phase, synaptic and axonal abnormalities occur before the degenerative loss of neuronal cell bodies. These early phenotypes can be attributed to synaptic accumulation of toxic α-Syn. Under physiological conditions, α-Syn functions in its native conformation as a soluble monomer. However, PD patient brains are characterized by intracellular inclusions of insoluble fibrils. Yet, oligomers and protofibrils of α-Syn have been identified to be the most toxic species, with their accumulation at presynaptic terminals affecting several steps of neurotransmitter release. First, high levels of α-Syn alter the size of synaptic vesicle pools and impair their trafficking. Second, α-Syn overexpression can either misregulate or redistribute proteins of the presynaptic SNARE complex. This leads to deficient tethering, docking, priming and fusion of synaptic vesicles at the active zone (AZ). Third, α-Syn inclusions are found within the presynaptic AZ, accompanied by a decrease in AZ protein levels. Furthermore, α-Syn overexpression reduces the endocytic retrieval of synaptic vesicle membranes during vesicle recycling. These presynaptic alterations mediated by accumulation of α-Syn, together impair neurotransmitter exocytosis and neuronal communication. Although α-Syn is expressed throughout the brain and enriched at presynaptic terminals, DA neurons are the most vulnerable in PD, likely because α-Syn directly regulates dopamine levels. Indeed, evidence suggests that α-Syn is a negative modulator of dopamine by inhibiting enzymes responsible for its synthesis. In addition, α-Syn is able to interact with and reduce the activity of VMAT2 and DAT. The resulting dysregulation of dopamine levels directly contributes to the formation of toxic α-Syn oligomers. Together these data suggest a vicious cycle of accumulating α-Syn and deregulated dopamine that triggers synaptic dysfunction and impaired neuronal communication, ultimately causing synaptopathy and progressive neurodegeneration in Parkinson's disease. PMID:29515354

Epothilone D inhibits microglia-mediated spread of alpha-synuclein aggregates.

PubMed

Valdinocci, Dario; Grant, Gary D; Dickson, Tracey C; Pountney, Dean L

2018-04-16

Multiple System Atrophy (MSA) is a progressive neurodegenerative disease characterized by chronic neuroinflammation and widespread α-synuclein (α-syn) cytoplasmic inclusions. Neuroinflammation associated with microglial cells is typically located in brain regions with α-syn deposits. The potential link between microglial cell migration and the transport of pathological α-syn protein in MSA was investigated. Qualitative analysis via immunofluorescence of MSA cases (n = 4) revealed microglial cells bearing α-syn inclusions distal from oligodendrocytes bearing α-syn cytoplasmic inclusions, as well as close interactions between microglia and oligodendrocytes bearing α-syn, suggestive of a potential transfer mechanism between microglia and α-syn bearing cells in MSA and the possibility of microglia acting as a mobile vehicle to spread α-syn between anatomically connected brain regions. Further In vitro experiments using microglial-like differentiated THP-1 cells were conducted to investigate if microglial cells could act as potential transporters of α-syn. Monomeric or aggregated α-syn was immobilized at the centre of glass coverslips and treated with either cell free medium, undifferentiated THP-1 cells or microglial-like phorbol-12-myristate-13-acetate differentiated THP-1 cells (48 h; n = 3). A significant difference in residual immobilized α-syn density was observed between cell free controls and differentiated (p = 0.016) as well as undifferentiated and differentiated THP-1 cells (p = 0.032) when analysed by quantitative immunofluorescence. Furthermore, a significantly greater proportion of differentiated cells were observed bearing α-syn aggregates distal from the immobilized protein than their non-differentiated counterparts (p = 0.025). Similar results were observed with Highly Aggressive Proliferating Immortalised (HAPI) microglial cells, with cells exposed to aggregated α-syn yielding lower residual immobilized α-syn (p = 0.004) and a higher proportion of α-syn positive distal cells (p = 0.001) than cells exposed to monomeric α-syn. Co-treatment of THP-1 groups with the tubulin depolymerisation inhibitor, Epothilone D (EpoD; 10 nM), was conducted to investigate if inhibition of microtubule activity had an effect on cell migration and residual immobilized α-syn density. There was a significant increase in both residual immobilized α-syn between EpoD treated and non-treated differentiated cells exposed to monomeric (p = 0.037) and aggregated (p = 0.018) α-syn, but not with undifferentiated cells. Differentiated THP-1 cells exposed to immobilized aggregated α-syn showed a significant difference in the proportion of distal aggregate bearing cells between EpoD treated and untreated (p = 0.027). The results suggest microglia could play a role in α-syn transport in MSA, a role which could potentially be inhibited therapeutically by EpoD. Copyright © 2018 Elsevier Inc. All rights reserved.

High-speed atomic force microscopy reveals structural dynamics of α -synuclein monomers and dimers

NASA Astrophysics Data System (ADS)

Zhang, Yuliang; Hashemi, Mohtadin; Lv, Zhengjian; Williams, Benfeard; Popov, Konstantin I.; Dokholyan, Nikolay V.; Lyubchenko, Yuri L.

2018-03-01

α-Synuclein (α-syn) is the major component of the intraneuronal inclusions called Lewy bodies, which are the pathological hallmark of Parkinson's disease. α-Syn is capable of self-assembly into many different species, such as soluble oligomers and fibrils. Even though attempts to resolve the structures of the protein have been made, detailed understanding about the structures and their relationship with the different aggregation steps is lacking, which is of interest to provide insights into the pathogenic mechanism of Parkinson's disease. Here we report the structural flexibility of α-syn monomers and dimers in an aqueous solution environment as probed by single-molecule time-lapse high-speed AFM. In addition, we present the molecular basis for the structural transitions using discrete molecular dynamics (DMD) simulations. α-Syn monomers assume a globular conformation, which is capable of forming tail-like protrusions over dozens of seconds. Importantly, a globular monomer can adopt fully extended conformations. Dimers, on the other hand, are less dynamic and show a dumbbell conformation that experiences morphological changes over time. DMD simulations revealed that the α-syn monomer consists of several tightly packed small helices. The tail-like protrusions are also helical with a small β-sheet, acting as a "hinge". Monomers within dimers have a large interfacial interaction area and are stabilized by interactions in the non-amyloid central (NAC) regions. Furthermore, the dimer NAC-region of each α-syn monomer forms a β-rich segment. Moreover, NAC-regions are located in the hydrophobic core of the dimer.

Suppression of Murine Retrovirus Polypeptide Termination: Effect of Amber Suppressor tRNA on the Cell-Free Translation of Rauscher Murine Leukemia Virus, Moloney Murine Leukemia Virus, and Moloney Murine Sarcoma Virus 124 RNA

PubMed Central

Murphy, Edwin C.; Wills, Norma; Arlinghaus, Ralph B.

1980-01-01

The effect of suppressor tRNA's on the cell-free translation of several leukemia and sarcoma virus RNAs was examined. Yeast amber suppressor tRNA (amber tRNA) enhanced the synthesis of the Rauscher murine leukemia virus and clone 1 Moloney murine leukemia virus Pr200gag-pol polypeptides by 10- to 45-fold, but at the same time depressed the synthesis of Rauscher murine leukemia virus Pr65gag and Moloney murine leukemia virus Pr63gag. Under suppressor-minus conditions, Moloney murine leukemia virus Pr70gag was present as a closely spaced doublet. Amber tRNA stimulated the synthesis of the “upper” Moloney murine leukemia virus Pr70gag polypeptide. Yeast ochre suppressor tRNA appeared to be ineffective. Quantitative analyses of the kinetics of viral precursor polypeptide accumulation in the presence of amber tRNA showed that during linear protein synthesis, the increase in accumulated Moloney murine leukemia virus Pr200gag-pol coincided closely with the molar loss of Pr63gag. Enhancement of Pr200gag-pol and Pr70gag by amber tRNA persisted in the presence of pactamycin, a drug which blocks the initiation of protein synthesis, thus arguing for the addition of amino acids to the C terminus of Pr63gag as the mechanism behind the amber tRNA effect. Moloney murine sarcoma virus 124 30S RNA was translated into four major polypeptides, Pr63gag, P42, P38, and P23. In the presence of amber tRNA, a new polypeptide, Pr67gag, appeared, whereas Pr63gag synthesis was decreased. Quantitative estimates indicated that for every 1 mol of Pr67gag which appeared, 1 mol of Pr63gag was lost. Images PMID:7373716

Structural development of the onshore Otway passive margin (Australia): the interaction of rotating syn-sedimentary faults

NASA Astrophysics Data System (ADS)

Tanner, David C.; Ziesch, Jennifer; Krawczyk, Charlotte M.

2017-04-01

Within the context of long-term CO2 storage integrity, we interpreted the faults within the 2.2 km thick, syn-rift, Late Cretaceous to Recent sediments below the CO2CRC Otway Project site in Australia using a detailed interpretation of a 3-D reflection seismic cube (32.3 km×14.35 km × 4100 ms TWT). All the faults in the onshore Otway passive margin basin in this area were active to varying degrees during sedimentation, between ca. 120 and 50 Ma, before they died out. From analysis of fault juxtaposition and fault tip-line propagation maps, as well as analysis of individual stratigraphic thickness maps, we determine the direction and incremental amount of syn-sedimentary movement on each fault. Thickening of the hanging-walls of the faults occurred, as is typical for syn-sedimentary faults. However, we also determine that substantial local footwall thinning took place. Although the syn-sedimentary behaviour of the faults was constantly maintained until 50 Ma, there were two main phases of footwall thinning, separated by a quiescent phase. We postulate that these phases of footwall thinning represent rotation of the fault blocks that correlate with prograding sediment pulses within the passive margin. The rotation of the fault blocks occurred simultaneously, i.e., they could only rotate if they interacted.

Identification of bovine sperm acrosomal proteins that interact with a 32-kDa acrosomal matrix protein.

PubMed

Nagdas, Subir K; Smith, Linda; Medina-Ortiz, Ilza; Hernandez-Encarnacion, Luisa; Raychoudhury, Samir

2016-03-01

Mammalian fertilization is accomplished by the interaction between sperm and egg. Previous studies from this laboratory have identified a stable acrosomal matrix assembly from the bovine sperm acrosome termed the outer acrosomal membrane-matrix complex (OMC). This stable matrix assembly exhibits precise binding activity for acrosin and N-acetylglucosaminidase. A highly purified OMC fraction comprises three major (54, 50, and 45 kDa) and several minor (38-19 kDa) polypeptides. The set of minor polypeptides (38-19 kDa) termed "OMCrpf polypeptides" is selectively solubilized by high-pH extraction (pH 10.5), while the three major polypeptides (55, 50, and 45 kDa) remain insoluble. Proteomic identification of the OMC32 polypeptide (32 kDa polypeptide isolated from high-pH soluble fraction of OMC) yielded two peptides that matched the NCBI database sequence of acrosin-binding protein. Anti-OMC32 recognized an antigenically related family of polypeptides (OMCrpf polypeptides) in the 38-19-kDa range with isoelectric points ranging between 4.0 and 5.1. Other than glycohydrolases, OMC32 may also be complexed to other acrosomal proteins. The present study was undertaken to identify and localize the OMC32 binding polypeptides and to elucidate the potential role of the acrosomal protein complex in sperm function. OMC32 affinity chromatography of a detergent-soluble fraction of bovine cauda sperm acrosome followed by mass spectrometry-based identification of bound proteins identified acrosin, lactadherin, SPACA3, and IZUMO1. Co-immunoprecipitation analysis also demonstrated the interaction of OMC32 with acrosin, lactadherin, SPACA3, and IZUMO1. Our immunofluorescence studies revealed the presence of SPACA3 and lactadherin over the apical segment, whereas IZUMO1 is localized over the equatorial segment of Triton X-100 permeabilized cauda sperm. Immunoblot analysis showed that a significant portion of SPACA3 was released after the lysophosphatidylcholine (LPC)-induced acrosome reaction, whereas the IZUMO1 and lactadherin polypeptides remain associated to the particulate fraction. Almost entire population of bovine sperm IZUMO1 relocates to the equatorial segment during the LPC-induced acrosome reaction. We propose that the interaction of OMC32 matrix polypeptide with detergent-soluble acrosomal proteins regulates the release of hydrolases/other acrosomal protein(s) during the acrosome reaction.

α-Synuclein Heterocomplexes with β-Amyloid Are Increased in Red Blood Cells of Parkinson’s Disease Patients and Correlate with Disease Severity

PubMed Central

Daniele, Simona; Frosini, Daniela; Pietrobono, Deborah; Petrozzi, Lucia; Lo Gerfo, Annalisa; Baldacci, Filippo; Fusi, Jonathan; Giacomelli, Chiara; Siciliano, Gabriele; Trincavelli, Maria Letizia; Franzoni, Ferdinando; Ceravolo, Roberto; Martini, Claudia; Bonuccelli, Ubaldo

2018-01-01

Neurodegenerative disorders (NDs) are characterized by abnormal accumulation/misfolding of specific proteins, primarily α-synuclein (α-syn), β-amyloid1–42 (Aβ1–42) and tau, in both brain and peripheral tissues. In addition to oligomers, the role of the interactions of α-syn with Aβ or tau has gradually emerged. Nevertheless, despite intensive research, NDs have no accepted peripheral markers for biochemical diagnosis. In this respect, Red Blood Cells (RBCs) are emerging as a valid peripheral model for the study of aging-related pathologies. Herein, a small cohort (N = 28) of patients affected by Parkinson’s disease (PD) and age-matched controls were enrolled to detect the content of α-syn (total and oligomeric), Aβ1–42 and tau (total and phosphorylated) in RBCs. Moreover, the presence of α-syn association with tau and Aβ1–42 was explored by co-immunoprecipitation/western blotting in the same cells, and quantitatively confirmed by immunoenzymatic assays. For the first time, PD patients were demonstrated to exhibit α-syn heterocomplexes with Aβ1–42 and tau in peripheral tissues; interestingly, α-syn-Aβ1–42 concentrations were increased in PD subjects with respect to healthy controls (HC), and directly correlated with disease severity and motor deficits. Moreover, total-α-syn levels were decreased in PD subjects and inversely related to their motor deficits. Finally, an increase of oligomeric-α-syn and phosphorylated-tau was observed in RBCs of the enrolled patients. The combination of three parameters (total-α-syn, phosphorylated-tau and α-syn-Aβ1–42 concentrations) provided the best fitting predictive index for discriminating PD patients from controls. Nevertheless further investigations should be required, overall, these data suggest α-syn hetero-aggregates in RBCs as a putative tool for the diagnosis of PD. PMID:29520218

From the Macro to the Micro: Gel Mapping to Differentiate between Sporozoites of Two Immunologically Distinct Strains of Eimeria maxima (Strains M6 and Guelph)

PubMed Central

Liu, Hongbin; Al Nasr, Ibrahim; Liu, Xianyong; Suo, Xun; Barta, John

2015-01-01

Two immunologically distinct strains of E. maxima were examined in this study: the M6 strain and the Guelph strain. The differential expression between the sporozoites of the two strains of E. maxima was determined by image analysis of 100 μg of protein from each strain separated by standard one- and conventional two-dimensional polyacrylamide gel electrophoresis. In addition to differences in both molecular weight and the electrophoretic mobility, differences in the intensity of polypeptide bands for example, GS 136.4 and M6 169 were explored. Pooled gels were prepared from each strain. A representative 2D-PAGE gel spanning a non-linear pH range of 3–10 of E. maxima strain M6 consisted of approximately 694 polypeptide spots with about 67 (9.6%) of the polypeptide spots being unique relative to the other strain. E. maxima strain GS had about 696 discernable polypeptide spots with 69 spots (9.9%) that differed from those of the M6 strain. In-depth characterization of the variable polypeptide spots; unique polypeptide spots (absence or presence) and shared polypeptide spots with modifications may lead to novel vaccine target in the form of multi-component, multi-stage, multi-immunovariant strains, multi-species subunit vaccine, and diagnostic probe for E. maxima. PMID:26641262

From the Macro to the Micro: Gel Mapping to Differentiate between Sporozoites of Two Immunologically Distinct Strains of Eimeria maxima (Strains M6 and Guelph).

PubMed

El-Ashram, Saeed; Yin, Qing; Liu, Hongbin; Al Nasr, Ibrahim; Liu, Xianyong; Suo, Xun; Barta, John

2015-01-01

Two immunologically distinct strains of E. maxima were examined in this study: the M6 strain and the Guelph strain. The differential expression between the sporozoites of the two strains of E. maxima was determined by image analysis of 100 μg of protein from each strain separated by standard one- and conventional two-dimensional polyacrylamide gel electrophoresis. In addition to differences in both molecular weight and the electrophoretic mobility, differences in the intensity of polypeptide bands for example, GS 136.4 and M6 169 were explored. Pooled gels were prepared from each strain. A representative 2D-PAGE gel spanning a non-linear pH range of 3-10 of E. maxima strain M6 consisted of approximately 694 polypeptide spots with about 67 (9.6%) of the polypeptide spots being unique relative to the other strain. E. maxima strain GS had about 696 discernable polypeptide spots with 69 spots (9.9%) that differed from those of the M6 strain. In-depth characterization of the variable polypeptide spots; unique polypeptide spots (absence or presence) and shared polypeptide spots with modifications may lead to novel vaccine target in the form of multi-component, multi-stage, multi-immunovariant strains, multi-species subunit vaccine, and diagnostic probe for E. maxima.

Crystal structure and magnetic properties of two isomeric three-dimensional pyromellitate-containing cobalt(II) complexes.

PubMed

Fabelo, Oscar; Pasán, Jorge; Cañadillas-Delgado, Laura; Delgado, Fernando S; Lloret, Francesc; Julve, Miguel; Ruiz-Pérez, Catalina

2008-09-15

The hydrothermal preparation, crystal structure determination, and magnetic study of two isomers made up of 1,2,4,5-benzenetetracarboxylate and high-spin Co(II) ions of formula [Co2(bta)(H2O)4]n x 2n H2O (1 and 2; H4bta = 1,2,4,5-benzenetetracarboxylic acid) are reported. 1 and 2 are three-dimensional compounds whose structures can be described as (4,4) rectangular layers of trans-diaquacobalt(II) units with the bta(4-) anion acting as tetrakis-monodentate ligand through the four carboxylate groups, which are further connected through other trans-[Co(H2O)2](2+) (1) and planar [Co(H2O)4](2+) (2) entities, with the bridging units being a carboxylate group in either the anti-syn (1) or syn-syn (2) conformations and a water molecule (2). The study of the magnetic properties of 1 and 2 in the temperature range 1.9-300 K shows the occurrence of weak antiferromagnetic interactions between the high-spin Co(II) ions, with the strong decrease of chi(M)T upon cooling being mainly due to the depopulation of the higher energy Kramers doublets of the six-coordinated Co(II) ions. The computed values of the exchange coupling between the Co(II) ions across anti-syn carboxylate (1) and syn-syn carboxylate/water (2) bridges are J = -0.060 (1) and -1.90 (2) cm(-1) (with the Hamiltonian being defined as H = -Jsigma(i,j)S(i) x S(j)). These values follow the different conformations of the carboxylate bridge in 1 (anti-syn) and 2 (syn-syn) with the occurrence of a double bridge in 2 (water/carboxylate).

Purification of α-Synuclein from Human Brain Reveals an Instability of Endogenous Multimers as the Protein Approaches Purity

PubMed Central

2015-01-01

Despite two decades of research, the structure–function relationships of endogenous, physiological forms of α-synuclein (αSyn) are not well understood. Most in vitro studies of this Parkinson’s disease-related protein have focused on recombinant αSyn that is unfolded and monomeric, assuming that this represents its state in the normal human brain. Recently, we have provided evidence that αSyn exists in considerable part in neurons, erythrocytes, and other cells as a metastable multimer that principally sizes as a tetramer. In contrast to recombinant αSyn, physiological tetramers purified from human erythrocytes have substantial α-helical content and resist pathological aggregation into β-sheet rich fibers. Here, we report the first method to fully purify soluble αSyn from the most relevant source, human brain. We describe protocols that purify αSyn to homogeneity from nondiseased human cortex using ammonium sulfate precipitation, gel filtration, and ion exchange, hydrophobic interaction, and affinity chromatographies. Cross-linking of the starting material and the partially purified chromatographic fractions revealed abundant αSyn multimers, including apparent tetramers, but these were destabilized in large part to monomers during the final purification step. The method also fully purified the homologue β-synuclein, with a similar outcome. Circular dichroism spectroscopy showed that purified, brain-derived αSyn can display more helical content than the recombinant protein, but this result varied. Collectively, our data suggest that purifying αSyn to homogeneity destabilizes native, α-helix-rich multimers that exist in intact and partially purified brain samples. This finding suggests existence of a stabilizing cofactor (e.g., a small lipid) present inside neurons that is lost during final purification. PMID:25490121

Excimer-based peptide beacons: a convenient experimental approach for monitoring polypeptide-protein and polypeptide-oligonucleotide interactions.

PubMed

Oh, Kenneth J; Cash, Kevin J; Plaxco, Kevin W

2006-11-01

While protein-polypeptide and nucleic acid-polypeptide interactions are of significant experimental interest, quantitative methods for the characterization of such interactions are often cumbersome. Here we described a relatively simple means of optically monitoring such interactions using excimer-based peptide beacons (PBs). The design of PBs is based on the observation that, whereas short peptides are almost invariably unfolded and highly dynamic, they become rigid when complexed with macromolecular targets. Using this binding-induced folding to segregate two pyrene moieties and therefore inhibit excimer formation, we have produced PBs directed against both anti-HIV antibodies and the retroviral transactive response (TAR) RNA hairpin. For both polypeptides, target recognition is accompanied by a roughly 2-fold decrease in excimer emission, thus allowing the detection of their respective targets at concentrations of a few nanomolar. Because excimer emission requires the formation of a tight, precisely oriented pyrene dimer, even relatively trivial binding-induced segregation reduces fluorescence significantly. This suggests that the PB approach will be suitable for monitoring a wide range of peptide-macromolecule recognition events. Moreover, the synthesis of excimer-based PBs utilizes commercially available modified pyrenes in a simple and well-established protocol, making the approach well suited for routine laboratory applications.

Peppytides: Interactive Models of Polypeptide Chains

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zuckermann, Ron; Chakraborty, Promita; Derisi, Joe

2014-01-21

Peppytides are scaled, 3D-printed models of polypeptide chains that can be folded into accurate protein structures. Designed and created by Berkeley Lab Researcher, Promita Chakraborty, and Berkeley Lab Senior Scientist, Dr. Ron Zuckermann, Peppytides are accurate physical models of polypeptide chains that anyone can interact with and fold intro various protein structures - proving to be a great educational tool, resulting in a deeper understanding of these fascinating structures and how they function. Build your own Peppytide model and learn about how nature's machines fold into their intricate architectures!

Peppytides: Interactive Models of Polypeptide Chains

ScienceCinema

Zuckermann, Ron; Chakraborty, Promita; Derisi, Joe

2018-06-08

Peppytides are scaled, 3D-printed models of polypeptide chains that can be folded into accurate protein structures. Designed and created by Berkeley Lab Researcher, Promita Chakraborty, and Berkeley Lab Senior Scientist, Dr. Ron Zuckermann, Peppytides are accurate physical models of polypeptide chains that anyone can interact with and fold intro various protein structures - proving to be a great educational tool, resulting in a deeper understanding of these fascinating structures and how they function. Build your own Peppytide model and learn about how nature's machines fold into their intricate architectures!

c-Abl phosphorylates α-synuclein and regulates its degradation: implication for α-synuclein clearance and contribution to the pathogenesis of Parkinson's disease

PubMed Central

Mahul-Mellier, Anne-Laure; Fauvet, Bruno; Gysbers, Amanda; Dikiy, Igor; Oueslati, Abid; Georgeon, Sandrine; Lamontanara, Allan J.; Bisquertt, Alejandro; Eliezer, David; Masliah, Eliezer; Halliday, Glenda; Hantschel, Oliver; Lashuel, Hilal A.

2014-01-01

Increasing evidence suggests that the c-Abl protein tyrosine kinase could play a role in the pathogenesis of Parkinson's disease (PD) and other neurodegenerative disorders. c-Abl has been shown to regulate the degradation of two proteins implicated in the pathogenesis of PD, parkin and α-synuclein (α-syn). The inhibition of parkin's neuroprotective functions is regulated by c-Abl-mediated phosphorylation of parkin. However, the molecular mechanisms by which c-Abl activity regulates α-syn toxicity and clearance remain unknown. Herein, using NMR spectroscopy, mass spectrometry, in vitro enzymatic assays and cell-based studies, we established that α-syn is a bona fide substrate for c-Abl. In vitro studies demonstrate that c-Abl directly interacts with α-syn and catalyzes its phosphorylation mainly at tyrosine 39 (pY39) and to a lesser extent at tyrosine 125 (pY125). Analysis of human brain tissues showed that pY39 α-syn is detected in the brains of healthy individuals and those with PD. However, only c-Abl protein levels were found to be upregulated in PD brains. Interestingly, nilotinib, a specific inhibitor of c-Abl kinase activity, induces α-syn protein degradation via the autophagy and proteasome pathways, whereas the overexpression of α-syn in the rat midbrains enhances c-Abl expression. Together, these data suggest that changes in c-Abl expression, activation and/or c-Abl-mediated phosphorylation of Y39 play a role in regulating α-syn clearance and contribute to the pathogenesis of PD. PMID:24412932

Hippocampal neuronal cells that accumulate α-synuclein fragments are more vulnerable to Aβ oligomer toxicity via mGluR5 – implications for dementia with Lewy bodies

PubMed Central

2014-01-01

Background In dementia with Lewy bodies (DLB) abnormal interactions between α-synuclein (α-syn) and beta amyloid (Aβ) result in selective degeneration of neurons in the neocortex, limbic system and striatum. However, factors rendering these neurons selectively vulnerable have not been fully investigated. The metabotropic glutamate receptor 5 (mGluR5) has been shown to be up regulated in DLB and might play a role as a mediator of the neurotoxic effects of Aβ and α-syn in vulnerable neuronal populations. In this context, the main objective of the present study was to investigate the role of mGluR5 as a mediator of the neurotoxic effects of α-syn and Aβ in the hippocampus. Results We generated double transgenic mice over-expressing amyloid precursor protein (APP) and α-syn under the mThy1 cassette and investigated the relationship between α-syn cleavage, Aβ, mGluR5 and neurodegeneration in the hippocampus. We found that compared to the single tg mice, the α-syn/APP tg mice displayed greater accumulation of α-syn and mGluR5 in the CA3 region of the hippocampus compared to the CA1 and other regions. This was accompanied by loss of CA3 (but not CA1) neurons in the single and α-syn/APP tg mice and greater loss of MAP 2 and synaptophysin in the CA3 in the α-syn/APP tg. mGluR5 gene transfer using a lentiviral vector into the hippocampus CA1 region resulted in greater α-syn accumulation and neurodegeneration in the single and α-syn/APP tg mice. In contrast, silencing mGluR5 with a lenti-shRNA protected neurons in the CA3 region of tg mice. In vitro, greater toxicity was observed in primary hippocampal neuronal cultures treated with Aβ oligomers and over-expressing α-syn; this effect was attenuated by down-regulating mGluR5 with an shRNA lentiviral vector. In α-syn-expressing neuronal cells lines, Aβ oligomers promoted increased intracellular calcium levels, calpain activation and α-syn cleavage resulting in caspase-3-dependent cell death. Treatment with pharmacological mGluR5 inhibitors such as 2-Methyl-6-(phenylethynyl)pyridine (MPEP) and 3-((2-Methyl-4-thiazolyl)ethynyl)pyridine (MTEP) attenuated the toxic effects of Aβ in α-syn-expressing neuronal cells. Conclusions Together, these results support the possibility that vulnerability of hippocampal neurons to α-syn and Aβ might be mediated via mGluR5. Moreover, therapeutical interventions targeting mGluR5 might have a role in DLB. PMID:24885390

Two novel Mesocestoides vogae fatty acid binding proteins--functional and evolutionary implications.

PubMed

Alvite, Gabriela; Canclini, Lucía; Corvo, Ileana; Esteves, Adriana

2008-01-01

This work describes two new fatty acid binding proteins (FABPs) identified in the parasite platyhelminth Mesocestoides vogae (syn. corti). The corresponding polypeptide chains share 62% identical residues and overall 90% similarity according to CLUSTALX default conditions. Compared with Cestoda FABPs, these proteins share the highest similarity score with the Taenia solium protein. M. vogae FABPs are also phylogenetically related to the FABP3/FABP4 mammalian FABP subfamilies. The native proteins were purified by chromatographical procedures, and apparent molecular mass and isoelectric point were determined. Immunolocalization studies determined the localization of the expression of these proteins in the larval form of the parasite. The genomic exon-intron organization of both genes is also reported, and supports new insights on intron evolution. Consensus motifs involved in splicing were identified.

α-Synuclein in Central Nervous System and from Erythrocytes, Mammalian Cells, and Escherichia coli Exists Predominantly as Disordered Monomer*

PubMed Central

Fauvet, Bruno; Mbefo, Martial K.; Fares, Mohamed-Bilal; Desobry, Carole; Michael, Sarah; Ardah, Mustafa T.; Tsika, Elpida; Coune, Philippe; Prudent, Michel; Lion, Niels; Eliezer, David; Moore, Darren J.; Schneider, Bernard; Aebischer, Patrick; El-Agnaf, Omar M.; Masliah, Eliezer; Lashuel, Hilal A.

2012-01-01

Since the discovery and isolation of α-synuclein (α-syn) from human brains, it has been widely accepted that it exists as an intrinsically disordered monomeric protein. Two recent studies suggested that α-syn produced in Escherichia coli or isolated from mammalian cells and red blood cells exists predominantly as a tetramer that is rich in α-helical structure (Bartels, T., Choi, J. G., and Selkoe, D. J. (2011) Nature 477, 107–110; Wang, W., Perovic, I., Chittuluru, J., Kaganovich, A., Nguyen, L. T. T., Liao, J., Auclair, J. R., Johnson, D., Landeru, A., Simorellis, A. K., Ju, S., Cookson, M. R., Asturias, F. J., Agar, J. N., Webb, B. N., Kang, C., Ringe, D., Petsko, G. A., Pochapsky, T. C., and Hoang, Q. Q. (2011) Proc. Natl. Acad. Sci. 108, 17797–17802). However, it remains unknown whether or not this putative tetramer is the main physiological form of α-syn in the brain. In this study, we investigated the oligomeric state of α-syn in mouse, rat, and human brains. To assess the conformational and oligomeric state of native α-syn in complex mixtures, we generated α-syn standards of known quaternary structure and conformational properties and compared the behavior of endogenously expressed α-syn to these standards using native and denaturing gel electrophoresis techniques, size-exclusion chromatography, and an oligomer-specific ELISA. Our findings demonstrate that both human and rodent α-syn expressed in the central nervous system exist predominantly as an unfolded monomer. Similar results were observed when human α-syn was expressed in mouse and rat brains as well as mammalian cell lines (HEK293, HeLa, and SH-SY5Y). Furthermore, we show that α-syn expressed in E. coli and purified under denaturing or nondenaturing conditions, whether as a free protein or as a fusion construct with GST, is monomeric and adopts a disordered conformation after GST removal. These results do not rule out the possibility that α-syn becomes structured upon interaction with other proteins and/or biological membranes. PMID:22315227

Elucidating the Role of Site-Specific Nitration of α-Synuclein in the Pathogenesis of Parkinson's Disease via Protein Semisynthesis and Mutagenesis.

PubMed

Burai, Ritwik; Ait-Bouziad, Nadine; Chiki, Anass; Lashuel, Hilal A

2015-04-22

Parkinson's disease (PD) is characterized by the loss of dopaminergic neurons in the substantia nigra and the presence of intraneuronal inclusions consisting of aggregated and post-translationally modified α-synuclein (α-syn). Despite advances in the chemical synthesis of α-syn and other proteins, the generation of site-specifically nitrated synthetic proteins has not been reported. Consequently, it has not been possible to determine the roles of nitration at specific residues in regulating the physiological and pathogenic properties of α-syn. Here we report, for the first time, the site-specific incorporation of 3-nitrotyrosine at different regions of α-syn using native chemical ligation combined with a novel desulfurization strategy. This strategy enabled us to investigate the role of nitration at single or multiple tyrosine residues in regulating α-syn structure, membrane binding, oligomerization, and fibrils formation. We demonstrate that different site-specifically nitrated α-syn species exhibit distinct structural and aggregation properties and exhibit reduced affinity to negatively charged vesicle membranes. We provide evidence that intermolecular interactions between the N- and C-terminal regions of α-syn play critical roles in mediating nitration-induced α-syn oligomerization. For example, when Y39 is not available for nitration (Y39F and Y39/125F), the extent of cross-linking is limited mostly to dimer formation, whereas mutants in which Y39 along with one or multiple C-terminal tyrosines (Y125F, Y133F, Y136F and Y133/136F) can still undergo nitration readily to form higher-order oligomers. Our semisynthetic strategy for generating site-specifically nitrated proteins opens up new possibilities for investigating the role of nitration in regulating protein structure and function in health and disease.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mishra, Arjun K.; Agnihotri, Pragati; Srivastava, Vijay Kumar

Highlights: • L. donovani spermidine synthase and S-adenosylmethionine decarboxylase have been cloned and purified. • S-adenosylmethionine decarboxylase has autocatalytic property. • GST pull down assay shows the two proteins to form a metabolon. • Isothermal titration calorimetry shows that binding was exothermic having K{sub d} value of 0.4 μM. • Interaction confirmed by fluorescence spectroscopy and size exclusion chromatography. - Abstract: Polyamine biosynthesis pathway has long been considered an essential drug target for trypanosomatids including Leishmania. S-adenosylmethionine decarboxylase (AdoMetDc) and spermidine synthase (SpdSyn) are enzymes of this pathway that catalyze successive steps, with the product of the former, decarboxylated S-adenosylmethioninemore » (dcSAM), acting as an aminopropyl donor for the latter enzyme. Here we have explored the possibility of and identified the protein–protein interaction between SpdSyn and AdoMetDc. The protein–protein interaction has been identified using GST pull down assay. Isothermal titration calorimetry reveals that the interaction is thermodynamically favorable. Fluorescence spectroscopy studies also confirms the interaction, with SpdSyn exhibiting a change in tertiary structure with increasing concentrations of AdoMetDc. Size exclusion chromatography suggests the presence of the complex as a hetero-oligomer. Taken together, these results suggest that the enzymes indeed form a heteromer. Computational analyses suggest that this complex differs significantly from the corresponding human complex, implying that this complex could be a better therapeutic target than the individual enzymes.« less

Combined Effects and Cross-Interactions of Different Antibiotics and Polypeptides in Salmonella bredeney.

PubMed

Ju, Xiangyu; Zhu, Mengjiao; Han, Jinzhi; Lu, Zhaoxin; Zhao, Haizhen; Bie, Xiaomei

2018-05-24

Salmonella spp. are health-threatening foodborne pathogens. The increasingly common spread of antibiotic-resistant Salmonella spp. is a major public healthcare issue worldwide. In this study, we wished to explore (1) antibiotic or polypeptide combinations to inhibit multidrug-resistant Salmonella bredeney and (2) the regulation of cross-resistance and collateral sensitivity of antibiotics and polypeptides. We undertook a study to select antibiotic combinations. Then, we promoted drug-resistant strains of S. bredeney after 15 types of antibiotic treatment. From each evolving population, the S. bredeney strain was exposed to a particular single drug. Then, we analyzed how the evolved S. bredeney strains acquired resistance or susceptibility to other drugs. A total of 105 combinations were tested against S. bredeney following the protocols of CLSI-2016 and EUCAST-2017. The synergistic interactions between drug pairings were diverse. Notably, polypeptides were more likely to be linked to synergistic combinations: 56% (19/34) of the synergistic pairings were relevant to polypeptides. Simultaneously, macrolides demonstrated antagonism toward polypeptides. The latter were more frequently related to collateral sensitivity than the other drugs because the other 13 drugs sensitized S. bredeney to polypeptides. In an experimental evolution involving 15 drugs, single drug-evolved strains were examined against the other 14 drugs, and the results were compared with the minimal inhibitory concentration of the ancestral strain. Single drug-evolved S. bredeney strains could alter the sensitivity to other drugs, and S. bredeney evolution against antibiotics could sensitize it to polypeptides.

α-Synuclein Activates Innate Immunity but Suppresses Interferon-γ Expression in Murine Astrocytes.

PubMed

Wang, Jintang; Chen, Zheng; Walston, Jeremy; Gao, Peisong; Gao, Maolong; Leng, Sean X

2018-05-19

Glial activation and neuroinflammation contribute to pathogenesis of neurodegenerative diseases, linked to neuron loss and dysfunction. α-Synuclein (α-syn), as a metabolite of neuron, can induce microglia activation to trigger innate immune response. However, whether α-syn, as well as its mutants (A53T, A30P and E46K), induces astrocyte activation and inflammatory response is not fully elucidated. In this study, we used A53T mutant and wildtype α-syns to stimulate primary astrocytes in dose- and time-dependent manners (0.5, 2, 8 and 20 μg/mL for 24 hour or 3, 12, 24 and 48 hour at 2 μg/mL), and evaluated activation of several canonical inflammatory pathway components. The results showed that A53T mutant or wildtype α-syn significantly upregulated mRNA expression of toll-like receptor (TLR)2, TLR3, nuclear factor-κB and interleukin (IL)-1β, displaying a pattern of positive dose-effect correlation or negative time-effect correlation. Such upregulation was confirmed at protein levels of TLR2 (at 20 μg/mL), TLR3 (at most doses) and IL-1β (at 3 hour) by western blotting. Blockage of TLR2 other than TLR4 inhibited TLR3 and IL-1β mRNA expressions. By contrast, interferon (IFN)-γ was significantly downregulated at mRNA, protein and protein release levels, especially at high concentrations of α-syns or early time-points. These findings indicate that α-syn was a TLRs-mediated immunogenic agent (A53T mutant stronger than wildtype α-syn). The stimulation patterns suggest that persistent release and accumulation of α-syn is required for maintenance of innate immunity activation, and IFN-γ expression inhibition by α-syn suggests a novel immune molecule interaction mechanism underlying pathogenesis of neurodegenerative diseases. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

Dispersion interactions between neighboring Bi atoms in (BiH3 )2 and Te(BiR2 )2.

PubMed

Haack, Rebekka; Schulz, Stephan; Jansen, Georg

2018-03-13

Triggered by the observation of a short Bi⋯Bi distance and a BiTeBi bond angle of only 86.6° in the crystal structure of bis(diethylbismuthanyl)tellurane quantum chemical computations on interactions between neighboring Bi atoms in Te(BiR 2 ) 2 molecules (R = H, Me, Et) and in (BiH 3 ) 2 were undertaken. Bi⋯Bi distances atoms were found to significantly shorten upon inclusion of the d shells of the heavy metal atoms into the electron correlation treatment, and it was confirmed that interaction energies from spin component-scaled second-order Møller-Plesset theory (SCS-MP2) agree well with coupled-cluster singles and doubles theory including perturbative triples (CCSD(T)). Density functional theory-based symmetry-adapted perturbation theory (DFT-SAPT) was used to study the anisotropy of the interplay of dispersion attraction and steric repulsion between the Bi atoms. Finally, geometries and relative stabilities of syn-syn and syn-anti conformers of Te(BiR 2 ) 2 (R = H, Me, Et) and interconversion barriers between them were computed. © 2018 Wiley Periodicals, Inc. © 2018 Wiley Periodicals, Inc.

Aggregates assembled from overexpression of wild-type alpha-synuclein are not toxic to human neuronal cells.

PubMed

Ko, Li-Wen; Ko, Hwai-Hwa C; Lin, Wen-Lang; Kulathingal, Jayanranyan G; Yen, Shu-Hui C

2008-11-01

Filamentous alpha-synuclein (alpha-syn) aggregates form Lewy bodies (LBs), the neuropathologic hallmarks of Parkinson disease and related alpha-synucleinopathies. To model Lewy body-associated neurodegeneration, we generated transfectant 3D5 of human neuronal-type in which expression of human wild-type alpha-syn is regulated by the tetracycline off (TetOff)-inducible mechanism. Retinoic acid-elicited differentiation promoted assembly of alpha-syn aggregates after TetOff induction in 3D5 cells. The aggregates accumulated 14 days after TetOff induction were primarily soluble and showed augmented thioflavin affinity with concomitant phosphorylation and nitration of alpha-syn. Extension of the induction led to the formation of sarkosyl-insoluble aggregates that appeared concurrently with thioflavin-positive inclusions. Immunoelectron microscopy revealed that the inclusions consist of dense bundles of 8- to 12-nm alpha-syn fibrils that congregate in the perikarya and resemble Lewy bodies. Most importantly, accumulation of soluble and insoluble aggregates after TetOff induction for 14 and 28 days was reversible and did not compromise the viability of the cells or their subsequent survival. Thus, this chemically defined culture paradigm provides a useful means to elucidate how oxidative injuries and other insults that are associated with aging promote alpha-syn to self-assemble or interact with other molecules leading to neuronal degeneration in alpha-synucleinopathies.

The Caenorhabditis elegans gene unc-89, required fpr muscle M-line assembly, encodes a giant modular protein composed of Ig and signal transduction domains

PubMed Central

1996-01-01

Mutations in the Caenorhabditis elegans gene unc-89 result in nematodes having disorganized muscle structure in which thick filaments are not organized into A-bands, and there are no M-lines. Beginning with a partial cDNA from the C. elegans sequencing project, we have cloned and sequenced the unc-89 gene. An unc-89 allele, st515, was found to contain an 84-bp deletion and a 10-bp duplication, resulting in an in- frame stop codon within predicted unc-89 coding sequence. Analysis of the complete coding sequence for unc-89 predicts a novel 6,632 amino acid polypeptide consisting of sequence motifs which have been implicated in protein-protein interactions. UNC-89 begins with 67 residues of unique sequences, SH3, dbl/CDC24, and PH domains, 7 immunoglobulins (Ig) domains, a putative KSP-containing multiphosphorylation domain, and ends with 46 Ig domains. A polyclonal antiserum raised to a portion of unc-89 encoded sequence reacts to a twitchin-sized polypeptide from wild type, but truncated polypeptides from st515 and from the amber allele e2338. By immunofluorescent microscopy, this antiserum localizes to the middle of A-bands, consistent with UNC-89 being a structural component of the M-line. Previous studies indicate that myofilament lattice assembly begins with positional cues laid down in the basement membrane and muscle cell membrane. We propose that the intracellular protein UNC-89 responds to these signals, localizes, and then participates in assembling an M-line. PMID:8603916

α-Synuclein-induced lysosomal dysfunction occurs through disruptions in protein trafficking in human midbrain synucleinopathy models.

PubMed

Mazzulli, Joseph R; Zunke, Friederike; Isacson, Ole; Studer, Lorenz; Krainc, Dimitri

2016-02-16

Parkinson's disease (PD) is an age-related neurodegenerative disorder characterized by the accumulation of protein aggregates comprised of α-synuclein (α-syn). A major barrier in treatment discovery for PD is the lack of identifiable therapeutic pathways capable of reducing aggregates in human neuronal model systems. Mutations in key components of protein trafficking and cellular degradation machinery represent important risk factors for PD; however, their precise role in disease progression and interaction with α-syn remains unclear. Here, we find that α-syn accumulation reduced lysosomal degradation capacity in human midbrain dopamine models of synucleinopathies through disrupting hydrolase trafficking. Accumulation of α-syn at the cell body resulted in aberrant association with cis-Golgi-tethering factor GM130 and disrupted the endoplasmic reticulum-Golgi localization of rab1a, a key mediator of vesicular transport. Overexpression of rab1a restored Golgi structure, improved hydrolase trafficking and activity, and reduced pathological α-syn in patient neurons. Our work suggests that enhancement of lysosomal hydrolase trafficking may prove beneficial in synucleinopathies and indicates that human midbrain disease models may be useful for identifying critical therapeutic pathways in PD and related disorders.

Transcriptional profiling of striatal neurons in response to single or concurrent activation of dopamine D2, adenosine A(2A) and metabotropic glutamate type 5 receptors: focus on beta-synuclein expression.

PubMed

Canela, Laia; Selga, Elisabet; García-Martínez, Juan Manuel; Amaral, Olavo B; Fernández-Dueñas, Víctor; Alberch, Jordi; Canela, Enric I; Franco, Rafael; Noé, Véronique; Lluís, Carme; Ciudad, Carlos J; Ciruela, Francisco

2012-10-25

G protein-coupled receptor oligomerization is a concept which is changing the understanding of classical pharmacology. Both, oligomerization and functional interaction between adenosine A(2A,) dopamine D(2) and metabotropic glutamate type 5 receptors have been demonstrated in the striatum. However, the transcriptional consequences of receptors co-activation are still unexplored. We aim here to determine the changes in gene expression of striatal primary cultured neurons upon isolated or simultaneous receptor activation. Interestingly, we found that 95 genes of the total analyzed (15,866 transcripts and variants) changed their expression in response to simultaneous stimulation of all three receptors. Among these genes, we focused on the β-synuclein (β-Syn) gene (SCNB). Quantitative PCR verified the magnitude and direction of change in expression of SCNB. Since β-Syn belongs to the homologous synuclein family and may be considered a natural regulator of α-synuclein (α-Syn), it has been proposed that β-Syn might act protectively against α-Syn neuropathology. Copyright © 2012 Elsevier B.V. All rights reserved.

A "push and slide" mechanism allows sequence-insensitive translocation of secretory proteins by the SecA ATPase.

PubMed

Bauer, Benedikt W; Shemesh, Tom; Chen, Yu; Rapoport, Tom A

2014-06-05

In bacteria, most secretory proteins are translocated across the plasma membrane by the interplay of the SecA ATPase and the SecY channel. How SecA moves a broad range of polypeptide substrates is only poorly understood. Here we show that SecA moves polypeptides through the SecY channel by a "push and slide" mechanism. In its ATP-bound state, SecA interacts through a two-helix finger with a subset of amino acids in a substrate, pushing them into the channel. A polypeptide can also passively slide back and forth when SecA is in the predominant ADP-bound state or when SecA encounters a poorly interacting amino acid in its ATP-bound state. SecA performs multiple rounds of ATP hydrolysis before dissociating from SecY. The proposed push and slide mechanism is supported by a mathematical model and explains how SecA allows translocation of a wide range of polypeptides. This mechanism may also apply to hexameric polypeptide-translocating ATPases. Copyright © 2014 Elsevier Inc. All rights reserved.

Phosphorylated α-Synuclein Accumulations and Lewy Body-like Pathology Distributed in Parkinson's Disease-Related Brain Areas of Aged Rhesus Monkeys Treated with MPTP.

PubMed

Huang, Baihui; Wu, Shihao; Wang, Zhengbo; Ge, Longjiao; Rizak, Joshua D; Wu, Jing; Li, Jiali; Xu, Lin; Lv, Longbao; Yin, Yong; Hu, Xintian; Li, Hao

2018-05-21

Phosphorylation of α-synuclein at serine 129 (P-Ser 129 α-syn) is involved in the pathogenesis of Parkinson's disease (PD) and Lewy body (LB) formation. However, there is no clear evidence indicates the quantitative relation of P-Ser 129 α-syn accumulation and dopaminergic cell loss, LBs pathology and the affected brain areas in PD monkeys. Here, pathological changes in the substantia nigra (SN) and PD-related brain areas were measured in aged monkeys treated with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) utilizing a modeling-recovery-remodeling strategy. Compared to age-matched controls, the MPTP-treated monkeys showed significantly reduced tyrosine hydroxylase (TH)-positive neurons and increased P-Ser 129 α-syn-positive aggregations in the SN. Double-labeling Immunofluorescence found some TH-positive neurons to be co-localized with P-Ser129 α-syn in the SN, suggesting the inverse correlation between P-Ser 129 α-syn aggregations and dopaminergic cell loss in the SN may represent an interactive association related to the progression of the PD symptoms in the model. P-Ser 129 α-syn aggregations or LB-like pathology was also found in the midbrain and the neocortex, specifically in the oculomotor nucleus (CN III), temporal cortex (TC), prefrontal cortex (PFC) and in cells surrounding the third ventricle. Notably, the occipital cortex (OC) was P-Ser 129 α-syn negative. The findings of LB-like pathologies, dopaminergic cell loss and the stability of the PD symptoms in this model suggest that the modeling-recovery-remodeling strategy in aged monkeys may provide a new platform for biomedical investigations into the pathogenesis of PD and potential therapeutic development. Copyright © 2018 IBRO. Published by Elsevier Ltd. All rights reserved.

Immunity to East Coast fever in cattle induced by a polypeptide fragment of the major surface coat protein of Theileria parva sporozoites.

PubMed

Bishop, Richard; Nene, Vishvanath; Staeyert, Jan; Rowlands, John; Nyanjui, John; Osaso, Julius; Morzaria, Subhash; Musoke, Antony

2003-03-07

Full-length recombinant versions of p67, the 709 amino acid major surface protein of Theileria parva sporozoites, induce immunity to East Coast fever (ECF) in cattle. We show that a soluble Escherichia coli recombinant version of p67 (p67(635)), in which a prokaryotic signal peptide replaces the eukaryotic one, confers protection comparable to that induced by the full-length molecule, but is unstable. Peptides encoding 80 (p67C) and 205 (p67N) amino acid fragments of p67, containing epitopes recognised by sporozoite neutralising monoclonal antibodies, exhibit improved stability in E. coli. Antibodies raised against the central region of p67 (p67M) neutralise sporozoite infectivity in vitro. The p67C peptide induced immunity against ECF in cattle, at a level equivalent to p67(635), suggesting that a synthetic peptide vaccine might be achievable. Copyright 2002 Elsevier Science Ltd.

A de novo designed 11 kDa polypeptide: model for amyloidogenic intrinsically disordered proteins.

PubMed

Topilina, Natalya I; Ermolenkov, Vladimir V; Sikirzhytski, Vitali; Higashiya, Seiichiro; Lednev, Igor K; Welch, John T

2010-07-01

A de novo polypeptide GH(6)[(GA)(3)GY(GA)(3)GE](8)GAH(6) (YE8) has a significant number of identical weakly interacting beta-strands with the turns and termini functionalized by charged amino acids to control polypeptide folding and aggregation. YE8 exists in a soluble, disordered form at neutral pH but is responsive to changes in pH and ionic strength. The evolution of YE8 secondary structure has been successfully quantified during all stages of polypeptide fibrillation by deep UV resonance Raman (DUVRR) spectroscopy combined with other morphological, structural, spectral, and tinctorial characterization. The YE8 folding kinetics at pH 3.5 are strongly dependent on polypeptide concentration with a lag phase that can be eliminated by seeding with a solution of folded fibrillar YE8. The lag phase of polypeptide folding is concentration dependent leading to the conclusion that beta-sheet folding of the 11-kDa amyloidogenic polypeptide is completely aggregation driven.

Interactions between ethanol, syn-2,3-hexanediol, 3-hydroxyhexan-2-one, and 3-hydroxyoctan-2-one lures on trap catches of hardwood longhorn beetles in southeastern united states

Treesearch

D R Miller; C M Crowe; P D Mayo; L S Reid; P J Silk; J D Sweeney

2017-01-01

The effectiveness of a four-component “super lure” consisting of ethanol (E) and the cerambycid pheromones syn-2,3-hexanediol (D6), racemic 3-hydroxyhexan-2-one (K6), and racemic 3-hydroxyoctan-2-one (K8) on trap catches of Cerambycidae (Coleoptera) was determined in southeast United States with seven trapping experiments in 2011–2013. We captured...

Proof of principle in a de novo designed protein maquette: an allosterically regulated, charge-activated conformational switch in a tetra-alpha-helix bundle.

PubMed

Grosset, A M; Gibney, B R; Rabanal, F; Moser, C C; Dutton, P L

2001-05-08

New understanding of the engineering and allosteric regulation of natural protein conformational switches (such as those that couple chemical and ionic signals, mechanical force, and electro/chemical free energy for biochemical activation, catalysis, and motion) can be derived from simple de novo designed synthetic protein models (maquettes). We demonstrate proof of principle of both reversible switch action and allosteric regulation in a tetra-alpha-helical bundle protein composed of two identical di-helical subunits containing heme coordinated at a specific position close to the disulfide loop region. Individual bundles assume one of two switch states related by large-scale mechanical changes: a syn-topology (helices of the different subunits parallel) or anti-topology (helices antiparallel). Both the spectral properties of a coproporphyrin probe appended to the loop region and the distance-dependent redox interaction between the hemes identify the topologies. Beginning from a syn-topology, introduction of ferric heme in each subunit (either binding or redox change) shifts the topological balance by 25-50-fold (1.9-2.3 kcal/mol) to an anti-dominance. Charge repulsion between the two internal cationic ferric hemes drives the syn- to anti-switch, as demonstrated in two ways. When fixed in the syn-topology, the second ferric heme binding is 25-80-fold (1.9-2.6 kcal/mol) weaker than the first, and adjacent heme redox potentials are split by 80 mV (1.85 kcal/mol), values that energetically match the shift in topological balance. Allosteric and cooperative regulation of the switch by ionic strength exploits the shielded charge interactions between the two hemes and the exposed, cooperative interactions between the coproporphyrin carboxylates.

Interaction of β-sheet folds with a gold surface.

PubMed

Hoefling, Martin; Monti, Susanna; Corni, Stefano; Gottschalk, Kay Eberhard

2011-01-01

The adsorption of proteins on inorganic surfaces is of fundamental biological importance. Further, biomedical and nanotechnological applications increasingly use interfaces between inorganic material and polypeptides. Yet, the underlying adsorption mechanism of polypeptides on surfaces is not well understood and experimentally difficult to analyze. Therefore, we investigate here the interactions of polypeptides with a gold(111) surface using computational molecular dynamics (MD) simulations with a polarizable gold model in explicit water. Our focus in this paper is the investigation of the interaction of polypeptides with β-sheet folds. First, we concentrate on a β-sheet forming model peptide. Second, we investigate the interactions of two domains with high β-sheet content of the biologically important extracellular matrix protein fibronectin (FN). We find that adsorption occurs in a stepwise mechanism both for the model peptide and the protein. The positively charged amino acid Arg facilitates the initial contact formation between protein and gold surface. Our results suggest that an effective gold-binding surface patch is overall uncharged, but contains Arg for contact initiation. The polypeptides do not unfold on the gold surface within the simulation time. However, for the two FN domains, the relative domain-domain orientation changes. The observation of a very fast and strong adsorption indicates that in a biological matrix, no bare gold surfaces will be present. Hence, the bioactivity of gold surfaces (like bare gold nanoparticles) will critically depend on the history of particle administration and the proteins present during initial contact between gold and biological material. Further, gold particles may act as seeds for protein aggregation. Structural re-organization and protein aggregation are potentially of immunological importance.

Chirality-selected phase behaviour in ionic polypeptide complexes

DOE PAGES

Perry, Sarah L.; Leon, Lorraine; Hoffmann, Kyle Q.; ...

2015-01-14

In this study, polyelectrolyte complexes present new opportunities for self-assembled soft matter. Factors determining whether the phase of the complex is solid or liquid remain unclear. Ionic polypeptides enable examination of the effects of stereochemistry on complex formation. Here we demonstrate that chirality determines the state of polyelectrolyte complexes, formed from mixing dilute solutions of oppositely charged polypeptides, via a combination of electrostatic and hydrogen-bonding interactions. Fluid complexes occur when at least one of the polypeptides in the mixture is racemic, which disrupts backbone hydrogen-bonding networks. Pairs of purely chiral polypeptides, of any sense, form compact, fibrillar solids with amore » β-sheet structure. Analogous behaviour occurs in micelles formed from polypeptide block copolymers with polyethylene oxide, where assembly into aggregates with either solid or fluid cores, and eventually into ordered phases at high concentrations, is possible. Chirality is an exploitable tool for manipulating material properties in polyelectrolyte complexation.« less

Characterization of Insulin-Immunoreactive Cells and Endocrine Cells Within the Duct System of the Adult Human Pancreas.

PubMed

Li, Rong; Zhang, Xiaoxi; Yu, Lan; Zou, Xia; Zhao, Hailu

2016-01-01

The adult pancreatic duct system accommodates endocrine cells that have the potential to produce insulin. Here we report the characterization and distribution of insulin-immunoreactive cells and endocrine cells within the ductal units of adult human pancreas. Sequential pancreas sections from 12 nondiabetic adults were stained with biomarkers of ductal epithelial cells (cytokeratin 19), acinar cells (amylase), endocrine cells (chromogranin A; neuron-specific enolase), islet hormones (insulin, glucagon, somatostatin, pancreatic polypeptide), cell proliferation (Ki-67), and neogenesis (CD29). The number of islet hormone-immunoreactive cells increased from large ducts to the terminal branches. The insulin-producing cells outnumbered endocrine cells reactive for glucagon, somatostatin, or pancreatic polypeptide. The proportions of insulin-immunoreactive count compared with local islets (100% as a baseline) were 1.5% for the main ducts, 7.2% for interlobular ducts, 24.8% for intralobular ducts, 67.9% for intercalated ducts, and 348.9% for centroacinar cells. Both Ki-67- and CD29-labeled cells were predominantly localized in the terminal branches around the islets. The terminal branches also showed cells coexpressing islet hormones and cytokeratin 19. The adult human pancreatic ducts showed islet hormone-producing cells. The insulin-reactive cells predominantly localized in terminal branches where they may retain potential capability for β-cell neogenesis.

Cooperative polymerization of α-helices induced by macromolecular architecture

NASA Astrophysics Data System (ADS)

Baumgartner, Ryan; Fu, Hailin; Song, Ziyuan; Lin, Yao; Cheng, Jianjun

2017-07-01

Catalysis observed in enzymatic processes and protein polymerizations often relies on the use of supramolecular interactions and the organization of functional elements in order to gain control over the spatial and temporal elements of fundamental cellular processes. Harnessing these cooperative interactions to catalyse reactions in synthetic systems, however, remains challenging due to the difficulty in creating structurally controlled macromolecules. Here, we report a polypeptide-based macromolecule with spatially organized α-helices that can catalyse its own formation. The system consists of a linear polymeric scaffold containing a high density of initiating groups from which polypeptides are grown, forming a brush polymer. The folding of polypeptide side chains into α-helices dramatically enhances the polymerization rate due to cooperative interactions of macrodipoles between neighbouring α-helices. The parameters that affect the rate are elucidated by a two-stage kinetic model using principles from nucleation-controlled protein polymerizations; the key difference being the irreversible nature of this polymerization.

H3K9me2/3 Binding of the MBT Domain Protein LIN-61 Is Essential for Caenorhabditis elegans Vulva Development

PubMed Central

Koester-Eiserfunke, Nora; Fischle, Wolfgang

2011-01-01

MBT domain proteins are involved in developmental processes and tumorigenesis. In vitro binding and mutagenesis studies have shown that individual MBT domains within clustered MBT repeat regions bind mono- and dimethylated histone lysine residues with little to no sequence specificity but discriminate against the tri- and unmethylated states. However, the exact function of promiscuous histone methyl-lysine binding in the biology of MBT domain proteins has not been elucidated. Here, we show that the Caenorhabditis elegans four MBT domain protein LIN-61, in contrast to other MBT repeat factors, specifically interacts with histone H3 when methylated on lysine 9, displaying a strong preference for di- and trimethylated states (H3K9me2/3). Although the fourth MBT repeat is implicated in this interaction, H3K9me2/3 binding minimally requires MBT repeats two to four. Further, mutagenesis of residues conserved with other methyl-lysine binding MBT regions in the fourth MBT repeat does not abolish interaction, implicating a distinct binding mode. In vivo, H3K9me2/3 interaction of LIN-61 is required for C. elegans vulva development within the synMuvB pathway. Mutant LIN-61 proteins deficient in H3K9me2/3 binding fail to rescue lin-61 synMuvB function. Also, previously identified point mutant synMuvB alleles are deficient in H3K9me2/3 interaction although these target residues that are outside of the fourth MBT repeat. Interestingly, lin-61 genetically interacts with two other synMuvB genes, hpl-2, an HP1 homologous H3K9me2/3 binding factor, and met-2, a SETDB1 homologous H3K9 methyl transferase (H3K9MT), in determining C. elegans vulva development and fertility. Besides identifying the first sequence specific and di-/trimethylation binding MBT domain protein, our studies imply complex multi-domain regulation of ligand interaction of MBT domains. Our results also introduce a mechanistic link between LIN-61 function and biology, and they establish interplay of the H3K9me2/3 binding proteins, LIN-61 and HPL-2, as well as the H3K9MT MET-2 in distinct developmental pathways. PMID:21437264

On the photoisomerization of 5-hydroxytropolone: An ab initio and nuclear wave function study

NASA Astrophysics Data System (ADS)

Paz, Juan J.; Moreno, Miquel; Lluch, José M.

1997-10-01

In this paper we perform ab initio calculations for the stable conformations and the transition states for the isomerization processes in 5-hydroxytropolone in both the ground (S0) and first excited (S1) singlet electronic states. The Hartree-Fock self-consistent field (SCF) level and a complete active space SCF (CASSCF) level for S0 are considered, whereas the configuration interaction all single excitation method (CIS) and the CASSCF levels are used to deal with the S1 state. Energies are reevaluated at all levels through perturbation theory up to second order: Møller-Plesset for the Hartree-Fock and CIS methods, and the CASPT2 method for CAS results. The ab initio results are then used to perform different monodimensional fits to the potential energy surfaces in order to analyze the wave functions for the nuclear motions in both electronic states. Our best results predict that for the S0 state two stable conformers, syn and anti, can exist in thermal equilibrium. In accordance with experimental expectations the syn isomer is the most stable. As for the S1 state, and again in accord with experimental spectroscopical data, the order of stability reverses, the anti being the most stable. A more interesting result is that analysis of the nuclear wave functions shows an important syn-anti mixing in the S1 state that does not appear in S0. This result explains the appearance of syn-anti and anti-syn crossover transitions observed in the electronic spectra of 5-hydroxytropolone so that syn-anti reaction may take place through photoisomerization.

Interaction of β-Sheet Folds with a Gold Surface

PubMed Central

Hoefling, Martin; Monti, Susanna; Corni, Stefano; Gottschalk, Kay Eberhard

2011-01-01

The adsorption of proteins on inorganic surfaces is of fundamental biological importance. Further, biomedical and nanotechnological applications increasingly use interfaces between inorganic material and polypeptides. Yet, the underlying adsorption mechanism of polypeptides on surfaces is not well understood and experimentally difficult to analyze. Therefore, we investigate here the interactions of polypeptides with a gold(111) surface using computational molecular dynamics (MD) simulations with a polarizable gold model in explicit water. Our focus in this paper is the investigation of the interaction of polypeptides with β-sheet folds. First, we concentrate on a β-sheet forming model peptide. Second, we investigate the interactions of two domains with high β-sheet content of the biologically important extracellular matrix protein fibronectin (FN). We find that adsorption occurs in a stepwise mechanism both for the model peptide and the protein. The positively charged amino acid Arg facilitates the initial contact formation between protein and gold surface. Our results suggest that an effective gold-binding surface patch is overall uncharged, but contains Arg for contact initiation. The polypeptides do not unfold on the gold surface within the simulation time. However, for the two FN domains, the relative domain-domain orientation changes. The observation of a very fast and strong adsorption indicates that in a biological matrix, no bare gold surfaces will be present. Hence, the bioactivity of gold surfaces (like bare gold nanoparticles) will critically depend on the history of particle administration and the proteins present during initial contact between gold and biological material. Further, gold particles may act as seeds for protein aggregation. Structural re-organization and protein aggregation are potentially of immunological importance. PMID:21687744

Effects of hydrophobic and dipole-dipole interactions on the conformational transitions of a model polypeptide

NASA Astrophysics Data System (ADS)

Mu, Yan; Gao, Yi Qin

2007-09-01

We studied the effects of hydrophobicity and dipole-dipole interactions between the nearest-neighbor amide planes on the secondary structures of a model polypeptide by calculating the free energy differences between different peptide structures. The free energy calculations were performed with low computational costs using the accelerated Monte Carlo simulation (umbrella sampling) method, with a bias-potential method used earlier in our accelerated molecular dynamics simulations. It was found that the hydrophobic interaction enhances the stability of α helices at both low and high temperatures but stabilizes β structures only at high temperatures at which α helices are not stable. The nearest-neighbor dipole-dipole interaction stabilizes β structures under all conditions, especially in the low temperature region where α helices are the stable structures. Our results indicate clearly that the dipole-dipole interaction between the nearest neighboring amide planes plays an important role in determining the peptide structures. Current research provides a more unified and quantitative picture for understanding the effects of different forms of interactions on polypeptide structures. In addition, the present model can be extended to describe DNA/RNA, polymer, copolymer, and other chain systems.

Pancreatic cholera syndrome: effect of a synthetic somatostatin analog on intestinal water and ion transport.

PubMed

Santangelo, W C; O'Dorisio, T M; Kim, J G; Severino, G; Krejs, G J

1985-09-01

The effect of a synthetic somatostatin analog was studied in a patient with severe secretory diarrhea due to pancreatic cholera syndrome. Basal intestinal perfusion studies indicated an absence of water and sodium absorption, and active chloride secretion in the small bowel. Intravenous administration of the somatostatin analog (1 microgram/kg.h) changed zero net water movement to absorption (122 mL/30 cm of the jejunum per hour). Chloride secretion changed to absorption (5.0 to 7.9 meq/30 cm.h), and plasma vasoactive intestinal polypeptide concentration was reduced from 330 to 45 pmol/L (normal, less than 51). When the analog was given subcutaneously, 100 micrograms twice daily, stool weight decreased, and plasma vasoactive intestinal polypeptide concentration fell toward the normal range (67 pmol/L). Plasma concentration of pancreatic polypeptide was initially elevated and dropped during intravenous infusion of somatostatin analog but returned to baseline on maintenance therapy with the analog delivered subcutaneously. The patient has not had further diarrhea during 9 months of therapy.

Thermodynamic Approach to Enhanced Dispersion and Physical Properties in a Carbon Nanotube/Polypeptide Nanocomposite

NASA Technical Reports Server (NTRS)

Lovell, Conrad S.; Wise, Kristopher E.; Kim, Jae-Woo; Lillehei, Peter T.; Harrison, Joycelyn S.; Park, Cheol

2009-01-01

A high molecular weight synthetic polypeptide has been designed which exhibits favorable interactions with single wall carbon nanotubes (SWCNTs). The enthalpic and entropic penalties of mixing between these two molecules are reduced due to the polypeptide's aromatic sidechains and helical secondary structure, respectively. These enhanced interactions result in a well dispersed SWCNT/Poly (L-Leucine-ran-L-Phenylalanine) nanocomposite with enhanced mechanical and electrical properties using only shear mixing and sonication. At 0.5 wt% loading of SWCNT filler, the nanocomposite exhibits simultaneous increases in the Young's modulus, failure strain, and toughness of 8%, 120%, and 144%, respectively. At one kHz, the same nanotube loading level also enhances the dielectric constant from 2.95 to 22.81, while increasing the conductivity by four orders of magnitude.

Molecular description of the LCST behavior of an elastin-like polypeptide.

PubMed

Li, Nan K; García Quiroz, Felipe; Hall, Carol K; Chilkoti, Ashutosh; Yingling, Yaroslava G

2014-10-13

Elastin-like polypeptides (ELPs) with the repeat sequence of VPGVG are widely used as a model system for investigation of lower critical solution temperature (LCST) transition behavior. In this paper, the effect of temperature on the structure, dynamics and association of (VPGVG)18 in aqueous solution is investigated using atomistic molecular dynamics simulations. Our simulations show that as the temperature increases the ELP backbones undergo gradual conformational changes, which are attributed to the formation of more ordered secondary structures such as β-strands. In addition, increasing temperature changes the hydrophobicity of the ELP by exposure of hydrophobic valine-side chains to the solvent and hiding of proline residues. Based on our simulations, we conclude that the transition behavior of (VPGVG)18 can be attributed to a combination of thermal disruption of the water network that surrounds the polypeptide, reduction of solvent accessible surface area of the polypeptide, and increase in its hydrophobicity. Simulations of the association of two (VPGVG)18 molecules demonstrated that the observed gradual changes in the structural properties of the single polypeptide chain are enough to cause the aggregation of polypeptides above the LCST. These results lead us to propose that the LCST phase behavior of poly(VPGVG) is a collective phenomenon that originates from the correlated gradual changes in single polypeptide structure and the abrupt change in properties of hydration water around the peptide and is a result of a competition between peptide-peptide and peptide-water interactions. This is a computational study of an important intrinsically disordered peptide system that provides an atomic-level description of structural features and interactions that are relevant in the LCST phase behavior.

Battling the un-dead: the status of the Diptera genus-group names originally proposed in Johann Wilhelm Meigen's 1800 pamphlet.

PubMed

Evenhuis, Neal L; Pape, Thomas

2017-06-08

The work of Meigen 1800 was suppressed by the ICZN Commission in 1963 for the purposes of zoological nomenclature. The work as such is still to be treated as having been published and it remains available as a source of published descriptions and illustrations. Therefore, while the names in Meigen (1800) are deemed unavailable, a subsequent usage of any of the names may be considered a novel proposal. We review the first post-Meigen 1800 occurrence of each name, its first date of availability and authorship, and determine status and synonymy.        Designations of type species are given for the following genus-group names: Coryneta Hendel, 1908 [Hybotidae]; Cyanea Hendel, 1908 [Hippoboscidae].        Acting as First Reviser, we select the following as the correct original spelling from multiple original spellings: Calirrhoe Hendel, 1908.        New synonymies are proposed for the following: Ablabesmyia Johannsen, 1905 under Pelopia Latreille, 1802, n. syn. [Limoniidae]; Amasia Meigen in Hendel, 1908 under Penthetria Meigen, 1803, n. syn. [Bibionidae]; Amphinome Meigen in Hendel, 1908 under Limonia Meigen, 1803, n. syn. [Limoniidae]; Antiopa Meigen in Hendel, 1908 under Chrysotoxum Meigen, 1803, n. syn. [Syrphidae]; Apivora Meigen in Hendel, 1908 under Volucella Geoffroy, 1762, n. syn. [Syrphidae]; Atalanta Meigen in Hendel, 1908 under Clinocera Meigen, 1803, n. syn. [Empididae]; Calirrhoe Meigen & Hendel in Hendel, 1908 under Prosena Le Peletier & Audinet-Serville, 1828, n. syn. [Tachinidae]; Chrysozona Hendel, 1903 under Haematopota Meigen, 1803, n. syn. [Tabanidae]; Cinxia Meigen in Hendel, 1908 under Sericomyia Meigen, 1803, n. syn. [Syrphidae]; Cleona Meigen in Hendel 1908 under Callomyia Meigen, 1804, n. syn. [Platypezidae]; Clythia Hendel, 1903 under Platypeza Meigen, 1803, n. syn. [Platypezidae]; Coryneta Meigen in Hendel, 1908 under Tachydromia Meigen, 1803, n. syn. [Hybotidae]; Crocuta Bezzi, 1907 under Siphona Meigen, 1803, n. syn. [Tachinidae]; Cyanea Meigen in Hendel, 1908 under Melophagus Latreille, 1802, n. syn. [Hippoboscidae]; Cypsela Meigen in Hendel, 1908 under Sphaerocera Latreille, 1804, n. syn. [Sphaeroceridae]; Dionnaea Meigen in Hendel, 1908 under Rhamphomyia Meigen, 1822, n. syn. [Empididae]; Dorilas Meigen in Hendel, 1908 under Pipunculus Latreille, 1802, n. syn. [Pipunculidae]; Echinodes Meigen in Hendel, 1908 under Eriothrix Meigen, 1803, n. syn. [Tachinidae]; Erinna Hendel, 1903 under Xylophagus Meigen, 1803, n. syn. [Xylophagidae]; Eulalia Hendel, 1903 under Odontomyia Meigen, 1803, n. syn. [Stratiomyidae]; Euphrosyne Meigen in Hendel, 1908 under Macrocera Meigen, 1803, n. syn. [Keroplatidae]; Flabellifera Osten Sacken, 1882 under Tanyptera Latreille, 1804, n. syn. [Tipulidae]; Fungivora Meigen in Hendel, 1908 under Mycetophila Meigen, 1803, n. syn. [Mycetophilidae]; Helea Osten Sacken, 1882 under Ceratopogon Meigen, 1803, n. syn. [Ceratopogonidae]; Hermione Bezzi, 1908 under Oxycera Meigen, 1803, n. syn. [Stratiomyidae]; Itonida Bezzi, 1908 under Cecidomyia Meigen, 1803, n. syn. [Cecidomyiidae]; Lampetia Meigen in Hendel, 1908 under Merodon Meigen, 1803, n. syn. [Syrphidae]; Laphria Bezzi, 1907 under Laphria Meigen, 1803, n. syn. [Asilidae]; Lapria Bezzi, 1907 under Laphria Meigen, 1803, n. syn. [Asilidae]; Larvaevora Meigen in Hendel, 1908 under Tachina Meigen, 1803, n. syn. [Tachinidae]; Liriope Meigen in Hendel, 1908 under Ptychoptera Meigen, 1803, n. syn. [Ptychopteridae]; Lycoria Latreille, 1802 under Sylvicola Harris, 1776, n. syn. [Anisopodidae]; Melusina Meigen in Hendel, 1908 under Trichocera Meigen, 1803, n. syn. [Trichoceridae]; Musidora Meigen in Hendel, 1908 under Lonchoptera Meigen, 1803, n. syn. [Lonchopteridae]; Noeza Meigen in Hendel, 1908 under Hybos Meigen, 1803, n. syn. [Hybotidae]; Omphrale Meigen in Hendel, 1908 under Scenopinus Latreille, 1802, n. syn. [Scenopinidae]; Pales Bezzi, 1906 under Nephrotoma Meigen, 1803, n. syn . [Tipulidae]; Penthesilea Meigen in Hendel, 1908 under Blera Billberg, 1820, n. syn. [Syrphidae]; Petaurista Meigen in Hendel, 1908 under Trichocera Meigen, 1803, n. syn. [Trichoceridae]; Phalaenula Desmarest, 1818 under Psychoda Latreille, 1797, n. syn. [Psychodidae]; Philia Meigen in Hendel, 1908 under Dilophus Meigen, 1803, n. syn. [Bibionidae]; Phryne Meigen in Hendel, 1908 under Sylvicola Harris, 1776, n. syn. [Anisopodidae]; Polymeda Meigen in Hendel, 1908 under Erioptera Meigen, 1803, n. syn. [Limoniidae]; Polyxena Meigen in Hendel, 1908 under Cordyla Meigen, 1803, n. syn. [Mycetophilidae]; Potamida Hendel, 1903 under Clitellaria Meigen, 1803, n. syn. [Stratiomyidae]; Rhodogyne Meigen in Hendel, 1908 under Gymnosoma Meigen, 1803, n. syn. [Tachinidae]; Salmacia Meigen in Hendel, 1908 under Gonia Meigen, 1803, n. syn. [Tachinidae]; Scathophaga Meigen, 1803 under Scopeuma Latreille, 1802, n. syn. [Scathophagidae]; Coremacera Rondani, 1856 under Statinia Latreille, 1802, n. syn. [Sciomyzidae]; Tendipes Meigen in Hendel, 1908 under Chironomus Meigen, 1803, n. syn. [Chironomidae]; Titania Meigen in Hendel, 1908 under Chlorops Meigen, 1803, n. syn. [Chloropidae]; Trepidaria Meigen in Hendel, 1908 under Calobata Meigen, 1803, n. syn. [Micropezidae]; Tritonia Meigen in Hendel, 1908 under Temnostoma Le Peletier & Audinet-Serville, 1828, n. syn. [Syrphidae]; Tubifera Meigen in Hendel, 1908 under Eristalis Latreille, 1804, n. syn. [Syrphidae]; Urophora Robineau-Desvoidy, 1830 under Euribia Latreille, 1802, n. syn. [Tephritidae]; Zelima Hendel, 1903 under Xylota Meigen, 1822, n. syn. [Syrphidae]; Zelmira Meigen in Hendel, 1908 under Orfelia Costa, 1857, n. syn. [Keroplatidae].        The following three names have not been found to be synonymous with any other taxon, and are treated here as nomina dubia: Orithea Meigen in Hendel, 1908; Salpyga Meigen in Hendel, 1908; Titia Meigen in Hendel, 1908 (preoccupied).      The following four names are found to be senior synonyms of more commonly used genus-group names: Euribia Latreille, 1802 of Urophora Robineau-Desvoidy, 1830, n. syn. [Tephritidae]; Pelopia Latreille, 1802 of Ablabesmyia Johannsen, 1905, n. syn.; Scopeuma Latreille, 1802 of Scathophaga Meigen, 1803, n. syn. [Scathophagidae]; Statinia Latreille, 1802 of Coremacera Rondani, 1856, n. syn. [Sciomyzidae]. If they are construed as threatening stability of nomenclature and/or taxonomy, applications to the ICZN Commission may be warranted to request suppression of these names in favor of their junior synonyms.

A Bio-Inspired Two-Layer Sensing Structure of Polypeptide and Multiple-Walled Carbon Nanotube to Sense Small Molecular Gases

PubMed Central

Wang, Li-Chun; Su, Tseng-Hsiung; Ho, Cheng-Long; Yang, Shang-Ren; Chiu, Shih-Wen; Kuo, Han-Wen; Tang, Kea-Tiong

2015-01-01

In this paper, we propose a bio-inspired, two-layer, multiple-walled carbon nanotube (MWCNT)-polypeptide composite sensing device. The MWCNT serves as a responsive and conductive layer, and the nonselective polypeptide (40 mer) coating the top of the MWCNT acts as a filter into which small molecular gases pass. Instead of using selective peptides to sense specific odorants, we propose using nonselective, peptide-based sensors to monitor various types of volatile organic compounds. In this study, depending on gas interaction and molecular sizes, the randomly selected polypeptide enabled the recognition of certain polar volatile chemical vapors, such as amines, and the improved discernment of low-concentration gases. The results of our investigation demonstrated that the polypeptide-coated sensors can detect ammonia at a level of several hundred ppm and barely responded to triethylamine. PMID:25751078

Structural Effects of Two Camelid Nanobodies Directed to Distinct C-Terminal Epitopes on α-Synuclein.

PubMed

El-Turk, Farah; Newby, Francisco N; De Genst, Erwin; Guilliams, Tim; Sprules, Tara; Mittermaier, Anthony; Dobson, Christopher M; Vendruscolo, Michele

2016-06-07

α-Synuclein is an intrinsically disordered protein whose aggregation is associated with Parkinson's disease and other related neurodegenerative disorders. Recently, two single-domain camelid antibodies (nanobodies) were shown to bind α-synuclein with high affinity. Herein, we investigated how these two nanobodies (NbSyn2 and NbSyn87), which are directed to two distinct epitopes within the C-terminal domain of α-synuclein, affect the conformational properties of this protein. Our results suggest that nanobody NbSyn2, which binds to the five C-terminal residues of α-synuclein (residues 136-140), does not disrupt the transient long-range interactions that generate a degree of compaction within the native structural ensemble of α-synuclein. In contrast, the data that we report indicate that NbSyn87, which targets a central region within the C-terminal domain (residues 118-128), has more substantial effects on the fluctuating secondary and tertiary structure of the protein. These results are consistent with the different effects that the two nanobodies have on the aggregation behavior of α-synuclein in vitro. Our findings thus provide new insights into the type of effects that nanobodies can have on the conformational ensemble of α-synuclein.

Competition between surface adsorption and folding of fibril-forming polypeptides

NASA Astrophysics Data System (ADS)

Ni, Ran; Kleijn, J. Mieke; Abeln, Sanne; Cohen Stuart, Martien A.; Bolhuis, Peter G.

2015-02-01

Self-assembly of polypeptides into fibrillar structures can be initiated by planar surfaces that interact favorably with certain residues. Using a coarse-grained model, we systematically studied the folding and adsorption behavior of a β -roll forming polypeptide. We find that there are two different folding pathways depending on the temperature: (i) at low temperature, the polypeptide folds in solution into a β -roll before adsorbing onto the attractive surface; (ii) at higher temperature, the polypeptide first adsorbs in a disordered state and folds while on the surface. The folding temperature increases with increasing attraction as the folded β -roll is stabilized by the surface. Surprisingly, further increasing the attraction lowers the folding temperature again, as strong attraction also stabilizes the adsorbed disordered state, which competes with folding of the polypeptide. Our results suggest that to enhance the folding, one should use a weakly attractive surface. They also explain the recent experimental observation of the nonmonotonic effect of charge on the fibril formation on an oppositely charged surface [C. Charbonneau et al., ACS Nano 8, 2328 (2014), 10.1021/nn405799t].

A versatile expression vector for the growth and amplification of unmodified phage display polypeptides.

PubMed

Winton, Alexander J; Baptiste, Janae L; Allen, Mark A

2018-09-01

Proteins and polypeptides represent nature's most complex and versatile polymer. They provide complicated shapes, diverse chemical functionalities, and tightly regulated and controlled sizes. Several disease states are related to the misfolding or overproduction of polypeptides and yet polypeptides are present in several therapeutic molecules. In addition to biological roles; short chain polypeptides have been shown to interact with and drive the bio-inspired synthesis or modification of inorganic materials. This paper outlines the development of a versatile cloning vector which allows for the expression of a short polypeptide by controlling the incorporation of a desired DNA coding insert. As a demonstration of the efficacy of the expression system, a solid binding polypeptide identified from M13 phage display was expressed and purified. The solid binding polypeptide was expressed as a soluble 6xHis-SUMO tagged construct. Expression was performed in E. coli using auto-induction followed by Ni-NTA affinity chromatography and ULP1 protease cleavage. Methodology demonstrates the production of greater than 8 mg of purified polypeptide per liter of E. coli culture. Isotopic labeling of the peptide is also demonstrated. The versatility of the designed cloning vector, use of the 6xHis-SUMO solubility partner, bacterial expression in auto-inducing media and the purification methodology make this expressionun vector a readily scalable and user-friendly system for the creation of desired peptide domains. Copyright © 2018. Published by Elsevier Inc.

Characterization of antibodies that selectively detect alpha-synuclein in pathological inclusions.

PubMed

Waxman, Elisa A; Duda, John E; Giasson, Benoit I

2008-07-01

Sensitive detection of alpha-synuclein (alpha-syn) pathology is important in the diagnosis of disorders like Parkinson's disease, dementia with Lewy bodies, and multiple system atrophy and in providing better insights into the etiology of these diseases. Several monoclonal antibodies that selectively react with aggregated alpha-syn in pathological inclusions and reveal extensive and underappreciated alpha-syn pathology in the brains of diseased patients were previously reported by Duda et al. (Ann Neurol 52:205-210, 2002). We sought to characterize the specificity of some of these antibodies (Syn 505, Syn 506 and Syn 514); using C-terminal and N-terminal truncations of alpha-syn, all three antibodies were determined to require N-terminal epitopes that minimally comprise amino acids 2-4, but possibly extend to amino acid 12 of alpha-syn. The selectivity of these antibodies was further assessed using biochemical analysis of human brains and reactivity to altered recombinant alpha-syn proteins with duplication variants of amino acids 1-12. In addition, by expressing wild-type or a double mutant (E46K/A53T) of alpha-syn in cultured cells and by comparing their immunoreactivities to another antibody (SNL-4), which has a similar primary epitope, it was determined that Syn 505, Syn 506 and Syn 514 recognize conformational variants of alpha-syn that is enhanced by the presence of the double mutations. These studies indicate that antibodies Syn 505, Syn 506 and Syn 514 preferentially recognize N-terminal epitopes in complex conformations, consistent with the dramatic conformational change associated with the polymerization of alpha-synuclein into amyloid fibrils that form pathological inclusions.

Statistical thermodynamics of protein folding: Comparison of a mean-field theory with Monte Carlo simulations

NASA Astrophysics Data System (ADS)

Hao, Ming-Hong; Scheraga, Harold A.

1995-01-01

A comparative study of protein folding with an analytical theory and computer simulations, respectively, is reported. The theory is based on an improved mean-field formalism which, in addition to the usual mean-field approximations, takes into account the distributions of energies in the subsets of conformational states. Sequence-specific properties of proteins are parametrized in the theory by two sets of variables, one for the energetics of mean-field interactions and one for the distribution of energies. Simulations are carried out on model polypeptides with different sequences, with different chain lengths, and with different interaction potentials, ranging from strong biases towards certain local chain states (bond angles and torsional angles) to complete absence of local conformational preferences. Theoretical analysis of the simulation results for the model polypeptides reveals three different types of behavior in the folding transition from the statistical coiled state to the compact globular state; these include a cooperative two-state transition, a continuous folding, and a glasslike transition. It is found that, with the fitted theoretical parameters which are specific for each polypeptide under a different potential, the mean-field theory can describe the thermodynamic properties and folding behavior of the different polypeptides accurately. By comparing the theoretical descriptions with simulation results, we verify the basic assumptions of the theory and, thereby, obtain new insights about the folding transitions of proteins. It is found that the cooperativity of the first-order folding transition of the model polypeptides is determined mainly by long-range interactions, in particular the dipolar orientation; the local interactions (e.g., bond-angle and torsion-angle potentials) have only marginal effect on the cooperative characteristic of the folding, but have a large impact on the difference in energy between the folded lowest-energy structure and the unfolded conformations of a protein.

Epigallocatechin Gallate (EGCG) Inhibits Alpha-Synuclein Aggregation: A Potential Agent for Parkinson's Disease.

PubMed

Xu, Yan; Zhang, Yanyan; Quan, Zhenzhen; Wong, Winnie; Guo, Jianping; Zhang, Rongkai; Yang, Qinghu; Dai, Rongji; McGeer, Patrick L; Qing, Hong

2016-10-01

Protein aggregation is a prominent feature of many neurodegenerative disorders including Parkinson's disease (PD). Aggregation of alpha-synuclein (SNCA) may underlie the pathology of PD. They are the main components of Lewy bodies and dystrophic neurites that are the intraneuronal inclusions characteristic of the disease. We have demonstrated that the polyphenol (-)-epi-gallocatechine gallate (EGCG) inhibited SNCA aggregation, which made it a candidate for therapeutic intervention in PD. Three methods were used: SNCA fibril formation inhibition by EGCG in incubates; inhibition of the SNCA fluorophore A-Syn-HiLyte488 binding to plated SNCA in microwells; and inhibition of the A-Syn-HiLyte488 probe binding to aggregated SNCA in postmortem PD tissue. Recombinant human SNCA was incubated under conditions that result in fibril formation. The aggregation was blocked by 100 nM EGCG in a concentration-dependent manner, as shown by an absence of thioflavin T binding. In the microplate assay system, the ED 50 of EGCG inhibition of A-Syn-HiLyte488 binding to coated SNCA was 250 nM. In the PD tissue based assay, SNCA aggregates were recognized by incubation with 7 nM of A-Syn-HiLyte488. This binding was blocked by EGCG in a concentration dependent manner. The SNCA amino acid sites, which potentially interacted with EGCG, were detected on peptide membranes. It was implicated that EGCG binds to SNCA by instable hydrophobic interactions. In this study, we suggested that EGCG could be a potent remodeling agent of SNCA aggregates and a potential disease modifying drug for the treatment of PD and other α-synucleinopathies.

CERES SYN1deg Ed4 Product Removal

Atmospheric Science Data Center

2016-04-18

... The CERES Synoptic One Degree Edition4A product family, SYN1deg-1Hour, SYN1deg-3Hour, SYN1deg-MHour, SYN1deg-Day, and SYN1deg-Month with Configuration Code 400404 was pulled from public view after two errors were identified.   The first issue ...

Effects of Serine 129 Phosphorylation on α-Synuclein Aggregation, Membrane Association, and Internalization*

PubMed Central

Samuel, Filsy; Flavin, William P.; Iqbal, Sobia; Pacelli, Consiglia; Sri Renganathan, Sri Dushyaanthan; Trudeau, Louis-Eric; Campbell, Edward M.; Fraser, Paul E.; Tandon, Anurag

2016-01-01

Although trace levels of phosphorylated α-synuclein (α-syn) are detectable in normal brains, nearly all α-syn accumulated within Lewy bodies in Parkinson disease brains is phosphorylated on serine 129 (Ser-129). The role of the phosphoserine residue and its effects on α-syn structure, function, and intracellular accumulation are poorly understood. Here, co-expression of α-syn and polo-like kinase 2 (PLK2), a kinase that targets Ser-129, was used to generate phosphorylated α-syn for biophysical and biological characterization. Misfolding and fibril formation of phosphorylated α-syn isoforms were detected earlier, although the fibrils remained phosphatase- and protease-sensitive. Membrane binding of α-syn monomers was differentially affected by phosphorylation depending on the Parkinson disease-linked mutation. WT α-syn binding to presynaptic membranes was not affected by phosphorylation, whereas A30P α-syn binding was greatly increased, and A53T α-syn was slightly lower, implicating distal effects of the carboxyl- on amino-terminal membrane binding. Endocytic vesicle-mediated internalization of pre-formed fibrils into non-neuronal cells and dopaminergic neurons matched the efficacy of α-syn membrane binding. Finally, the disruption of internalized vesicle membranes was enhanced by the phosphorylated α-syn isoforms, a potential means for misfolded extracellular or lumenal α-syn to access cytosolic α-syn. Our results suggest that the threshold for vesicle permeabilization is evident even at low levels of α-syn internalization and are relevant to therapeutic strategies to reduce intercellular propagation of α-syn misfolding. PMID:26719332

Measurement of lipocalin-2 and syndecan-4 levels to differentiate bacterial from viral infection in children with community-acquired pneumonia.

PubMed

Esposito, Susanna; Bianchini, Sonia; Gambino, Monia; Madini, Barbara; Di Pietro, Giada; Umbrello, Giulia; Presicce, Maria Lory; Ruggiero, Luca; Terranova, Leonardo; Principi, Nicola

2016-07-20

In this study, we evaluated the lipocalin-2 (LIP2) and syndecan-4 (SYN4) levels in children who were hospitalized for radiologically confirmed CAP in order to differentiate bacterial from viral infection. The results regarding the LIP2 and SYN4 diagnostic outcomes were compared with the white blood cell (WBC) count and C reactive protein (CRP) levels. A total of 110 children <14 years old who were hospitalized for radiologically confirmed CAP were enrolled. Serum samples were obtained upon admission and on day 5 to measure the levels of LIP2, SYN4, and CRP as well as the WBC. Polymerase chain reaction of the respiratory secretions and tests on blood samples were performed to detect respiratory viruses, Streptococcus pneumoniae, and Mycoplasma pneumoniae. CAP was considered to be due to a probable bacterial infection in 74 children (67.3 %) and due to a probable viral infection in 16 children (14.5 %). Overall, 84 children (76.4 %) were diagnosed with severe CAP. The mean values of the WBC count and the LIP2 and SYN4 levels did not differ among the probable bacterial, probable viral, and undetermined cases. However, the CRP serum concentrations were significantly higher in children with probable bacterial CAP than in those with probable viral disease (32.2 ± 55.5 mg/L vs 9.4 ± 17.0 mg/L, p < 0.05). The WBC count was the best predictor of severe CAP, but the differences among the studied variables were marginal. The WBC count was significantly lower on day 5 in children with probable bacterial CAP (p < 0.01) and in those with an undetermined etiology (p < 0.01). The CRP and LIP2 levels were significantly lower 5 days after enrollment in all of the studied groups, independent of the supposed etiology of CAP (p < 0.01 for all comparisons). No statistically significant variation was observed for SYN4. Measuring the LIP2 and SYN4 levels does not appear to solve the problem of the poor reliability of routine laboratory tests in defining the etiology and severity of pediatric CAP. Currently, the CRP levels and WBC, when combined with evaluation of clinical data, can be used to limit the overuse of antibiotics as much as possible and to provide the best treatment to the patient.

Crystallization of calcium oxalates is controlled by molecular hydrophilicity and specific polyanion-crystal interactions.

PubMed

Grohe, Bernd; Taller, Adam; Vincent, Peter L; Tieu, Long D; Rogers, Kem A; Heiss, Alexander; Sørensen, Esben S; Mittler, Silvia; Goldberg, Harvey A; Hunter, Graeme K

2009-10-06

To gain more insight into protein structure-function relationships that govern ectopic biomineralization processes in kidney stone formation, we have studied the ability of urinary proteins (Tamm-Horsfall protein, osteopontin (OPN), prothrombin fragment 1 (PTF1), bikunin, lysozyme, albumin, fetuin-A), and model compounds (a bikunin fragment, recombinant-, milk-, bone osteopontin, poly-L-aspartic acid (poly asp), poly-L-glutamic acid (poly glu)) in modulating precipitation reactions of kidney stone-related calcium oxalate mono- and dihydrates (COM, COD). Combining scanning confocal microscopy and fluorescence imaging, we determined the crystal faces of COM with which these polypeptides interact; using scanning electron microscopy, we characterized their effects on crystal habits and precipitated volumes. Our findings demonstrate that polypeptide adsorption to COM crystals is dictated first by the polypeptide's affinity for the crystal followed by its preference for a crystal face: basic and relatively hydrophobic macromolecules show no adsorption, while acidic and more hydrophilic polypeptides adsorb either nonspecifically to all faces of COM or preferentially to {100}/{121} edges and {100} faces. However, investigating calcium oxalates grown in the presence of these polypeptides showed that some acidic proteins that adsorb to crystals do not affect crystallization, even if present in excess of physiological concentrations. These proteins (albumin, bikunin, PTF1, recombinant OPN) have estimated total hydrophilicities from 200 to 850 kJ/mol and net negative charges from -9 to -35, perhaps representing a "window" in which proteins adsorb and coat urinary crystals (support of excretion) without affecting crystallization. Strongest effects on crystallization were observed for polypeptides that are either highly hydrophilic (>950 kJ/mol) and highly carboxylated (poly asp, poly glu), or else highly hydrophilic and highly phosphorylated (native OPN isoforms), suggesting that highly hydrophilic proteins strongly affect precipitation processes in the urinary tract. Therefore, the level of hydrophilicity and net charge is a critical factor in the ability of polypeptides to affect crystallization and to regulate biomineralization processes.

Selective lowering of synapsins induced by oligomeric α-synuclein exacerbates memory deficits.

PubMed

Larson, Megan E; Greimel, Susan J; Amar, Fatou; LaCroix, Michael; Boyle, Gabriel; Sherman, Mathew A; Schley, Hallie; Miel, Camille; Schneider, Julie A; Kayed, Rakez; Benfenati, Fabio; Lee, Michael K; Bennett, David A; Lesné, Sylvain E

2017-06-06

Mounting evidence indicates that soluble oligomeric forms of amyloid proteins linked to neurodegenerative disorders, such as amyloid-β (Aβ), tau, or α-synuclein (αSyn) might be the major deleterious species for neuronal function in these diseases. Here, we found an abnormal accumulation of oligomeric αSyn species in AD brains by custom ELISA, size-exclusion chromatography, and nondenaturing/denaturing immunoblotting techniques. Importantly, the abundance of αSyn oligomers in human brain tissue correlated with cognitive impairment and reductions in synapsin expression. By overexpressing WT human αSyn in an AD mouse model, we artificially enhanced αSyn oligomerization. These bigenic mice displayed exacerbated Aβ-induced cognitive deficits and a selective decrease in synapsins. Following isolation of various soluble αSyn assemblies from transgenic mice, we found that in vitro delivery of exogenous oligomeric αSyn but not monomeric αSyn was causing a lowering in synapsin-I/II protein abundance. For a particular αSyn oligomer, these changes were either dependent or independent on endogenous αSyn expression. Finally, at a molecular level, the expression of synapsin genes SYN1 and SYN2 was down-regulated in vivo and in vitro by αSyn oligomers, which decreased two transcription factors, cAMP response element binding and Nurr1, controlling synapsin gene promoter activity. Overall, our results demonstrate that endogenous αSyn oligomers can impair memory by selectively lowering synapsin expression.

Millimeter and submillimeter wave spectroscopy of propanal

NASA Astrophysics Data System (ADS)

Zingsheim, Oliver; Müller, Holger S. P.; Lewen, Frank; Jørgensen, Jes K.; Schlemmer, Stephan

2017-12-01

The rotational spectra of the two stable conformers syn- and gauche-propanal (CH3CH2CHO) were studied in the millimeter and submillimeter wave regions from 75 to 500 GHz with the Cologne (Sub-)Millimeter wave Spectrometer. Furthermore, the first excited states associated with the aldehyde torsion and with the methyl torsion, respectively, of the syn-conformer were analyzed. The newly obtained spectroscopic parameters yield better predictions, thus fulfill sensitivity and resolution requirements in new astronomical observations in order to unambiguously assign pure rotational transitions of propanal. This is demonstrated on a radio astronomical spectrum from the Atacama Large Millimeter/submillimeter Array Protostellar Interferometric Line Survey (ALMA-PILS). In particular, an accurate description of observed splittings, caused by internal rotation of the methyl group in the syn-conformer and by tunneling rotation interaction from two stable degenerate gauche-conformers, is reported. The rotational spectrum of propanal is of additional interest because of its two large amplitude motions pertaining to the methyl and the aldehyde group, respectively.

Quantitative assessments of the distinct contributions of polypeptide backbone amides versus sidechain groups to chain expansion via chemical denaturation

PubMed Central

Holehouse, Alex S.; Garai, Kanchan; Lyle, Nicholas; Vitalis, Andreas; Pappu, Rohit V.

2015-01-01

In aqueous solutions with high concentrations of chemical denaturants such as urea and guanidinium chloride (GdmCl) proteins expand to populate heterogeneous conformational ensembles. These denaturing environments are thought to be good solvents for generic protein sequences because properties of conformational distributions align with those of canonical random coils. Previous studies showed that water is a poor solvent for polypeptide backbones and therefore backbones form collapsed globular structures in aqueous solvents. Here, we ask if polypeptide backbones can intrinsically undergo the requisite chain expansion in aqueous solutions with high concentrations of urea and GdmCl. We answer this question using a combination of molecular dynamics simulations and fluorescence correlation spectroscopy. We find that the degree of backbone expansion is minimal in aqueous solutions with high concentrations denaturants. Instead, polypeptide backbones sample conformations that are denaturant-specific mixtures of coils and globules, with a persistent preference for globules. Therefore, typical denaturing environments cannot be classified as good solvents for polypeptide backbones. How then do generic protein sequences expand in denaturing environments? To answer this question, we investigated the effects of sidechains using simulations of two archetypal sequences with amino acid compositions that are mixtures of charged, hydrophobic, and polar groups. We find that sidechains lower the effective concentration of backbone amides in water leading to an intrinsic expansion of polypeptide backbones in the absence of denaturants. Additional dilution of the effective concentration of backbone amides is achieved through preferential interactions with denaturants. These effects lead to conformational statistics in denaturing environments that are congruent with those of canonical random coils. Our results highlight the role of sidechain-mediated interactions as determinants of the conformational properties of unfolded states in water and in influencing chain expansion upon denaturation. PMID:25664638

Anti-α-synuclein immunotherapy reduces α-synuclein propagation in the axon and degeneration in a combined viral vector and transgenic model of synucleinopathy.

PubMed

Spencer, Brian; Valera, Elvira; Rockenstein, Edward; Overk, Cassia; Mante, Michael; Adame, Anthony; Zago, Wagner; Seubert, Peter; Barbour, Robin; Schenk, Dale; Games, Dora; Rissman, Robert A; Masliah, Eliezer

2017-01-13

Neurodegenerative disorders such as Parkinson's Disease (PD), PD dementia (PDD) and Dementia with Lewy bodies (DLB) are characterized by progressive accumulation of α-synuclein (α-syn) in neurons. Recent studies have proposed that neuron-to-neuron propagation of α-syn plays a role in the pathogenesis of these disorders. We have previously shown that antibodies against the C-terminus of α-syn reduce the intra-neuronal accumulation of α-syn and related deficits in transgenic models of synucleinopathy, probably by abrogating the axonal transport and accumulation of α-syn in in vivo models. Here, we assessed the effect of passive immunization against α-syn in a new mouse model of axonal transport and accumulation of α-syn. For these purpose, non-transgenic, α-syn knock-out and mThy1-α-syn tg (line 61) mice received unilateral intra-cerebral injections with a lentiviral (LV)-α-syn vector construct followed by systemic administration of the monoclonal antibody 1H7 (recognizes amino acids 91-99) or control IgG for 3 months. Cerebral α-syn accumulation and axonopathy was assessed by immunohistochemistry and effects on behavior were assessed by Morris water maze. Unilateral LV-α-syn injection resulted in axonal propagation of α-syn in the contra-lateral site with subsequent behavioral deficits and axonal degeneration. Passive immunization with 1H7 antibody reduced the axonal accumulation of α-syn in the contra-lateral side and ameliorated the behavioral deficits. Together this study supports the notion that immunotherapy might improve the deficits in models of synucleinopathy by reducing the axonal propagation and accumulation of α-syn. This represents a potential new mode of action through which α-syn immunization might work.

A novel cold-inducible gene from Pak-choi (Brassica campestris ssp. chinensis), BcWRKY46, enhances the cold, salt and dehydration stress tolerance in transgenic tobacco.

PubMed

Wang, Feng; Hou, Xilin; Tang, Jun; Wang, Zhen; Wang, Shuming; Jiang, Fangling; Li, Ying

2012-04-01

WRKY TFs belong to one of the largest families of transcriptional regulators in plants and form integral parts of signaling webs that modulate many plant processes. BcWRKY46, a cDNA clone encoding a polypeptide of 284 amino acids and exhibited the structural features of group III of WRKY protein family, was isolated from the cold-treated leaves of Pak-choi (Brassica campestris ssp. chinensis Makino, syn. B. rapa ssp. chinensis) using the cDNA-AFLP technique. Expression of this gene was induced quickly and strongly in response to various environmental stresses, including low temperatures, ABA, salt and dehydration. Constitutive expression of BcWRKY46 in tobacco under the control of the CaMV35S promoter reduced the susceptibility of transgenic tobacco to freezing, ABA, salt and dehydration stresses. Our studies suggest that BcWRKY46 plays an important role in responding to ABA and abiotic stress.

Polypeptide composition of bacterial cyclic diguanylic acid-dependent cellulose synthase and the occurrence of immunologically crossreacting proteins in higher plants

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mayer, R.; Ross, P.; Weinhouse, H.

1991-06-15

To comprehend the catalytic and regulatory mechanism of the cyclic diguanylic acid (c-di-GMP)-dependent cellulose synthase of Acetobacter xylinum and its relatedness to similar enzymes in other organisms, the structure of this enzyme was analyzed at the polypeptide level. The enzyme, purified 350-fold by enzyme-product entrapment, contains three major peptides (90, 67, and 54 kDa), which, based on direct photoaffinity and immunochemical labeling and amino acid sequence analysis, are constituents of the native cellulose synthase. Labeling of purified synthase with either ({sup 32}P)c-di-GMP or ({alpha}-{sup 32}P)UDP-glucose indicates that activator- and substrate-specific binding sites are most closely associated with the 67- andmore » 54-kDa peptides, respectively, whereas marginal photolabeling is detected in the 90-k-Da peptide. However, antibodies raised against a protein derived from the cellulose synthase structural gene (bcsB) specifically label all three peptides. The authors suggest that the structurally related 67- and 54-kDa peptides are fragments proteolytically derived from the 90-kDa peptide encoded by bcsB. The anti-cellulose synthase antibodies crossreact with a similar set of peptides derived from other cellulose-producing microorganisms and plants such as Agrobacterium tumefaciens, Rhizobium leguminosarum, mung bean, peas, barley, and cotton. The occurrence of such cellulose synthase-like structures in plant species suggests that a common enzymatic mechanism for cellulose biogenesis is employed throughout nature.« less

Experimental and theoretical IR study of methyl thioglycolate, CH3OC(O)CH2SH, in different phases: Evidence of a dimer formation

NASA Astrophysics Data System (ADS)

Bava, Yanina B.; Tamone, Luciana M.; Juncal, Luciana C.; Seng, Samantha; Tobón, Yeny A.; Sobanska, Sophie; Picone, A. Lorena; Romano, Rosana M.

2017-07-01

The IR spectrum of methyl thioglycolate (MTG) was studied in three different phases, and interpreted with the aid of DFT calculations. The gas phase IR spectrum was explainable by the presence of the most stable conformer (syn-gauche-(-)gauche) only, while the IR spectrum of the liquid reveals strong intermolecular interactions, coincident with the formation of a dimeric form. The matrix-isolated spectra allow the identification of the second conformer (syn-gauche-gauche), in addition to the most stable form. The MTG dimer was also isolated by increasing the proportion of MTG in the matrix. The theoretical most stable structure of the dimer, which calculated IR spectrum agrees very well with the experimental one, is stabilized by a double interaction of the lone pair of the O atom of each of the Cdbnd O groups with the antibonding orbitals σ* (Ssbnd H).

Fibril growth and seeding capacity play key roles in α-synuclein-mediated apoptotic cell death

PubMed Central

Mahul-Mellier, A-L; Vercruysse, F; Maco, B; Ait-Bouziad, N; De Roo, M; Muller, D; Lashuel, H A

2015-01-01

The role of extracellular α-synuclein (α-syn) in the initiation and the spreading of neurodegeneration in Parkinson's disease (PD) has been studied extensively over the past 10 years. However, the nature of the α-syn toxic species and the molecular mechanisms by which they may contribute to neuronal cell loss remain controversial. In this study, we show that fully characterized recombinant monomeric, fibrillar or stabilized forms of oligomeric α-syn do not trigger significant cell death when added individually to neuroblastoma cell lines. However, a mixture of preformed fibrils (PFFs) with monomeric α-syn becomes toxic under conditions that promote their growth and amyloid formation. In hippocampal primary neurons and ex vivo hippocampal slice cultures, α-syn PFFs are capable of inducing a moderate toxicity over time that is greatly exacerbated upon promoting fibril growth by addition of monomeric α-syn. The causal relationship between α-syn aggregation and cellular toxicity was further investigated by assessing the effect of inhibiting fibrillization on α-syn-induced cell death. Remarkably, our data show that blocking fibril growth by treatment with known pharmacological inhibitor of α-syn fibrillization (Tolcapone) or replacing monomeric α-syn by monomeric β-synuclein in α-syn mixture composition prevent α-syn-induced toxicity in both neuroblastoma cell lines and hippocampal primary neurons. We demonstrate that exogenously added α-syn fibrils bind to the plasma membrane and serve as nucleation sites for the formation of α-syn fibrils and promote the accumulation and internalization of these aggregates that in turn activate both the extrinsic and intrinsic apoptotic cell death pathways in our cellular models. Our results support the hypothesis that ongoing aggregation and fibrillization of extracellular α-syn play central roles in α-syn extracellular toxicity, and suggest that inhibiting fibril growth and seeding capacity constitute a viable strategy for protecting against α-syn-induced toxicity and slowing the progression of neurodegeneration in PD and other synucleinopathies. PMID:26138444

Reinventing Cell Penetrating Peptides Using Glycosylated Methionine Sulfonium Ion Sequences.

PubMed

Kramer, Jessica R; Schmidt, Nathan W; Mayle, Kristine M; Kamei, Daniel T; Wong, Gerard C L; Deming, Timothy J

2015-05-27

Cell penetrating peptides (CPPs) are intriguing molecules that have received much attention, both in terms of mechanistic analysis and as transporters for intracellular therapeutic delivery. Most CPPs contain an abundance of cationic charged residues, typically arginine, where the amino acid compositions, rather than specific sequences, tend to determine their ability to enter cells. Hydrophobic residues are often added to cationic sequences to create efficient CPPs, but typically at the penalty of increased cytotoxicity. Here, we examined polypeptides containing glycosylated, cationic derivatives of methionine, where we found these hydrophilic polypeptides to be surprisingly effective as CPPs and to also possess low cytotoxicity. X-ray analysis of how these new polypeptides interact with lipid membranes revealed that the incorporation of sterically demanding hydrophilic cationic groups in polypeptides is an unprecedented new concept for design of potent CPPs.

Reinventing cell penetrating peptides using glycosylated methionine sulfonium ion sequences

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kramer, Jessica R.; Schmidt, Nathan W.; Mayle, Kristine M.

2015-04-15

Cell penetrating peptides (CPPs) are intriguing molecules that have received much attention, both in terms of mechanistic analysis and as transporters for intracellular therapeutic delivery. Most CPPs contain an abundance of cationic charged residues, typically arginine, where the amino acid compositions, rather than specific sequences, tend to determine their ability to enter cells. Hydrophobic residues are often added to cationic sequences to create efficient CPPs, but typically at the penalty of increased cytotoxicity. Here, we examined polypeptides containing glycosylated, cationic derivatives of methionine, where we found these hydrophilic polypeptides to be surprisingly effective as CPPs and to also possess lowmore » cytotoxicity. X-ray analysis of how these new polypeptides interact with lipid membranes revealed that the incorporation of sterically demanding hydrophilic cationic groups in polypeptides is an unprecedented new concept for design of potent CPPs.« less

Differential effects of immunotherapy with antibodies targeting α-synuclein oligomers and fibrils in a transgenic model of synucleinopathy.

PubMed

El-Agnaf, Omar; Overk, Cassia; Rockenstein, Edward; Mante, Michael; Florio, Jazmin; Adame, Anthony; Vaikath, Nishant; Majbour, Nour; Lee, Seung-Jae; Kim, Changyoun; Masliah, Eliezer; Rissman, Robert A

2017-08-01

Disorders with progressive accumulation of α-synuclein (α-syn) are a common cause of dementia and parkinsonism in the aging population. Accumulation and propagation of α-syn play a role in the pathogenesis of these disorders. Previous studies have shown that immunization with antibodies that recognize C-terminus of α-syn reduces the intra-neuronal accumulation of α-syn and related deficits in transgenic models of synucleinopathy. These studies employed antibodies that recognize epitopes within monomeric and aggregated α-syn that were generated through active immunization or administered via passive immunization. However, it is possible that more specific effects might be achieved with antibodies recognizing selective species of the α-syn aggregates. In this respect we recently developed antibodies that differentially recognized various oligomers (Syn-O1, -O2, and -O4) and fibrilar (Syn-F1 and -F2) forms of α-syn. For this purpose wild-type α-syn transgenic (line 61) mice were immunized with these 5 different antibodies and neuropathologically and biochemically analyzed to determine which was most effective at reducing α-syn accumulation and related deficits. We found that Syn-O1, -O4 and -F1 antibodies were most effective at reducing accumulation of α-syn oligomers in multiple brain regions and at preventing neurodegeneration. Together this study supports the notion that selective antibodies against α-syn might be suitable for development new treatments for synucleinopathies such as PD and DLB. Published by Elsevier Inc.

Differential effects of immunotherapy with antibodies targeting α-synuclein oligomers and fibrils in a transgenic model of synucleinopathy

PubMed Central

El-Agnaf, Omar; Overk, Cassia; Rockenstein, Edward; Mante, Michael; Florio, Jazmin; Adame, Anthony; Vaikath, Nishant; Majbour, Nour; Lee, Seung-Jae; Kim, Changyoun; Masliah, Eliezer; Rissman, Robert A.

2018-01-01

Disorders with progressive accumulation of α-synuclein (α-syn) are a common cause of dementia and parkinsonism in the aging population. Accumulation and propagation of α-syn play a role in the pathogenesis of these disorders. Previous studies have shown that immunization with antibodies that recognize C-terminus of α-syn reduces the intra-neuronal accumulation of α-syn and related deficits in transgenic models of synucleinopathy. These studies employed antibodies that recognize epitopes within monomeric and aggregated α-syn that were generated through active immunization or administered via passive immunization. However, it is possible that more specific effects might be achieved with antibodies recognizing selective species of the α-syn aggregates. In this respect we recently developed antibodies that differentially recognized various oligomers (Syn-O1, -O2, and -O4) and fibrilar (Syn-F1 and -F2) forms of α-syn. For this purpose wild-type α-syn transgenic (line 61) mice were immunized with these 5 different antibodies and neuropathologically and biochemically analyzed to determine which was most effective at reducing α-syn accumulation and related deficits. We found that Syn-O1, -O4 and -F1 antibodies were most effective at reducing accumulation of α-syn oligomers in multiple brain regions and at preventing neurodegeneration. Together this study supports the notion that selective antibodies against α-syn might be suitable for development new treatments for synucleinopathies such as PD and DLB. PMID:28476636

Different Conformations of 2'-Deoxycytidine in the Gas and Solid Phases: Competition between Intra- and Intermolecular Hydrogen Bonds.

PubMed

Ling, Sanliang; Gutowski, Maciej

2016-10-06

Computational results have been reported for 2'-deoxycytidine (dC), its gas phase isomers, tautomers, and their conformers, as well as for the crystalline phase. In addition to the neutral gas phase molecules, we have also considered associated radical anions and cations. The structural calculations were performed at the density functional and MP2 levels of theory. Vertical electron ionization energies and excess electron binding energies were determined using electron propagator theory. The α-anomer proved to be more stable by a fraction of kcal/mol than the biologically relevant canonical β-anomer. The conformational space of canonical dC has been systematically probed. dC in the crystalline phase or DNA structures favors canonical anti conformations. These structures were used in past computational studies to model gas phase characteristics of dC. Our findings indicate, however, that the gas phase dC favors syn conformations. It has repercussions for earlier interpretations of gas phase experimental results based on these computational results. The thermodynamic dominance of syn conformations results from the formation of an intramolecular O5'-H13···O2 hydrogen bond. The IR spectra of the most stable syn and anti canonical conformers differ markedly in the region of frequencies corresponding to NH/OH stretching modes. The MP2 value of deprotonation enthalpy of dC of 1411.7 kJ/mol is in very good agreement with the experimental value of 1409 ± 2.5 kJ/mol. The most stable valence anions are characterized by electron vertical detachment energies (VDE) in the 0.8-1.0 eV range, in good agreement with the experimental VDE of 0.87 eV. The barrier for the glycosidic bond cleavage is significant in the neutral canonical dC, 40.0 kcal/mol, and it is reduced to 22 and 16 kcal/mol for the anionic and cationic radicals of dC, respectively. The cleavage reaction is exothermic by 4 kcal/mol for dC - and endothermic by 7 and 9 kcal/mol for dC + and dC, respectively. We decomposed the crystal cohesive energy into repulsive one-body terms associated with the syn-anti conformational changes, and the attractive intermolecular interaction term. We exposed that the syn-anti conformational changes are very favorable for intermolecular interactions; in particular they make the imino-amino side of the cytosine residue accessible to intermolecular interactions.

Axonopathy in an α-Synuclein Transgenic Model of Lewy Body Disease Is Associated with Extensive Accumulation of C-Terminal–Truncated α-Synuclein

PubMed Central

Games, Dora; Seubert, Peter; Rockenstein, Edward; Patrick, Christina; Trejo, Margarita; Ubhi, Kiren; Ettle, Benjamin; Ghassemiam, Majid; Barbour, Robin; Schenk, Dale; Nuber, Silke; Masliah, Eliezer

2014-01-01

Progressive accumulation of α-synuclein (α-syn) in limbic and striatonigral systems is associated with the neurodegenerative processes in dementia with Lewy bodies (DLB) and Parkinson’s disease (PD). The murine Thy-1 (mThy1)-α-syn transgenic (tg) model recapitulates aspects of degenerative processes associated with α-syn accumulation in these disorders. Given that axonal and synaptic pathologies are important features of DLB and PD, we sought to investigate the extent and characteristics of these alterations in mThy1-α-syn tg mice and to determine the contribution of α-syn c-terminally cleaved at amino acid 122 (CT α-syn) to these abnormalities. We generated a novel polyclonal antibody (SYN105) against the c-terminally truncated sequence (amino acids 121 to 123) of α-syn (CT α-syn) and performed immunocytochemical and ultrastructural analyses in mThy1-α-syn tg mice. We found abundant clusters of dystrophic neurites in layers 2 to 3 of the neocortex, the stratum lacunosum, the dentate gyrus, and cornu ammonis 3 of the hippocampus, striatum, thalamus, midbrain, and pons. Dystrophic neurites displayed intense immunoreactivity detected with the SYN105 antibody. Double-labeling studies with antibodies to phosphorylated neurofilaments confirmed the axonal location of full-length and CT α-syn. α-Syn immunoreactive dystrophic neurites contained numerous electrodense laminated structures. These results show that neuritic dystrophy is a prominent pathologic feature of the mThy1-α-syn tg model and suggest that CT α-syn might play an important role in the process of axonal damage in these mice as well as in DLB and PD. PMID:23313024

Axonopathy in an α-synuclein transgenic model of Lewy body disease is associated with extensive accumulation of C-terminal-truncated α-synuclein.

PubMed

Games, Dora; Seubert, Peter; Rockenstein, Edward; Patrick, Christina; Trejo, Margarita; Ubhi, Kiren; Ettle, Benjamin; Ghassemiam, Majid; Barbour, Robin; Schenk, Dale; Nuber, Silke; Masliah, Eliezer

2013-03-01

Progressive accumulation of α-synuclein (α-syn) in limbic and striatonigral systems is associated with the neurodegenerative processes in dementia with Lewy bodies (DLB) and Parkinson's disease (PD). The murine Thy-1 (mThy1)-α-syn transgenic (tg) model recapitulates aspects of degenerative processes associated with α-syn accumulation in these disorders. Given that axonal and synaptic pathologies are important features of DLB and PD, we sought to investigate the extent and characteristics of these alterations in mThy1-α-syn tg mice and to determine the contribution of α-syn c-terminally cleaved at amino acid 122 (CT α-syn) to these abnormalities. We generated a novel polyclonal antibody (SYN105) against the c-terminally truncated sequence (amino acids 121 to 123) of α-syn (CT α-syn) and performed immunocytochemical and ultrastructural analyses in mThy1-α-syn tg mice. We found abundant clusters of dystrophic neurites in layers 2 to 3 of the neocortex, the stratum lacunosum, the dentate gyrus, and cornu ammonis 3 of the hippocampus, striatum, thalamus, midbrain, and pons. Dystrophic neurites displayed intense immunoreactivity detected with the SYN105 antibody. Double-labeling studies with antibodies to phosphorylated neurofilaments confirmed the axonal location of full-length and CT α-syn. α-Syn immunoreactive dystrophic neurites contained numerous electrodense laminated structures. These results show that neuritic dystrophy is a prominent pathologic feature of the mThy1-α-syn tg model and suggest that CT α-syn might play an important role in the process of axonal damage in these mice as well as in DLB and PD. Copyright © 2013 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

A New Method for Quantitative Immunoblotting of Endogenous α-Synuclein

PubMed Central

Newman, Andrew J.; Selkoe, Dennis; Dettmer, Ulf

2013-01-01

β-Sheet-rich aggregates of α-synuclein (αSyn) are the hallmark neuropathology of Parkinson’s disease and related synucleinopathies, whereas the principal native structure of αSyn in healthy cells - unfolded monomer or α-helically folded oligomer - is under debate. Our recent crosslinking analysis of αSyn in intact cells showed that a large portion of endogenous αSyn can be trapped as oligomers, most notably as apparent tetramers. One challenge in such studies is accurately quantifying αSyn Western blot signals among samples, as crosslinked αSyn trends toward increased immunoreactivity. Here, we analyzed this phenomenon in detail and found that treatment with the reducible amine-reactive crosslinker DSP strongly increased αSyn immunoreactivity even after cleavage with the reducing agent β-mercaptoethanol. The effect was observed with all αSyn antibodies tested and in all sample types from human brain homogenates to untransfected neuroblastoma cells, permitting easy detection of endogenous αSyn in the latter, which had long been considered impossible. Coomassie staining of blots before and after several hours of washing revealed complete retention of αSyn after DSP/β-mercaptoethanol treatment, in contrast to a marked loss of αSyn without this treatment. The treatment also enhanced immunodetection of the homologs β- and γ-synuclein and of histones, another group of small, lysine-rich proteins. We conclude that by neutralizing positive charges and increasing protein hydrophobicity, amine crosslinker treatment promotes adhesion of αSyn to blotting membranes. These data help explain the recent report of fixing αSyn blots with paraformaldehyde after transfer, which we find produces similar but weaker effects. DSP/β-mercaptoethanol treatment of Western blots should be particularly useful to quantify low-abundance αSyn forms such as extracellular and post-translationally modified αSyn and splice variants. PMID:24278419

Structural characterization of the N-terminal mineral modification domains from the molluscan crystal-modulating biomineralization proteins, AP7 and AP24.

PubMed

Wustman, Brandon A; Morse, Daniel E; Evans, John Spencer

2004-08-05

The AP7 and AP24 proteins represent a class of mineral-interaction polypeptides that are found in the aragonite-containing nacre layer of mollusk shell (H. rufescens). These proteins have been shown to preferentially interfere with calcium carbonate mineral growth in vitro. It is believed that both proteins play an important role in aragonite polymorph selection in the mollusk shell. Previously, we demonstrated the 1-30 amino acid (AA) N-terminal sequences of AP7 and AP24 represent mineral interaction/modification domains in both proteins, as evidenced by their ability to frustrate calcium carbonate crystal growth at step edge regions. In this present report, using free N-terminal, C(alpha)-amide "capped" synthetic polypeptides representing the 1-30 AA regions of AP7 (AP7-1 polypeptide) and AP24 (AP24-1 polypeptide) and NMR spectroscopy, we confirm that both N-terminal sequences possess putative Ca (II) interaction polyanionic sequence regions (2 x -DD- in AP7-1, -DDDED- in AP24-1) that are random coil-like in structure. However, with regard to the remaining sequences regions, each polypeptide features unique structural differences. AP7-1 possesses an extended beta-strand or polyproline type II-like structure within the A11-M10, S12-V13, and S28-I27 sequence regions, with the remaining sequence regions adopting a random-coil-like structure, a trait common to other polyelectrolyte mineral-associated polypeptide sequences. Conversely, AP24-1 possesses random coil-like structure within A1-S9 and Q14-N16 sequence regions, and evidence for turn-like, bend, or loop conformation within the G10-N13, Q17-N24, and M29-F30 sequence regions, similar to the structures identified within the putative elastomeric proteins Lustrin A and sea urchin spicule matrix proteins. The similarities and differences in AP7 and AP24 N-terminal domain structure are discussed with regard to joint AP7-AP24 protein modification of calcium carbonate growth. Copyright 2004 Wiley Periodicals, Inc.

Architecture effects on multivalent interactions by polypeptide-based multivalent ligands

NASA Astrophysics Data System (ADS)

Liu, Shuang

Multivalent interactions are characterized by the simultaneous binding between multiple ligands and multiple binding sites, either in solutions or at interfaces. In biological systems, most multivalent interactions occur between protein receptors and carbohydrate ligands through hydrogen-bonding and hydrophobic interactions. Compared with weak affinity binding between one ligand and one binding site, i.e. monovalent interaction, multivalent interactioins provide greater avidity and specificity, and therefore play unique roles in a broad range of biological activities. Moreover, the studies of multivalent interactions are also essential for producing effective inhibitors and effectors of biological processes that could have important therapeutic applications. Synthetic multivalent ligands have been designed to mimic the biological functions of natural multivalent interactions, and various types of scaffolds have been used to display multiple ligands, including small molecules, linear polymers, dendrimers, nanoparticle surfaces, monolayer surfaces and liposomes. Studies have shown that multivalent interactions can be highly affected by various architectural parameters of these multivalent ligands, including ligand identities, valencies, spacing, ligand densities, nature of linker arms, scaffold length and scaffold conformation. Most of these multivalent ligands are chemically synthesized and have limitations of controlling over sequence and conformation, which is a barrier for mimicking ordered and controlled natural biological systems. Therefore, multivalent ligands with precisely controlled architecture are required for improved structure-function relationship studies. Protein engineering methods with subsequent chemical coupling of ligands provide significant advantages of controlling over backbone conformation and functional group placement, and therefore have been used to synthesize recombinant protein-based materials with desired properties similar to natural protein materials, including structural as well as functional proteins. Therefore, polypeptide-based multivalent scaffolds are used to display ligands to assess the contribution of different architectural parameters to the multivalent binding events. In this work, a family of alanine-rich alpha-helical glycopolypeptides was designed and synthesized by a combination of protein engineering and chemical coupling, to display two types of saccharide ligands for two different multivalent binding systems. The valencies, chain length and spacing between adjacent ligands of these multivalent ligands were designed in order to study architecture effects on multivalent interactions. The polypeptides and their glycoconjugates were characterized via various methods, including SDS-PAGE, NMR, HPLC, amino acid analysis (AAA), MALDI, circular dichroism (CD) and GPC. In the first multivalent binding system, cholera toxin B pentamer (CT B5) was chosen to be the protein receptor due to its well-characterized structure, lack of significant steric interference of binding to multiple binding sites, and requirement of only simple monosaccharide as ligands. Galactopyranoside was incorporated into polypeptide scaffolds through amine-carboxylic acid coupling to the side chains of glutamic acid residues. The inhibition and binding to CT B5 of these glycopolypeptide ligands were evaluated by direct enzyme-linked assay (DELA). As a complement method, weak affinity chromatography (WAC) was also used to evaluate glycopolypeptides binding to a CT B5 immobilized column. The architecture effects on CT B 5 inhibition are discussed. In the second system, cell surface receptor L-selectin was targeted by polypeptide-based multivalent ligands containing disulfated galactopyranoside ligands, due to its important roles in various immunological activities. The effects of glycopolypeptide architectural variables L-selectin shedding were evaluated via ELISA-based assays. These polypeptide-based multivalent ligands are suggested to be useful for elucidating architecture effects on multivalent interactions, manipulating multivalent interactions and the subsequent cellular responses in different systems. These materials have great potential applications in therapeutics and could also provide guidelines for design of multivalent ligands for other protein receptors.

New Kunitz-Type HCRG Polypeptides from the Sea Anemone Heteractis crispa

PubMed Central

Gladkikh, Irina; Monastyrnaya, Margarita; Zelepuga, Elena; Sintsova, Oksana; Tabakmakher, Valentin; Gnedenko, Oksana; Ivanov, Alexis; Hua, Kuo-Feng; Kozlovskaya, Emma

2015-01-01

Sea anemones are a rich source of Kunitz-type polypeptides that possess not only protease inhibitor activity, but also Kv channels toxicity, analgesic, antihistamine, and anti-inflammatory activities. Two Kunitz-type inhibitors belonging to a new Heteractis crispa RG (HCRG) polypeptide subfamily have been isolated from the sea anemone Heteractis crispa. The amino acid sequences of HCRG1 and HCRG2 identified using the Edman degradation method share up to 95% of their identity with the representatives of the HCGS polypeptide multigene subfamily derived from H. crispa cDNA. Polypeptides are characterized by positively charged Arg at the N-terminus as well as P1 Lys residue at their canonical binding loop, identical to those of bovine pancreatic trypsin inhibitor (BPTI). These polypeptides are shown by our current evidence to be more potent inhibitors of trypsin than the known representatives of the HCGS subfamily with P1Thr. The kinetic and thermodynamic characteristics of the intermolecular interactions between inhibitors and serine proteases were determined by the surface plasmon resonance (SPR) method. Residues functionally important for polypeptide binding to trypsin were revealed using molecular modeling methods. Furthermore, HCRG1 and HCRG2 possess anti-inflammatory activity, reducing tumor necrosis factor-α (TNF-α) and interleukin 6 (IL-6) secretions, as well as proIL-1β expression in lipopolysaccharide (LPS)-activated macrophages. However, there was no effect on nitric oxide (NO) generation. PMID:26404319

Symptoms of anxiety and mood disturbance alter cardiac and peripheral autonomic control in patients with metabolic syndrome.

PubMed

Toschi-Dias, Edgar; Trombetta, Ivani C; da Silva, Valdo José Dias; Maki-Nunes, Cristiane; Alves, Maria Janieire N N; Angelo, Luciana F; Cepeda, Felipe X; Martinez, Daniel G; Negrão, Carlos Eduardo; Rondon, Maria Urbana P B

2013-03-01

Previous investigations show that metabolic syndrome (MetSyn) causes sympathetic hyperactivation. Symptoms of anxiety and mood disturbance (AMd) provoke sympatho-vagal imbalance. We hypothesized that AMd would alter even further the autonomic function in patients with MetSyn. Twenty-six never-treated patients with MetSyn (ATP-III) were allocated to two groups, according to the levels of anxiety and mood disturbance: (1) with AMd (MetSyn + AMd, n = 15), and (2) without AMd (MetSyn, n = 11). Ten healthy control subjects were also studied (C, n = 10). AMd was determined using quantitative questionnaires. Muscle sympathetic nerve activity (MSNA, microneurography), blood pressure (oscillometric beat-to-beat basis), and heart rate (ECG) were measured during a baseline 10-min period. Spectral analysis of RR interval and systolic arterial pressure were analyzed, and the power of low (LF) and high (HF) frequency bands were determined. Sympatho-vagal balance was obtained by LF/HF ratio. Spontaneous baroreflex sensitivity (BRS) was evaluated by calculation of α-index. MSNA was greater in patients with MetSyn + AMd compared with MetSyn and C. Patients with MetSyn + AMd showed higher LF and lower HF power compared with MetSyn and C. In addition, LF/HF balance was higher in MetSyn + AMd than in MetSyn and C groups. BRS was decreased in MetSyn + AMd compared with MetSyn and C groups. Anxiety and mood disturbance alter autonomic function in patients with MetSyn. This autonomic dysfunction may contribute to the increased cardiovascular risk observed in patients with mood alterations.

An allosterically regulated reversible mechanical molecular switch: A de novo protein maquette functions as a redox/ionic strength sensor coupling chemical binding energy or charge interactions to conformational change

NASA Astrophysics Data System (ADS)

Grosset, Anne Marie

2000-10-01

Switch-like structural rearrangements of subunits due to charge-interactions are common in the basic biological action of proteins that couple and transfer chemical and ionic signals, sensing and regulation, mechanical force and electrochemical free energy. A simple synthetic protein model (maquette) has been designed to better understand the engineering of natural switches. Basic thermodynamic principles define the two key elements required for biological or chemical function of a switch. First, there must be two well-defined states. In this case, the two conformational states must have an energetic difference (DeltaDeltaG°) that is spanned by the applied driving force. Second, there must be an external stimulus, which preferentially interacts with one of the two states. The external stimulus provides the driving force that shifts the equilibrium from the first state to the second state (≥10:1 shifting towards ≤1:10). The energetic difference between the states must be the same order of magnitude as the driving force. In this synthetic protein, the two conformational states correspond to parallel (syn) and antiparallel (anti) assembly of the two identical helix-ss-helix subunits that bind heme close to the di-sulfide loop region. Charge interactions between two ferric hemes bound to histidines provide a driving force on the order of 2 kcal/mol (corresponding in the syn-topology to the 75--100 mV split in the heme redox potentials, or the 25--80 times weaker binding for the second ferric heme). The tetra-alpha-helix bundle has been modified to have a DeltaG around 1.8--2.5 kcal/mol (a 50--80 fold difference in the anti/syn ratio). Therefore, oxidation and reduction of the heme, or the binding of a second charged ferric heme can reversibly switch between syn- and anti-topologies, providing a sensitive detector of redox state or heme concentration. External solution conditions (e.g. ionic composition) can act on the protein remotely from the primary internal switch action and confer a secondary level of allosteric regulation. Bifunctional ligands can link subunits to shift topology. Scanning redox potentiometry can monitor the kinetics of topological change. Point amino acid substitutions and computer repacking of the hydrophobic core can modulate both the kinetics and the energetics.

Reducing C-Terminal-Truncated Alpha-Synuclein by Immunotherapy Attenuates Neurodegeneration and Propagation in Parkinson's Disease-Like Models

PubMed Central

Games, Dora; Valera, Elvira; Spencer, Brian; Rockenstein, Edward; Mante, Michael; Adame, Anthony; Patrick, Christina; Ubhi, Kiren; Nuber, Silke; Sacayon, Patricia; Zago, Wagner; Seubert, Peter; Barbour, Robin; Schenk, Dale

2014-01-01

Parkinson's disease (PD) and dementia with Lewy bodies (DLB) are common neurodegenerative disorders of the aging population, characterized by progressive and abnormal accumulation of α-synuclein (α-syn). Recent studies have shown that C-terminus (CT) truncation and propagation of α-syn play a role in the pathogenesis of PD/DLB. Therefore, we explored the effect of passive immunization against the CT of α-syn in the mThy1-α-syn transgenic (tg) mouse model, which resembles the striato-nigral and motor deficits of PD. Mice were immunized with the new monoclonal antibodies 1H7, 5C1, or 5D12, all directed against the CT of α-syn. CT α-syn antibodies attenuated synaptic and axonal pathology, reduced the accumulation of CT-truncated α-syn (CT-α-syn) in axons, rescued the loss of tyrosine hydroxylase fibers in striatum, and improved motor and memory deficits. Among them, 1H7 and 5C1 were most effective at decreasing levels of CT-α-syn and higher-molecular-weight aggregates. Furthermore, in vitro studies showed that preincubation of recombinant α-syn with 1H7 and 5C1 prevented CT cleavage of α-syn. In a cell-based system, CT antibodies reduced cell-to-cell propagation of full-length α-syn, but not of the CT-α-syn that lacked the 118–126 aa recognition site needed for antibody binding. Furthermore, the results obtained after lentiviral expression of α-syn suggest that antibodies might be blocking the extracellular truncation of α-syn by calpain-1. Together, these results demonstrate that antibodies against the CT of α-syn reduce levels of CT-truncated fragments of the protein and its propagation, thus ameliorating PD-like pathology and improving behavioral and motor functions in a mouse model of this disease. PMID:25009275

Reducing C-terminal-truncated alpha-synuclein by immunotherapy attenuates neurodegeneration and propagation in Parkinson's disease-like models.

PubMed

Games, Dora; Valera, Elvira; Spencer, Brian; Rockenstein, Edward; Mante, Michael; Adame, Anthony; Patrick, Christina; Ubhi, Kiren; Nuber, Silke; Sacayon, Patricia; Zago, Wagner; Seubert, Peter; Barbour, Robin; Schenk, Dale; Masliah, Eliezer

2014-07-09

Parkinson's disease (PD) and dementia with Lewy bodies (DLB) are common neurodegenerative disorders of the aging population, characterized by progressive and abnormal accumulation of α-synuclein (α-syn). Recent studies have shown that C-terminus (CT) truncation and propagation of α-syn play a role in the pathogenesis of PD/DLB. Therefore, we explored the effect of passive immunization against the CT of α-syn in the mThy1-α-syn transgenic (tg) mouse model, which resembles the striato-nigral and motor deficits of PD. Mice were immunized with the new monoclonal antibodies 1H7, 5C1, or 5D12, all directed against the CT of α-syn. CT α-syn antibodies attenuated synaptic and axonal pathology, reduced the accumulation of CT-truncated α-syn (CT-α-syn) in axons, rescued the loss of tyrosine hydroxylase fibers in striatum, and improved motor and memory deficits. Among them, 1H7 and 5C1 were most effective at decreasing levels of CT-α-syn and higher-molecular-weight aggregates. Furthermore, in vitro studies showed that preincubation of recombinant α-syn with 1H7 and 5C1 prevented CT cleavage of α-syn. In a cell-based system, CT antibodies reduced cell-to-cell propagation of full-length α-syn, but not of the CT-α-syn that lacked the 118-126 aa recognition site needed for antibody binding. Furthermore, the results obtained after lentiviral expression of α-syn suggest that antibodies might be blocking the extracellular truncation of α-syn by calpain-1. Together, these results demonstrate that antibodies against the CT of α-syn reduce levels of CT-truncated fragments of the protein and its propagation, thus ameliorating PD-like pathology and improving behavioral and motor functions in a mouse model of this disease. Copyright © 2014 the authors 0270-6474/14/349441-14$15.00/0.

A Plant Small Polypeptide Is a Novel Component of DNA-Binding Protein Phosphatase 1-Mediated Resistance to Plum pox virus in Arabidopsis1[C][W

PubMed Central

Castelló, María José; Carrasco, Jose Luis; Navarrete-Gómez, Marisa; Daniel, Jacques; Granot, David; Vera, Pablo

2011-01-01

DNA-binding protein phosphatases (DBPs) have been identified as a novel class of plant-specific regulatory factors playing a role in plant-virus interactions. NtDBP1 from tobacco (Nicotiana tabacum) was shown to participate in transcriptional regulation of gene expression in response to virus infection in compatible interactions, and AtDBP1, its closest relative in the model plant Arabidopsis (Arabidopsis thaliana), has recently been found to mediate susceptibility to potyvirus, one of the most speciose taxa of plant viruses. Here, we report on the identification of a novel family of highly conserved small polypeptides that interact with DBP1 proteins both in tobacco and Arabidopsis, which we have designated DBP-interacting protein 2 (DIP2). The interaction of AtDIP2 with AtDBP1 was demonstrated in vivo by bimolecular fluorescence complementation, and AtDIP2 was shown to functionally interfere with AtDBP1 in yeast. Furthermore, reducing AtDIP2 gene expression leads to increased susceptibility to the potyvirus Plum pox virus and to a lesser extent also to Turnip mosaic virus, whereas overexpression results in enhanced resistance. Therefore, we describe a novel family of conserved small polypeptides in plants and identify AtDIP2 as a novel host factor contributing to resistance to potyvirus in Arabidopsis. PMID:22021419

Synapsin I and Synapsin II regulate neurogenesis in the dentate gyrus of adult mice.

PubMed

Barbieri, Raffaella; Contestabile, Andrea; Ciardo, Maria Grazia; Forte, Nicola; Marte, Antonella; Baldelli, Pietro; Benfenati, Fabio; Onofri, Franco

2018-04-10

Adult neurogenesis is emerging as an important player in brain functions and homeostasis, while impaired or altered adult neurogenesis has been associated with a number of neuropsychiatric diseases, such as depression and epilepsy. Here we investigated the possibility that synapsins (Syns) I and II, beyond their known functions in developing and mature neurons, also play a role in adult neurogenesis. We performed a systematic evaluation of the distinct stages of neurogenesis in the hippocampal dentate gyrus of Syn I and Syn II knockout (KO) mice, before (2-months-old) and after (6-months-old) the appearance of the epileptic phenotype. We found that Syns I and II play an important role in the regulation of adult neurogenesis. In juvenile mice, Syn II deletion was associated with a specific decrease in the proliferation of neuronal progenitors, whereas Syn I deletion impaired the survival of newborn neurons. These defects were reverted after the appearance of the epileptic phenotype, with Syn I KO and Syn II KO mice exhibiting significant increases in survival and proliferation, respectively. Interestingly, long-term potentiation dependent on newborn neurons was present in both juvenile Syn mutants while, at later ages, it was only preserved in Syn II KO mice that also displayed an increased expression of brain-derived neurotrophic factor. This study suggests that Syns I and II play a role in adult neurogenesis and the defects in neurogenesis associated with Syn deletion may contribute to the alterations of cognitive functions observed in Syn-deficient mice.

Antiapoptotic property of human alpha-synuclein in neuronal cell lines is associated with the inhibition of caspase-3 but not caspase-9 activity.

PubMed

Li, Wenxue; Lee, Michael K

2005-06-01

Abnormalities of alpha-synuclein (alpha-Syn) are mechanistically linked to Parkinson's disease (PD) and other alpha-synucleinopathies. To gain additional insights into the relationships between alpha-Syn expression and cell death, we examined the effects of expressing human alpha-Syn (Hualpha-Syn) variants on the cellular vulnerability to apoptotic stimuli. We show that the expression of wild-type (WT) and A30P mutant, but not A53T mutant, Hualpha-Syn leads to the protection of neuronal cell lines from apoptosis but not necrosis. Significantly, Hualpha-Syn did not protect non-neuronal cell lines from apoptosis. We also show that A53T mutant is a loss of function in regards to the antiapoptotic property since the expression of WT Hualpha-Syn with an excess of A53T mutant Hualpha-Syn leads to protection of the cells from apoptosis. The antiapoptotic property is specific to human alpha-Syn as neither beta-Syn nor mouse alpha-Syn protected cells from apoptosis, and the carboxy-terminal 20 amino acids are required for the antiapoptotic property. Analyses of capase-3 and caspase-9 activation reveal that the antiapoptotic property of Hualpha-Syn in neuronal cell lines is associated with the attenuation of caspase-3 activity without affecting the caspase-9 activity or the levels of cleaved, active caspase-3. We conclude that Hualpha-Syn modulates the activity of cleaved caspase-3 product in neuronal cell lines.

Reducing C-terminal truncation mitigates synucleinopathy and neurodegeneration in a transgenic model of multiple system atrophy

PubMed Central

Bassil, Fares; Fernagut, Pierre-Olivier; Bezard, Erwan; Pruvost, Alain; Leste-Lasserre, Thierry; Hoang, Quyen Q.; Ringe, Dagmar; Petsko, Gregory A.; Meissner, Wassilios G.

2016-01-01

Multiple system atrophy (MSA) is a sporadic orphan neurodegenerative disorder. No treatment is currently available to slow down the aggressive neurodegenerative process, and patients die within a few years after disease onset. The cytopathological hallmark of MSA is the accumulation of alpha-synuclein (α-syn) aggregates in affected oligodendrocytes. Several studies point to α-syn oligomerization and aggregation as a mediator of neurotoxicity in synucleinopathies including MSA. C-terminal truncation by the inflammatory protease caspase-1 has recently been implicated in the mechanisms that promote aggregation of α-syn in vitro and in neuronal cell models of α-syn toxicity. We present here an in vivo proof of concept of the ability of the caspase-1 inhibitor prodrug VX-765 to mitigate α-syn pathology and to mediate neuroprotection in proteolipid protein α-syn (PLP-SYN) mice, a transgenic mouse model of MSA. PLP-SYN and age-matched wild-type mice were treated for a period of 11 wk with VX-765 or placebo. VX-765 prevented motor deficits in PLP-SYN mice compared with placebo controls. More importantly, VX-765 was able to limit the progressive toxicity of α-syn aggregation by reducing its load in the striatum of PLP-SYN mice. Not only did VX-765 reduce truncated α-syn, but it also decreased its monomeric and oligomeric forms. Finally, VX-765 showed neuroprotective effects by preserving tyrosine hydroxylase-positive neurons in the substantia nigra of PLP-SYN mice. In conclusion, our results suggest that VX-765, a drug that was well tolerated in a 6 wk-long phase II trial in patients with epilepsy, is a promising candidate to achieve disease modification in synucleinopathies by limiting α-syn accumulation. PMID:27482103

Using porphyrin-amino acid pairs to model the electrochemistry of heme proteins: experimental and theoretical investigations.

PubMed

Samajdar, Rudra N; Manogaran, Dhivya; Yashonath, S; Bhattacharyya, Aninda J

2018-04-18

Quasi reversibility in electrochemical cycling between different oxidation states of iron is an often seen characteristic of iron containing heme proteins that bind dioxygen. Surprisingly, the system becomes fully reversible in the bare iron-porphyrin complex: hemin. This leads to the speculation that the polypeptide bulk (globin) around the iron-porphyrin active site in these heme proteins is probably responsible for the electrochemical quasi reversibility. To understand the effect of such polypeptide bulk on iron-porphyrin, we study the interaction of specific amino acids with the hemin center in solution. We choose three representative amino acids-histidine (a well-known iron coordinator in bio-inorganic systems), tryptophan (a well-known fluoroprobe for proteins), and cysteine (a redox-active organic molecule). The interactions of these amino acids with hemin are studied using electrochemistry, spectroscopy, and density functional theory. The results indicate that among these three, the interaction of histidine with the iron center is strongest. Further, histidine maintains the electrochemical reversibility of iron. On the other hand, tryptophan and cysteine interact weakly with the iron center but disturb the electrochemical reversibility by contributing their own redox active processes to the system. Put together, this study attempts to understand the molecular interactions that can control electrochemical reversibility in heme proteins. The results obtained here from the three representative amino acids can be scaled up to build a heme-amino acid interaction database that may predict the electrochemical properties of any protein with a defined polypeptide sequence.

Lewy Body-like α-Synuclein Aggregates Resist Degradation and Impair Macroautophagy*♦

PubMed Central

Tanik, Selcuk A.; Schultheiss, Christine E.; Volpicelli-Daley, Laura A.; Brunden, Kurt R.; Lee, Virginia M. Y.

2013-01-01

Cytoplasmic α-synuclein (α-syn) aggregates, referred to as Lewy bodies, are pathological hallmarks of a number of neurodegenerative diseases, most notably Parkinson disease. Activation of macroautophagy is suggested to facilitate degradation of certain proteinaceous inclusions, but it is unclear if this pathway is capable of degrading α-syn aggregates. Here, we examined this issue by utilizing cellular models in which intracellular Lewy body-like α-syn inclusions accumulate after internalization of pre-formed α-syn fibrils into α-syn-expressing HEK293 cells or cultured primary neurons. We demonstrate that α-syn inclusions cannot be effectively degraded, even though they co-localize with essential components of both the autophagic and proteasomal protein degradation pathways. The α-syn aggregates persist even after soluble α-syn levels have been substantially reduced, suggesting that once formed, the α-syn inclusions are refractory to clearance. Importantly, we also find that α-syn aggregates impair overall macroautophagy by reducing autophagosome clearance, which may contribute to the increased cell death that is observed in aggregate-bearing cells. PMID:23532841

Millimeter-wave spectroscopy of syn formyl azide (HC(O)N3) in seven vibrational states

NASA Astrophysics Data System (ADS)

Walters, Nicholas A.; Amberger, Brent K.; Esselman, Brian J.; Woods, R. Claude; McMahon, Robert J.

2017-01-01

Millimeter-wave spectra for formyl azide (HC(O)N3) were obtained from 240 to 360 GHz at ambient temperature. For the ground state of syn formyl azide, over 1500 independent rotational transitions were measured and least-squares fit to a complete S-reduced 8th order centrifugal distortion/rigid rotor Hamiltonian. The decomposition of formyl azide was monitored over a period of several hours, the half-life (t½ = 30 min) was determined, and its decomposition products were investigated. Transitions from five vibrational satellites of syn formyl azide (ν9, ν12, 2ν9, ν9 + ν12, and ν11) were observed, measured, and least-squares fit to complete or nearly complete octic centrifugally-distorted, single-state S-reduced models. A less complete single-state fit of 3ν9 (509.3 cm-1) was obtained from an unperturbed subset of its assignable transitions. This state is apparently coupled to the fundamental ν8 (489.4 cm-1) and the overtone 2ν12 (503.6 cm-1), but the coupling remains unanalyzed. Anharmonic CCSD(T)/ANO1 estimates of the vibrational frequencies of syn formyl azide were in close agreement with previously published experimental and computational values. Experimentally determined vibration-rotation interaction (αi) values were in excellent agreement with coupled-cluster predicted αi values for the fundamentals ν9, ν12, and ν11.

Soft-sediment deformation structures from an ice-marginal storm-tide interactive system, Permo-Carboniferous Talchir Formation, Talchir Coalbasin, India

NASA Astrophysics Data System (ADS)

Bhattacharya, H. N.; Bhattacharya, Biplab

2010-01-01

Permo-Carboniferous Talchir Formation, Talchir Coalbasin, India, records sedimentation during a phase of climatic amelioration in an ice-marginal storm-affected shelf. Evidences of subtidal processes are preserved only under thick mud drapes deposited during waning storm phases. Various soft-sediment deformation structures in some sandstone/siltstone-mudstone interbeds, like syn-sedimentary faults, deformed laminations, sand-silt flows, convolute laminations and various flame structures, suggest liquefaction and fluidization of the beds due to passage of syn-depositional seismic shocks. In the Late Paleozoic ice-marginal shelf, such earthquake tremors could be generated by crustal movements in response to glacioisostatic adjustments of the basin floor.

Nomenclatural Studies Toward a World List of Diptera Genus-Group Names. Part V: Pierre-Justin-Marie Macquart.

PubMed

Evenhuis, Neal L; Pape, Thomas; Pont, Adrian C

2016-09-30

The Diptera genus-group names of Pierre-Justin-Marie Macquart are reviewed and annotated. A total of 399 available genus-group names in 69 families of Diptera are listed alphabetically, for each name giving author, year and page of original publication, originally included species, type species and method of fixation, current status of the name, family placement, and a list of any emendations of it that have been found in the literature. Remarks are given to clarify nomenclatural or taxonomic information. In addition, an index to all the species-group names of Diptera proposed by Macquart (3,611, of which 3,543 are available) is given with bibliographic reference (year and page) to each original citation.        The following type species are designated herein: Agculocera nigra Macquart, 1855 for Onuxicera Macquart, 1855, present designation [Tachinidae]; Trixa imhoffi Macquart, 1834, for Semiomyia Macquart, 1848, present designation [Tachinidae].        The following type species are designated herein with fixation under ICZN Code Art. 70.3.2: Azelia nebulosa Robineau-Desvoidy, 1830 for Atomogaster Macquart, 1835, present designation [Muscidae]; Tachydromia vocatoria Fallén, 1816 for Chelipoda Macquart, 1835, present designation [Empididae]; Eriocera macquarti Enderlein, 1912 for Eriocera Macquart, 1838, present designation [Limoniidae]; Limosina acutangula Zetterstedt, 1847 for Heteroptera Macquart, 1835, present designation [Sphaeroceridae]; Phryxe pavoniae Robineau-Desvoidy, 1830 for Masicera Macquart, 1834, present designation [Tachinidae]; Pachymyia macquartii Townsend, 1916 for Pachymyia Macquart, 1844, present designation [Tachinidae].        Earlier valid subsequent type-species designations have been found in this study for the following: Anisophysa Macquart, 1835 [Sepsidae]; Diphysa Macquart, 1838 [Stratiomyidae]; Pachyrhina Macquart, 1834 [Tipulidae]; Silbomyia Macquart, 1844 [Calliphoridae].        One name is raised from synonymy: Czernyola Bezzi, 1907, n. stat. [Clusiidae].        Names previously treated as available but found in this work to be unavailable include the following: Genus-group names-Anodontina Macquart, 1838, n. stat. [Empididae]; Athricia Macquart, 1834, n. stat. [Tachinidae]; Blepharis Macquart, 1838, n. stat. [Asilidae]; Dichelocera Enderlein, 1922, n. stat. [Tabanidae]; Lepidoselaga Loew, 1869, n. stat. [Tabanidae]; Lemptopeza Macquart, 1828, n. stat. [Hybotidae]; Microphora Zetterstedt, 1842, n. stat. [Dolichopodidae]; Microphorus Macquart, 1834, n. stat. [Dolichopodidae]; Plagiocephala Macquart, 1844, n. stat. [Ulidiidae]; Stratiomyia Macquart, 1838, n. stat. [Stratiomyidae]; Taenioptera Agassiz, 1846, n. stat. [Micropezidae]; Tapigaster Bezzi, 1923, n. stat. [Heleomyzidae]; Trizota Macquart, 1829, n. stat. [Syrphidae]. Species-group names-Microstylum sinense Macquart, 1838, n. stat. [Asilidae].        Corrected or clarified included species and/or corrected or clarified type-species and methods of typification are given for: Anabarhynchus Macquart, 1848 [Therevidae]; Anacanthella Macquart, 1855 [Stratiomyidae]; Apeilesis Macquart, 1846 [Tipulidae]; Aplomera Macquart, 1838 [Empididae]; Aprotheca Macquart, 1851 [Tachinidae]; Ardoptera Macquart, 1828 [Empididae]; Blepharella Macquart, 1851 [Tachinidae]; Brachystylum Macquart, 1855 [Tachinidae]; Cadicera Macquart, 1855 [Tabanidae]; Calobatemyia Macquart, 1855 [Calliphoridae]; Catapicephala Macquart, 1851 [Calliphoridae]; Ceroptera Macquart, 1835 [Sphaeroceridae]; Cheligaster Macquart, 1835 [Sepsidae]; Chetogaster Macquart, 1851 [Tachinidae]; Chlorogaster Macquart, 1851 [Tachinidae]; Cleitamia Macquart, 1835 [Platystomatidae]; Craspedia Macquart, 1838 [Asilidae]; Craspedochoeta Macquart, 1851 [Anthomyiidae]; Crumomyia Macquart, 1835 [Sphaeroceridae]; Dasyomma Macquart, 1840 [Athericidae]; Demoticus Macquart, 1854 [Tachinidae]; Epicerella Macquart, 1851 [Pyrgotidae]; Epicerina Macquart, 1850 [Acroceridae]; Euprosopia Macquart, 1847 [Platystomatidae]; Grapholostylum Macquart, 1851 [Tachinidae]; Graphomyzina Macquart, 1835 [Sciomyzidae]; Gymnostylina Macquart, 1835 [Tachinidae]; Heterometopia Macquart, 1846 [Tachinidae]; Laxenecera Macquart, 1838 [Asilidae]; Leptomyza Macquart, 1835 [Anthomyzidae]; Megistogaster Macquart, 1851 [Tachinidae]; Microtrichodes Macquart, 1846 [Tachinidae]; Microtropesa Macquart, 1846 [Tachinidae]; Ogcodocera Macquart, 1840 [Bombyliidae]; Onuxicera Macquart, 1855 [Tachinidae]; Ozodicera Macquart, 1834 [Tipulidae]; Pachymerina Macquart, 1834 [Empididae]; Pachyrhina Macquart, 1834 [Tipulidae]; Pachystylum Macquart, 1848 [Tachinidae]; Physegaster Macquart, 1847 [Acroceridae]; Plesionevra Macquart, 1855 [Tachinidae]; Rhopalia Macquart, 1838 [Mydidae]; Semiomyia Macquart, 1848 [Tachinidae]; Senostoma Macquart, 1847 [Tachinidae]; Silbomyia Macquart, 1844 [Calliphoridae]; Sumpigaster Macquart, 1855 [Tachinidae]; Tapinocera Macquart, 1838 [Apioceridae]; Teretrophora Macquart, 1851 [Tachinidae]; Toxocnemis Macquart, 1855 [Tachinidae]; Toxotarsus Macquart, 1851 [Calliphoridae]; Trichostylum Macquart, 1851 [Tachinidae]; Trigonometopus Macquart, 1835 [Lauxaniidae]; Tritaxys Macquart, 1847 [Tachinidae]; Vermileo Macquart, 1834 [Vermileonidae].        Acting as First Reviser, the following correct original spellings for multiple original spellings are selected by us-(for genus-group names): Choeteprosopa Macquart, 1851 [Tachinidae]; Dichoetometopia Macquart, 1855 [Sarcophagidae]; Discocerina Macquart, 1835 [Ephydridae]; Dolichocephala Macquart, 1823 [Empididae]; Dolichomerus Macquart, 1850 [Syrphidae]; Graphalostylum Macquart, 1851 [Tachinidae]; Hemilampra Macquart, 1850 [Syrphidae]; Leptomyza Macquart, 1835 [Anthomyzidae]; Microcheilosia Macquart, 1855 [Tachinidae]; Phrissopodia Macquart, 1835 [Sarcophagidae]; Platytainia Macquart, 1851 [Tachinidae]; Polychaeta Macquart, 1851 [Tachinidae]; Stachynia Macquart, 1835 [Conopidae]-(for species-group names): Cadicera rubramarginata Macquart, 1855 [Tabanidae].        Previous First Reviser actions for multiple original spellings that were missed by other workers are given for the following: Amethysa Macquart, 1835 [Ulidiidae]; Anabarhynchus Macquart, 1848 [Therevidae]; Anacanthella Macquart, 1855 [Stratiomyidae]; Aulacigaster Macquart, 1835 [Aulacigastridae]; Cardiacera Macquart, 1847 [Pyrgotidae]; Comptosia Macquart, 1840 [Bombyliidae]; Craspedia Macquart, 1838 [Asilidae]; Cyclorhynchus Macquart, 1840 [Bombyliidae]; Ectinorhynchus Macquart, 1850 [Therevidae]; Euthinevra Macquart, 1836 [Hybotidae]; Gonistylum Macquart, 1851 [Tachinidae]; Heterostylum Macquart, 1848 [Bombyliidae]; Hoplistomera Macquart, 1838 [Asilidae]; Hystricephala Macquart, 1846 [Tachinidae]; Leptoxyda Macquart, 1835 [Tephritidae]; Nemopalpus Macquart, 1838 [Psychodidae]; Senotainia Macquart, 1846 [Sarcophagidae]; Spilogaster Macquart, 1835 [Muscidae]; Spogostylum Macquart, 1840 [Bombyliidae]; Stachynia Macquart, 1835 [Conopidae].        Invoking ICZN Code Article 33.3.1, the following is here considered a correct original spelling by being in prevailing usage: Leptopeza Macquart, 1828 [Empididae].        Reversal of Precedence (ICZN Code Article 23.9) is invoked to promote stability in nomenclature for the following cases of subjective synonymy: Atherigona Rondani, 1856, nomen protectum and Orthostylum Macquart, 1851, nomen oblitum [in Muscidae]; Clusiodes Coquillett, 1904, nomen protectum and Heteronevra Macquart, 1835, nomen oblitum [in Clusiidae]; Senotainia Macquart, 1846, nomen protectum and Megoera Macquart, 1834, nomen oblitum [in Sarcophagidae].        The following genus-group names, not listed in current regional catalogs, are treated here: Diasema Macquart, 1835 [Chloropidae]; Dichromyia Macquart, 1844 [Heleomyzidae]; Elomyia Macquart, 1834 [Tachinidae]; Eriosoma Macquart, 1838 [Acroceridae]; Eurypalpus Macquart, 1835 [Platystomatidae]; Notacanthina Macquart, 1835 [Ephydridae]; Pleurocerina Macquart, 1851[Conopidae]; Pteropexus Macquart, 1846 [Acroceridae]; Semiomyia Macquart, 1848 [Tachinidae]; Teremyia Macquart, 1835 [Lonchaeidae].        The following names are new synonymies of their respective senior synonyms: -genus-group names: Acemyia Macquart, 1834 of Acemya Robineau-Desvoidy, 1830, n. syn. [Tachinidae]; Acrochoeta Macquart, 1835 of Acrochaeta Wiedemann, 1830, n. syn. [Stratiomyidae]; Atractea Agassiz, 1846 of Atractia Macquart, 1838, n. syn. [Asilidae]; Aulacocephala Brauer, 1863 of Aulacephala Macquart, 1851, n. syn. [Tachinidae]; Beckeriella Williston, 1897 of Notacanthina Macquart, 1834, n. syn. [Ephydridae]; Caenosia Macquart, 1835 of Coenosia Meigen, 1826, n. syn. [Muscidae]; Ceromyia Macquart, 1834 of Ceromya Robineau-Desvoidy, 1830, n. syn. [Tachinidae]; Chiromysa Macquart, 1835 of Chiromyza Wiedemann, 1820, n. syn. [Stratiomyidae]; Chrisochlora Macquart, 1835 of Chrysochlora Latreille, 1829, n. syn. [Stratiomyidae]; Chrysopyla Macquart, 1840 of Chrysopilus Macquart, 1826, n. syn. [Rhagionidae]; Cleigaster Macquart, 1844 of Cleigastra Macquart, 1835, n. syn. [Scathophagidae]; Clyto Macquart, 1835 of Clytho Robineau-Desvoidy, 1830, n. syn. [Tachinidae]; Cordylura Macquart, 1835 of Cordilura Fallén, 1810, n. syn. [Scathophagidae]; Craspedochaeta Marschall, 1873 of Anthomyia Meigen, 1803, n. syn. [Anthomyiidae]; Cyrtonevra Agassiz, 1846 of Graphomya Robineau-Desvoidy, 1830, n. syn. [Muscidae]; Diaphora Macquart, 1834 of Diaphorus Meigen, 1824, n. syn. [Dolichopodidae]; Dichoeta Macquart, 1835 of Dichaeta Meigen, 1830, n. syn. [Ephydridae]; Dichromyia Macquart, 1844 of Dichromya Robineau-Desvoidy, 1830, n. syn. [Heleomyzidae]; Diphysa Macquart, 1838 of Archistratiomys Enderlein, 1913, n. syn. [Stratiomyidae]; Echinomyia Fischer von Waldheim, 1808 of Tachina Meigen, 1803, n. syn. [Tachinidae]; Egina Macquart, 1835 of Eginia Robineau-Desvoidy, 1830, n. syn. [Muscidae]; Hematobia Macquart, 1850 of Haematobia Le Peletier & Audinet-Serville, 1828, n. syn. [Muscidae]; Hemerodromyia Macquart, 1823 of Hemerodromia Meigen, 1822, n. syn. [Empididae]; Heteronevra Macquart, 1835 of Clusiodes Coquillett, 1904, n. syn. [Clusiidae]; Himastima Agassiz, 1846 of Mallota Meigen, 1822, n. syn. [Syrphidae]; Hoematopota Macquart, 1826 of Haematopota Meigen, 1803, n. syn. [Tabanidae]; Homalocephala Agassiz, 1846 of Setellia Robineau-Desvoidy, 1830, n. syn. [Sciomyzidae]; Hydrotoea Macquart, 1844 of Hydrotaea Robineau-Desvoidy, 1830, n. syn. [Muscidae]; Linnemyia Macquart, 1834 of Linnaemya Robineau-Desvoidy, 1830, n. syn. [Tachinidae]; Lonchoea Macquart, 1835 of Lonchaea Fallén, 1820b, n. syn. [Lonchaeidae]; Macromyia Macquart, 1835 of Macromya Robineau-Desvoidy, 1830, n. syn. [Tachinidae]; Megarhina Macquart, 1838 of Lynchiella Lahille, 1904, n. syn. [Culicidae]; Meriana Macquart, 1835 of Panzeria Robineau-Desvoidy, 1830, n. syn. [Tachinidae]; Microphorus Lundbeck, 1907 of Microphor Macquart, 1834, n. syn. [Dolichopodidae]; Nemoroea Macquart, 1844 of Nemoraea Robineau-Desvoidy, 1830, n. syn. [Tachinidae]; Ochthiphila Macquart, 1850 of Chamaemyia Meigen, 1803, n. syn. [Chamaemyiidae]; Ocydromyia Macquart, 1823 of Ocydromia Meigen, 1820, n. syn. [Hybotidae]; Oliviera Macquart, 1835 of Eriothrix Meigen, 1803, n. syn. [Tachinidae]; Ophilia Macquart, 1850 of Metopia Meigen, 1803, n. syn. [Sarcophagidae]; Ornithomyia Fischer von Waldheim, 1808 of Ornithomya Latreille, 1802, n. syn. [Hippoboscidae]; Orthochile Westwood, 1840 of Ortochile Latreille, 1809, n. syn. [Dolichopodidae]; Osmoea Macquart, 1834 of Triarthria Stephens, 1829, n. syn. [Tachinidae]; Pachyrrhina Osten Sacken, 1878 of Pachyrhina Macquart, 1834, n. syn. [Tipulidae]; Palis Macquart, 1850 of Pales Robineau-Desvoidy, 1830, n. syn. [Tachinidae]; Phanemia Macquart, 1835 of Clairvillia Robineau-Desvoidy, 1830, n. syn. [Tachinidae]; Phrissopoda Macquart, 1851 of Peckia Robineau-Desvoidy, 1830, n. syn. [Sarcophagidae]; Phyllomyia Macquart, 1834 of Phyllomya Robineau-Desvoidy, 1830, n. syn. [Tachinidae]; Physogenia Loew, 1862 of Physegenua Macquart, 1848, n. syn. [Lauxaniidae]; Physogenua Giglio-Tos, 1895 of Physegenua Macquart, 1848, n. syn. [Lauxaniidae]; Phytomiza Macquart, 1835 of Phytomyza Fallén, 1810, n. syn. [Agromyzidae]; Platipalpus Macquart, 1850 of Platypalpus Macquart, 1828, n. syn. [Hybotidae]; Platipeza Macquart, 1850 of Platypeza Meigen, 1803, n. syn. [Platypezidae]; Platynochoetus Macquart, 1834 of Platynochaetus Wiedemann, 1830 [Syrphidae]; Porphirops Macquart, 1838 of Porphyrops Meigen, 1824, n. syn [Dolichopodidae]; Rhinomyia Macquart, 1835 of Rhinomya Robineau-Desvoidy, 1830, n. syn. [Tachinidae]; Rhynomyia Macquart, 1834 of Rhinomya Robineau-Desvoidy, 1830, n. syn. [Tachinidae]; Scathopse Guérin-Méneville, 1839 of Scatopse Geoffroy, 1762, n. syn. [Scatopsidae]; Spherophoria Macquart, 1850 of Sphaerophoria Le Peletier & Audinet-Serville, 1828, n. syn. [Syrphidae]; Sphoerophoria Macquart, 1829 of Sphaerophoria Le Peletier & Audinet-Serville, 1828, n. syn. [Syrphidae]; Stenopteryx Schiner, 1864 of Stenepteryx Leach, 1817, n. syn. [Hippoboscidae]; Stenostoma Mik, 1890 of Senostoma Macquart, 1847, n. syn. [Tachinidae]; Tachydromyia Macquart, 1823 of Tachydromia Meigen, 1803, n. syn. [Hybotidae]; Taenioptera Mik, 1898 of Taeniaptera Macquart, 1835, n. syn. [Micropezidae]; Trinevra Macquart, 1835 of Phora Latreille, 1797, n. syn. [Phoridae]; Uramyia Macquart, 1844 of Uramya Robineau-Desvoidy, 1830, n. syn. [Tachinidae]; Xestomysa Macquart, 1851 of Xestomyza Wiedemann, 1820, n. syn. [Therevidae]; Zygonevra Macquart, 1834 of Zygoneura Meigen, 1803, n. syn. [Sciaridae]. -Species-group names: Calobatemyia nigra Macquart, 1855 of Musca doronici Scopoli, 1763, n. syn. [Calliphoridae]; Cyrtonevra protorum Macquart, 1850 of Musca pratorum Meigen, 1826, n. syn. [Muscidae]; Eumerus oeneus Macquart, 1850 of Eumerus aeneus Macquart, 1829, n. syn. [Syrphidae]; Lucilia ceserion Macquart, 1850 of Musca caesarion Scharfenberg, 1805, n. syn. [Calliphoridae]; Masicera sylvatica Macquart, 1850 of Tachina silvatica Fallén, 1810, n. syn. [Tachinidae]; Ophyra anolis Macquart, 1850 of Ophyra analis Macquart, 1846, n. syn. [Muscidae]; Pegomyia hyosciami Macquart, 1850 of Musca hyoscyami Panzer, 1798, n. syn. [Anthomyiidae]; Prosena syberita Macquart, 1850 of Stomoxys siberita Fabricius, 1775, n. syn. [Tachinidae]; Taxigramma heteronevra Macquart, 1850 of Miltogramma heteroneura Meigen, 1830, n. syn. [Sarcophagidae].

Knock Down of Heat Shock Protein 27 (HspB1) Induces Degradation of Several Putative Client Proteins

PubMed Central

Gibert, Benjamin; Eckel, Bénédicte; Fasquelle, Lydie; Moulin, Maryline; Bouhallier, Frantz; Gonin, Vincent; Mellier, Gregory; Simon, Stéphanie; Kretz-Remy, Carole; Arrigo, André-Patrick; Diaz-Latoud, Chantal

2012-01-01

Hsp27 belongs to the heat shock protein family and displays chaperone properties in stress conditions by holding unfolded polypeptides, hence avoiding their inclination to aggregate. Hsp27 is often referenced as an anti-cancer therapeutic target, but apart from its well-described ability to interfere with different stresses and apoptotic processes, its role in non-stressed conditions is still not well defined. In the present study we report that three polypeptides (histone deacetylase HDAC6, transcription factor STAT2 and procaspase-3) were degraded in human cancerous cells displaying genetically decreased levels of Hsp27. In addition, these proteins interacted with Hsp27 complexes of different native size. Altogether, these findings suggest that HDAC6, STAT2 and procaspase-3 are client proteins of Hsp27. Hence, in non stressed cancerous cells, the structural organization of Hsp27 appears to be a key parameter in the regulation by this chaperone of the level of specific polypeptides through client-chaperone type of interactions. PMID:22238643

Biomimetic Synthesis of Gelatin Polypeptide-Assisted Noble-Metal Nanoparticles and Their Interaction Study

NASA Astrophysics Data System (ADS)

Liu, Ying; Liu, Xiaoheng; Wang, Xin

2011-12-01

Herein, the generation of gold, silver, and silver-gold (Ag-Au) bimetallic nanoparticles was carried out in collagen (gelatin) solution. It first showed that the major ingredient in gelatin polypeptide, glutamic acid, acted as reducing agent to biomimetically synthesize noble metal nanoparticles at 80°C. The size of nanoparticles can be controlled not only by the mass ratio of gelatin to gold ion but also by pH of gelatin solution. Interaction between noble-metal nanoparticles and polypeptide has been investigated by TEM, UV-visible, fluorescence spectroscopy, and HNMR. This study testified that the degradation of gelatin protein could not alter the morphology of nanoparticles, but it made nanoparticles aggregated clusters array (opposing three-dimensional α-helix folding structure) into isolated nanoparticles stabilized by gelatin residues. This is a promising merit of gelatin to apply in the synthesis of nanoparticles. Therefore, gelatin protein is an excellent template for biomimetic synthesis of noble metal/bimetallic nanoparticle growth to form nanometer-sized device.

Biomimetic Synthesis of Gelatin Polypeptide-Assisted Noble-Metal Nanoparticles and Their Interaction Study

PubMed Central

2011-01-01

Herein, the generation of gold, silver, and silver–gold (Ag–Au) bimetallic nanoparticles was carried out in collagen (gelatin) solution. It first showed that the major ingredient in gelatin polypeptide, glutamic acid, acted as reducing agent to biomimetically synthesize noble metal nanoparticles at 80°C. The size of nanoparticles can be controlled not only by the mass ratio of gelatin to gold ion but also by pH of gelatin solution. Interaction between noble-metal nanoparticles and polypeptide has been investigated by TEM, UV–visible, fluorescence spectroscopy, and HNMR. This study testified that the degradation of gelatin protein could not alter the morphology of nanoparticles, but it made nanoparticles aggregated clusters array (opposing three-dimensional α-helix folding structure) into isolated nanoparticles stabilized by gelatin residues. This is a promising merit of gelatin to apply in the synthesis of nanoparticles. Therefore, gelatin protein is an excellent template for biomimetic synthesis of noble metal/bimetallic nanoparticle growth to form nanometer-sized device. PMID:27502645

Synapsin I and Synapsin II regulate neurogenesis in the dentate gyrus of adult mice

PubMed Central

Barbieri, Raffaella; Contestabile, Andrea; Ciardo, Maria Grazia; Forte, Nicola; Marte, Antonella; Baldelli, Pietro; Benfenati, Fabio; Onofri, Franco

2018-01-01

Adult neurogenesis is emerging as an important player in brain functions and homeostasis, while impaired or altered adult neurogenesis has been associated with a number of neuropsychiatric diseases, such as depression and epilepsy. Here we investigated the possibility that synapsins (Syns) I and II, beyond their known functions in developing and mature neurons, also play a role in adult neurogenesis. We performed a systematic evaluation of the distinct stages of neurogenesis in the hippocampal dentate gyrus of Syn I and Syn II knockout (KO) mice, before (2-months-old) and after (6-months-old) the appearance of the epileptic phenotype. We found that Syns I and II play an important role in the regulation of adult neurogenesis. In juvenile mice, Syn II deletion was associated with a specific decrease in the proliferation of neuronal progenitors, whereas Syn I deletion impaired the survival of newborn neurons. These defects were reverted after the appearance of the epileptic phenotype, with Syn I KO and Syn II KO mice exhibiting significant increases in survival and proliferation, respectively. Interestingly, long-term potentiation dependent on newborn neurons was present in both juvenile Syn mutants while, at later ages, it was only preserved in Syn II KO mice that also displayed an increased expression of brain-derived neurotrophic factor. This study suggests that Syns I and II play a role in adult neurogenesis and the defects in neurogenesis associated with Syn deletion may contribute to the alterations of cognitive functions observed in Syn-deficient mice. PMID:29721159

Regulatory peptide distribution in separated layers of the human jejunum.

PubMed

Ferri, G L; Adrian, T E; Soimero, L; McGregor, G P; Ghatei, M A; Morreale, R A; Rebecchi, L; Tonelli, L; Polak, J M; Bloom, S R

1987-01-01

The distribution of regulatory peptides was studied in the separated epithelium, lamina propria, submucosa and muscularis externa of the human jejunum. Gastrin, secretin, gastric inhibitory polypeptide, enteroglucagon and neurotensin immunoreactivity were almost confined to the endocrine cell-containing mucosal epithelium (greater than 98% of the total content), only minor amounts of motilin being detected in non-epithelial layers (3.6 +/- 0.7%, mean +/- SEM, n = 7). Conversely, vasoactive intestinal polypeptide, substance P and mammalian bombesin were virtually limited to non-epithelial layers (greater than 99%). Only somatostatin was found in all layers (44 +/- 6.7% in the epithelium, 34 +/- 5.2% in the lamina propria, 13 +/- 2.9% in the submucosa, and 7.9 +/- 2.8% in the muscularis). Substance P was found in higher concentrations in the mucosa, compared to submucosa and muscle (56 +/- 10, 30 +/- 4.0 and 29 +/- 4.0 pmol/g, respectively), while vasoactive intestinal polypeptide was more abundant in the muscle (411 +/- 52 pmol/g) compared to mucosa and submucosa (228 +/- 64 and 219 +/- 31 pmol/g, respectively). Only low levels of mammalian bombesin were measured, mainly in the muscle (6.9 +/- 1.5 pmol/g, or 89 +/- 3.6% of total content).

Polypeptide multilayer film co-delivers oppositely-charged drug molecules in sustained manners.

PubMed

Jiang, Bingbing; Defusco, Elizabeth; Li, Bingyun

2010-12-13

The current state-of-the-art for drug-carrying biomedical devices is mostly limited to those that release a single drug. Yet there are many situations in which more than one therapeutic agent is needed. Also, most polyelectrolyte multilayer films intended for drug delivery are loaded with active molecules only during multilayer film preparation. In this paper, we present the integration of capsules as vehicles within polypeptide multilayer films for sustained release of multiple oppositely charged drug molecules using layer-by-layer nanoassembly technology. Calcium carbonate (CaCO(3)) particles were impregnated with polyelectrolytes, shelled with polyelectrolyte multilayers, and then assembled onto polypeptide multilayer films using glutaraldehyde. Capsule-integrated polypeptide multilayer films were obtained after decomposition of CaCO(3) templates. Two oppositely charged drugs were loaded into capsules within polypeptide multilayer films postpreparation based on electrostatic interactions between the drugs and the polyelectrolytes impregnated within capsules. We determined that the developed innovative capsule-integrated polypeptide multilayer films could be used to load multiple drugs of very different properties (e.g., opposite charges) any time postpreparation (e.g., minutes before surgical implantation inside an operating room), and such capsule-integrated films allowed simultaneous delivery of two oppositely charged drug molecules and a sustained (up to two weeks or longer) and sequential release was achieved.

Polypeptide Multilayer Film Co-Delivers Oppositely-Charged Drug Molecules in Sustained Manners

PubMed Central

Jiang, Bingbing; DeFusco, Elizabeth; Li, Bingyun

2010-01-01

The current state-of-the-art for drug-carrying biomedical devices is mostly limited to those that release a single drug. Yet there are many situations in which more than one therapeutic agent is needed. Also, most polyelectrolyte multilayer films intending for drug delivery are loaded with active molecules only during multilayer film preparation. In this paper, we present the integration of capsules as vehicles within polypeptide multilayer films for sustained release of multiple oppositely-charged drug molecules using layer-by-layer nanoassembly technology. Calcium carbonate (CaCO3) particles were impregnated with polyelectrolytes, shelled with polyelectrolyte multilayers, and then assembled onto polypeptide multilayer films using glutaraldehyde. Capsule-integrated polypeptide multilayer films were obtained after decomposition of CaCO3 templates. Two oppositely-charged drugs were loaded into capsules within polypeptide multilayer films post-preparation based on electrostatic interactions between the drugs and the polyelectrolytes impregnated within capsules. We determined that the developed innovative capsule-integrated polypeptide multilayer films could be used to load multiple drugs of very different properties (e.g. opposite charges) any time post-preparation (e.g. minutes before surgical implantation inside an operating room), and such capsule-integrated films allowed simultaneous delivery of two oppositely-charged drug molecules and a sustained (up to two weeks or longer) and sequential release was achieved. PMID:21058719

Suppression of α-synuclein toxicity and vesicle trafficking defects by phosphorylation at S129 in yeast depends on genetic context

PubMed Central

Sancenon, Vicente; Lee, Sue-Ann; Patrick, Christina; Griffith, Janice; Paulino, Amy; Outeiro, Tiago F.; Reggiori, Fulvio; Masliah, Eliezer; Muchowski, Paul J.

2012-01-01

The aggregation of α-synuclein (αSyn) is a neuropathologic hallmark of Parkinson's disease and other synucleinopathies. In Lewy bodies, αSyn is extensively phosphorylated, predominantly at serine 129 (S129). Recent studies in yeast have shown that, at toxic levels, αSyn disrupts Rab homeostasis, causing an initial endoplasmic reticulum-to-Golgi block that precedes a generalized trafficking collapse. However, whether αSyn phosphorylation modulates trafficking defects has not been evaluated. Here, we show that constitutive expression of αSyn in yeast impairs late-exocytic, early-endocytic and/or recycling trafficking. Although members of the casein kinase I (CKI) family phosphorylate αSyn at S129, they attenuate αSyn toxicity and trafficking defects by an S129 phosphorylation-independent mechanism. Surprisingly, phosphorylation of S129 modulates αSyn toxicity and trafficking defects in a manner strictly determined by genetic background. Abnormal endosome morphology, increased levels of the endosome marker Rab5 and co-localization of mammalian CKI with αSyn aggregates are observed in brain sections from αSyn-overexpressing mice and human synucleinopathies. Our results contribute to evidence that suggests αSyn-induced defects in endocytosis, exocytosis and/or recycling of vesicles involved in these cellular processes might contribute to the pathogenesis of synucleinopathies. PMID:22357655

The N-Terminal Residues 43 to 60 Form the Interface for Dopamine Mediated α-Synuclein Dimerisation

PubMed Central

Leong, Su Ling; Hinds, Mark G.; Connor, Andrea R.; Smith, David P.; Illes-Toth, Eva; Pham, Chi L. L.; Barnham, Kevin J.; Cappai, Roberto

2015-01-01

α-synuclein (α-syn) is a major component of the intracellular inclusions called Lewy bodies, which are a key pathological feature in the brains of Parkinson’s disease patients. The neurotransmitter dopamine (DA) inhibits the fibrillisation of α-syn into amyloid, and promotes α-syn aggregation into SDS-stable soluble oligomers. While this inhibition of amyloid formation requires the oxidation of both DA and the methionines in α-syn, the molecular basis for these processes is still unclear. This study sought to define the protein sequences required for the generation of oligomers. We tested N- (α-syn residues 43–140) and C-terminally (1–95) truncated α-syn, and found that similar to full-length protein both truncated species formed soluble DA:α-syn oligomers, albeit 1–95 had a different profile. Using nuclear magnetic resonance (NMR), and the N-terminally truncated α-syn 43–140 protein, we analysed the structural characteristics of the DA:α-syn 43–140 dimer and α-syn 43–140 monomer and found the dimerisation interface encompassed residues 43 to 60. Narrowing the interface to this small region will help define the mechanism by which DA mediates the formation of SDS-stable soluble DA:α-syn oligomers. PMID:25679387

Method to determine transcriptional regulation pathways in organisms

DOEpatents

Gardner, Timothy S.; Collins, James J.; Hayete, Boris; Faith, Jeremiah

2012-11-06

The invention relates to computer-implemented methods and systems for identifying regulatory relationships between expressed regulating polypeptides and targets of the regulatory activities of such regulating polypeptides. More specifically, the invention provides a new method for identifying regulatory dependencies between biochemical species in a cell. In particular embodiments, provided are computer-implemented methods for identifying a regulatory interaction between a transcription factor and a gene target of the transcription factor, or between a transcription factor and a set of gene targets of the transcription factor. Further provided are genome-scale methods for predicting regulatory interactions between a set of transcription factors and a corresponding set of transcriptional target substrates thereof.

Identification of critical residues in Hepatitis E virus macro domain involved in its interaction with viral methyltransferase and ORF3 proteins

PubMed Central

Anang, Saumya; Subramani, Chandru; Nair, Vidya P.; Kaul, Sheetal; Kaushik, Nidhi; Sharma, Chandresh; Tiwari, Ashutosh; Ranjith-Kumar, CT; Surjit, Milan

2016-01-01

Hepatitis E virus (HEV) is a major cause of hepatitis in normal and organ transplant individuals. HEV open reading frame-1 encodes a polypeptide comprising of the viral nonstructural proteins as well as domains of unknown function such as the macro domain (X-domain), V, DUF3729 and Y. The macro domain proteins are ubiquitously present from prokaryotes to human and in many positive-strand RNA viruses, playing important roles in multiple cellular processes. Towards understanding the function of the HEV macro domain, we characterized its interaction partners among other HEV encoded proteins. Here, we report that the HEV X-domain directly interacts with the viral methyltransferase and the ORF3 proteins. ORF3 association with the X-domain was mediated through two independent motifs, located within its N-terminal 35aa (amino acids) and C-terminal 63-123aa. Methyltransferase interaction domain was mapped to N-terminal 30-90aa. The X-domain interacted with both ORF3 and methyltransferase through its C-terminal region, involving 66th,67th isoleucine and 101st,102nd leucine, conserved across HEV genotypes. Furthermore, ORF3 and methyltransferase competed with each other for associating with the X-domain. These findings provide molecular understanding of the interaction between the HEV macro domain, methyltransferase and ORF3, suggesting an important role of the macro domain in the life cycle of HEV. PMID:27113483

Taxonomic changes in African Stratiomyidae (Diptera).

PubMed

Hauser, Martin; Woodley, Norman E; Fachin, Diego A

2017-05-08

Thirteen new generic synonyms, nineteen species synonyms and forty-eight new combinations of African Stratiomyidae are proposed (senior synonym in parentheses):Arthronemina Lindner in James, 1980 syn. nov. (=Argyrobrithes Grünberg, 1915), Arthronema Lindner, 1966b syn. nov. (=Argyrobrithes Grünberg, 1915), Brachyphleps Lindner, 1965 syn. nov. (=Psapharomys Grünberg, 1915), Dinosargus Lindner, 1968 syn. nov. (=Gongrosargus Lindner, 1959), Dolichodema Kertész, 1916 syn. nov. (=Thorasena Macquart, 1838), Gobertina Bigot, 1879a syn. nov. (=Sternobrithes Loew, 1857), Himantochaeta Lindner, 1939 syn. nov. (=Nyplatys Séguy, 1938), Hypoxycera Lindner 1966a syn. nov. (=Hypoceromys Lindner, 1935), Leucacron Lindner, 1966b syn. nov. (=Ptilinoxus Lindner, 1966b), Lonchobrithes Lindner, 1968 syn. nov. (=Argyrobrithes Grünberg, 1915), Meristomeringella Lindner 1965 syn. nov. (=Hypoceromys Lindner, 1935), Physometopon Lindner, 1966b syn. nov. (=Cardopomyia Kertész, 1916), Psapharomydops Lindner, 1966a syn. nov. (=Steleoceromys Grünberg, 1915), Adoxomyia grisea (Séguy, 1931) syn. nov. (=Adoxomyia argenteofasciata (Bezzi, 1906)), Argyrobrithes argenteus Grünberg, 1915 syn. nov. (=Argyrobrithes fuscicornis (Bezzi, 1914)), Argyrobrithes crinitus Lindner, 1972 syn. nov. (=Argyrobrithes zernyi Lindner, 1943), Brachyphleps tristis Lindner, 1965 syn. nov. (=Psapharomys salebrosa Grünberg, 1915), Chrysochroma laetum Lindner, 1966b syn. nov. (=Ptectisargus abditus (Lindner, 1936), Dolichodema africana Kertész, 1916 syn. nov. (=Thorasena pectoralis (Wiedemann, 1838)), Gongrosargus distinguendus Lindner, 1966c syn. nov. (=Gongrosargus glaucus (Bigot, 1859)), Gongrosargus exclamationis Lindner, 1968 syn. nov. (=Gongrosargus pallidus (Macquart, 1838)), Gongrosargus univittatus Lindner, 1966b syn. nov. (=Gongrosargus pallidus (Macquart, 1838)), Hypoxycera simplex Lindner, 1966a syn. nov. (=Hypoceromys jamesi (Lindner, 1965)), Lonchobrithes modestus Lindner, 1968 syn. nov. (=Argyrobrithes curtilamellatum (Lindner, 1966)), Microptecticus clarus Lindner, 1968 syn. nov. (=Microptecticus ambiguus Lindner, 1966b), Neopachygaster umbrifera Lindner, 1966a syn. nov. (=Neopachygaster stigma Lindner, 1938), Odontomyia impressa Curran, 1928 syn. nov. (=Afrodontomyia gigas (Brunetti, 1926)), Odontomyia protrudens Curran, 1928 syn. nov. (=Afrodontomyia erecta (Brunetti, 1926)), Physometopon minor Lindner, 1968 syn. nov. (=Cardopomyia robusta Kertész, 1916), Platyna denudata Grünberg, 1915 syn. nov. (=Platyna hastata (Fabricius, 1805)), Ptectisargus lucidus Lindner, 1968 syn. nov. (=Ptectisargus abditus (Lindner, 1936)); Afrodontomyia erecta (Brunetti, 1926) comb. nov. (from Odontomyia), Afrodontomyia flammiventris (Brunetti, 1926) comb. nov. (from Odontomyia), Afrodontomyia rufiventris (Curran, 1928) comb. nov. (from Stratiomys), Argyrobrithes curtilamellatum (Lindner, 1966b) comb. nov. (from Arthronemina), Argyrobrithes fuscicornis (Bezzi, 1914) comb. nov. (from Sternobrithes), Cardopomyia parvicornis (Lindner, 1959) comb. nov. (from Pseudoxymyia Lindner, 1958), Cardopomyia vesicularis (Lindner, 1966b) comb. nov. (from Physometopon), Cephalochrysa bigoti (Lindner, 1968) comb. nov. (from Chrysochroma), Cephalochrysa flavum (Lindner, 1968) comb. nov. (from Chrysochroma), Cephalochrysa fortunatum (Lindner, 1966b) comb. nov. (from Chrysochroma), Cephalochrysa lapidis (Lindner, 1966b) comb. nov. (from Chrysochroma), Cephalochrysa latum (Lindner, 1966b) comb. nov. (from Chrysochroma), Cephalochrysa lucens (Lindner, 1968) comb. nov. (from Chrysochroma), Cephalochrysa matilei (Lindner, 1979) comb. nov. (from Chrysochroma), Cephalochrysa triste (Lindner, 1966b) comb. nov. (from Chrysochroma), Cephalochrysa turbidum (Lindner, 1965) comb. nov. (from Chrysochroma), Cephalochrysa vadoni (Lindner, 1966b) comb. nov. (from Chrysochroma), Gongrosargus flavipennis (Macquart, 1838) comb. nov. (from Sargus), Gongrosargus lateritius (Lindner, 1968) comb. nov. (from Dinosargus), Gongrosargus limbatus (Macquart, 1838) comb. nov. (from Sargus), Gongrosargus pallidus (Macquart, 1838) comb. nov. (from Sargus), Hypoceromys nigripes (Lindner, 1938) comb. nov. (from Pachygaster), Hypoceromys jamesi (Lindner, 1965) comb. nov. (from Meristomeringella), Microptecticus magnicornis (Lindner, 1936) comb. nov. (from Ptecticus), Microptecticus nigricoxa (Lindner, 1936) comb. nov. (from Microchrysa), Ptecticus lateritius (Rondani, 1863) comb. nov. (from Sargus), Ptectisargus abditus (Lindner, 1936) comb. nov. (from Ptecticus), Ptectisargus brunneus (Lindner, 1936) comb. nov. (from Ptecticus), Ptectisargus cingulatum (Lindner, 1968) comb. nov. (from Chrysochroma), Ptectisargus flavifrons (Lindner, 1968) comb. nov. (from Chrysochroma), Ptectisargus flavomarginatus (Loew, 1857) comb. nov. (from Chrysonotus), Ptectisargus gracilipes (Lindner, 1936) comb. nov. (from Ptecticus), Ptectisargus keiseri (Lindner, 1966b) comb. nov. (from Chrysochroma), Ptectisargus longestylum (Lindner, 1966b) comb. nov. (from Chrysochroma), Ptectisargus punctum (Lindner, 1968) comb. nov. (from Chrysochroma), Ptectisargus ranohira (Woodley, 2001) comb. nov. (from Chrysochroma), Ptectisargus unicolor (Lindner, 1968) comb. nov. (from Chrysochroma), Ptilinoxus interruptum (Lindner, 1966b) comb. nov. (from Leucacron), Sargus congoense (Lindner, 1965) comb. nov. (from Chrysochroma), Sargus flavipes (Lindner, 1966a) comb. nov. (from Chrysochroma), Sargus luctuosus (Lindner, 1938) comb. nov. (from Paraptecticus), Sargus opulentum (Grünberg, 1915) comb. nov. (from Chrysochroma), Sargus pallidiventre (Brunetti, 1926) comb. nov. (from Chrysochroma), Sargus ptecticoideum (Lindner, 1966a) comb. nov. (from Chrysochroma), Steleceromys procera (Lindner, 1966a) comb. nov. (from Psapharomydops), Sternobrithes mercurialis (Lindner, 1938) comb. nov. (from Gobertina), Sternobrithes picticornis (Bigot, 1879b). comb. nov. (from Gobertina), Thorasena pectoralis (Wiedemann, 1824) comb. nov. (from Hermetia), Thorasena fenestrata (James, 1949) comb. nov. (from Dolichodema). One genus was resurrected out of synonymy (Thorasena Macquart, 1838 stat. rev.) and one genus removed from the African fauna (Cyphomyia Wiedemann, 1819).

Probing the organic-mineral interface at the molecular level in model biominerals.

PubMed

Metzler, Rebecca A; Kim, Il Won; Delak, Katya; Evans, John Spencer; Zhou, Dong; Beniash, Elia; Wilt, Fred; Abrecht, Mike; Chiou, Jau-Wern; Guo, Jinghua; Coppersmith, Susan N; Gilbert, P U P A

2008-03-18

It is widely known that macromolecules, such as proteins, can control the nucleation and growth of inorganic solids in biomineralizing organisms. However, what is not known are the complementary molecular interactions, organization, and rearrangements that occur when proteins interact with inorganic solids during the formation of biominerals. The organic-mineral interface (OMI) is expected to be the site for these phenomena, and is therefore extraordinarily interesting to investigate. In this report, we employ X-ray absorption near edge (XANES) spectromicroscopy to investigate the electronic structure of both calcium carbonate mineral crystals and polypeptides, and detect changing bonds at the OMI during crystal growth in the presence of polypeptides. We acquired XANES spectra from calcium carbonate crystals grown in the presence of three mollusk nacre-associated polypeptides (AP7N, AP24N, n16N) and in the presence of a sea urchin spicule matrix protein, LSM34. All these model biominerals gave similar results, including the disruption of CO bonds in calcite and enhancement of the peaks associated with C-H bonds and C-O bonds in peptides, indicating ordering of the amino acid side chains in the mineral-associated polypeptides and carboxylate binding. This is the first evidence of the mutual effect of calcite on peptide chain and peptide chain on calcite during biomineralization. We also show that these changes do not occur when Asp and Glu are replaced in the n16N sequence with Asn and Gln, respectively, demonstrating that carboxyl groups in Asp and Glu do participate in polypeptide-mineral molecular associations.

SYN2 is an autism predisposing gene: loss-of-function mutations alter synaptic vesicle cycling and axon outgrowth

PubMed Central

Corradi, Anna; Fadda, Manuela; Piton, Amélie; Patry, Lysanne; Marte, Antonella; Rossi, Pia; Cadieux-Dion, Maxime; Gauthier, Julie; Lapointe, Line; Mottron, Laurent; Valtorta, Flavia; Rouleau, Guy A.; Fassio, Anna; Benfenati, Fabio; Cossette, Patrick

2014-01-01

An increasing number of genes predisposing to autism spectrum disorders (ASDs) has been identified, many of which are implicated in synaptic function. This ‘synaptic autism pathway’ notably includes disruption of SYN1 that is associated with epilepsy, autism and abnormal behavior in both human and mice models. Synapsins constitute a multigene family of neuron-specific phosphoproteins (SYN1-3) present in the majority of synapses where they are implicated in the regulation of neurotransmitter release and synaptogenesis. Synapsins I and II, the major Syn isoforms in the adult brain, display partially overlapping functions and defects in both isoforms are associated with epilepsy and autistic-like behavior in mice. In this study, we show that nonsense (A94fs199X) and missense (Y236S and G464R) mutations in SYN2 are associated with ASD in humans. The phenotype is apparent in males. Female carriers of SYN2 mutations are unaffected, suggesting that SYN2 is another example of autosomal sex-limited expression in ASD. When expressed in SYN2  knockout neurons, wild-type human Syn II fully rescues the SYN2 knockout phenotype, whereas the nonsense mutant is not expressed and the missense mutants are virtually unable to modify the SYN2 knockout phenotype. These results identify for the first time SYN2  as a novel predisposing gene for ASD and strengthen the hypothesis that a disturbance of synaptic homeostasis underlies ASD. PMID:23956174

Combined Active Humoral and Cellular Immunization Approaches for the Treatment of Synucleinopathies.

PubMed

Rockenstein, Edward; Ostroff, Gary; Dikengil, Fusun; Rus, Florentina; Mante, Michael; Florio, Jazmin; Adame, Anthony; Trinh, Ivy; Kim, Changyoun; Overk, Cassia; Masliah, Eliezer; Rissman, Robert A

2018-01-24

Dementia with Lewy bodies, Parkinson's disease, and Multiple System Atrophy are age-related neurodegenerative disorders characterized by progressive accumulation of α-synuclein (α-syn) and jointly termed synucleinopathies. Currently, no disease-modifying treatments are available for these disorders. Previous preclinical studies demonstrate that active and passive immunizations targeting α-syn partially ameliorate behavioral deficits and α-syn accumulation; however, it is unknown whether combining humoral and cellular immunization might act synergistically to reduce inflammation and improve microglial-mediated α-syn clearance. Since combined delivery of antigen plus rapamycin (RAP) in nanoparticles is known to induce antigen-specific regulatory T cells (Tregs), we adapted this approach to α-syn using the antigen-presenting cell-targeting glucan microparticle (GP) vaccine delivery system. PDGF-α-syn transgenic (tg) male and female mice were immunized with GP-alone, GP-α-syn (active humoral immunization), GP+RAP, or GP+RAP/α-syn (combined active humoral and Treg) and analyzed using neuropathological and biochemical markers. Active immunization resulted in higher serological total IgG, IgG1, and IgG2a anti-α-syn levels. Compared with mice immunized with GP-alone or GP-α-syn, mice vaccinated with GP+RAP or GP+RAP/α-syn displayed increased numbers of CD25-, FoxP3-, and CD4-positive cells in the CNS. GP-α-syn or GP+RAP/α-syn immunizations resulted in a 30-45% reduction in α-syn accumulation, neuroinflammation, and neurodegeneration. Mice immunized with GP+RAP/α-syn further rescued neurons and reduced neuroinflammation. Levels of TGF-β1 were increased with GP+RAP/α-syn immunization, while levels of TNF-α and IL-6 were reduced. We conclude that the observed effects of GP+RAP/α-syn immunization support the hypothesis that cellular immunization may enhance the effects of active immunotherapy for the treatment of synucleinopathies. SIGNIFICANCE STATEMENT We show that a novel vaccination modality combining an antigen-presenting cell-targeting glucan particle (GP) vaccine delivery system with encapsulated antigen (α-synuclein) + rapamycin (RAP) induced both strong anti-α-synuclein antibody titers and regulatory T cells (Tregs). This vaccine, collectively termed GP+RAP/α-syn, is capable of triggering neuroprotective Treg responses in synucleinopathy models, and the combined vaccine is more effective than the humoral or cellular immunization alone. Together, these results support the further development of this multifunctional vaccine approach for the treatment of synucleinopathies, such as Parkinson's disease, dementia with Lewy bodies, and multiple systems atrophy. Copyright © 2018 the authors 0270-6474/18/381000-15$15.00/0.

Conformational analysis of some N,N-diethyl-2-[(4‧-substituted) phenylthio] acetamides

NASA Astrophysics Data System (ADS)

Vinhato, Elisângela; Olivato, Paulo R.; Zukerman-Schpector, Julio; Dal Colle, Maurizio

2013-11-01

The conformational analysis of some N,N-diethyl-2[(4‧-substituted)phenylthio]acetamides bearing the substituents OMe 1, Me 2, H 3, Cl 4, Br 5 and NO26, was performed by νCO IR analysis, along with B3LYP/6-311++G(d,p) and Polarisable Continuum Model (PCM) calculations, as well as NBO analysis for 1, 3, and 6 and X-ray diffraction for 4. The results of the calculations indicated the existence of two stable conformation pairs, i.e. gauche (anti; syn) (most stable) and cis (anti; syn) in the gas phase. The gauche conformers were less polar with respect to the cis ones for 1 and 3, but more polar for 6. The most intense IR carbonyl doublet component observed at the lower frequency can be ascribed to the gauche conformers g(anti; syn) for 3-6 in n-C6H14, which is in agreement with the gauche and cis relative stabilities and frequencies resulting from the PCM calculations. Similarly, the single IR band for 1 and 2 in n-hexane may be attributed to the gauche conformers. The PCM calculations compared well with the IR data for the compounds in solution, showing that there is a progressive increase of the cis/gauche population ratio as the solvent polarity increases. The NBO analysis indicated that the gauche(anti; syn) conformation in the gas phase was stabilized by the relevant LPS4 → πC 2dbnd O 1∗, πC 2dbnd O 1 → σC 3sbnd S 4∗ , σC 3sbnd S 4 → πC 2dbnd O 1∗, πC 2dbnd O 1∗ → σC 3sbnd S 4∗, and LPO1 →σ∗ C11sbnd H28 orbital interactions, which were absent in the cis(anti; syn) conformer. On the contrary, the cis conformer for derivatives 1, 3, and 6 were stabilized by the σC 3 -S 4∗ →σ∗ C2sbnd N5 orbital interaction (through bond coupling), along with the additional LPO1 →σ∗ S4sbnd C10 interaction for 6. Moreover, the electrostatic repulsion between the Cδ+sbnd Sδ- and Cδ+dbnd Oδ- dipoles (Repulsive Field Effect) contributed to both the larger destabilization and increase of the νCO frequency of the cis conformer with respect to the gauche conformer. X-ray single crystal analysis indicates that compound 4 assumes the c2(anti) conformation in the solid state, which is the conformation obtained by compound 6 in the gas phase. To obtain the largest energy gain, the molecules were arranged in the crystal in a six-molecules synthon mediated by Csbnd H⋯O and Cl⋯Cl interactions, where the chlorine atoms were related by a crystallographic inversion center.

A Protein Aggregation Inhibitor, Leuco-Methylthioninium Bis(Hydromethanesulfonate), Decreases α-Synuclein Inclusions in a Transgenic Mouse Model of Synucleinopathy

PubMed Central

Schwab, Karima; Frahm, Silke; Horsley, David; Rickard, Janet E.; Melis, Valeria; Goatman, Elizabeth A.; Magbagbeolu, Mandy; Douglas, Morag; Leith, Michael G.; Baddeley, Thomas C.; Storey, John M. D.; Riedel, Gernot; Wischik, Claude M.; Harrington, Charles R.; Theuring, Franz

2018-01-01

α-Synuclein (α-Syn) aggregation is a pathological feature of synucleinopathies, neurodegenerative disorders that include Parkinson’s disease (PD). We have tested whether N,N,N′,N′-tetramethyl-10H-phenothiazine-3,7-diaminium bis(hydromethanesulfonate) (leuco-methylthioninium bis(hydromethanesulfonate); LMTM), a tau aggregation inhibitor, affects α-Syn aggregation in vitro and in vivo. Both cellular and transgenic models in which the expression of full-length human α-Syn (h-α-Syn) fused with a signal sequence peptide to promote α-Syn aggregation were used. Aggregated α-Syn was observed following differentiation of N1E-115 neuroblastoma cells transfected with h-α-Syn. The appearance of aggregated α-Syn was inhibited by LMTM, with an EC50 of 1.1 μM, with minimal effect on h-α-Syn mRNA levels being observed. Two independent lines of mice (L58 and L62) transgenic for the same fusion protein accumulated neuronal h-α-Syn that, with aging, developed into fibrillary inclusions characterized by both resistance to proteinase K (PK)-cleavage and their ability to bind thiazin red. There was a significant decrease in α-Syn-positive neurons in multiple brain regions following oral treatment of male and female mice with LMTM administered daily for 6 weeks at 5 and 15 mg MT/kg. The early aggregates of α-Syn and the late-stage fibrillar inclusions were both susceptible to inhibition by LMTM, a treatment that also resulted in the rescue of movement and anxiety-related traits in these mice. The results suggest that LMTM may provide a potential disease modification therapy in PD and other synucleinopathies through the inhibition of α-Syn aggregation. PMID:29375308

A Focus on the Beneficial Effects of Alpha Synuclein and a Re-Appraisal of Synucleinopathies

PubMed Central

Ryskalin, Larisa; Busceti, Carla L.; Limanaqi, Fiona; Biagioni, Francesca; Gambardella, Stefano; Fornai, Francesco

2018-01-01

Alpha synuclein (α-syn) belongs to a class of proteins which are commonly considered to play a detrimental role in neuronal survival. This assumption is based on the occurrence of a severe neuronal degeneration in patients carrying a multiplication of the α-syn gene (SNCA) and in a variety of experi-mental models, where overexpression of α-syn leads to cell death and neurological impairment. In these conditions, a higher amount of normally structured α-syn produces a damage, which is even worse com-pared with that produced by α-syn owning an abnormal structure (as occurring following point gene muta-tions). In line with this, knocking out the expression of α-syn is reported to protect from specific neurotox-ins such as 1-methyl, 4-phenyl 1,2,3,6-tetrahydropyridine (MPTP). In the present review we briefly dis-cuss these well-known detrimental effects but we focus on findings showing that, in specific conditions α-syn is beneficial for cell survival. This occurs during methamphetamine intoxication which is counteracted by endogenous α-syn. Similarly, the dysfunction of the chaperone cysteine-string protein-alpha leads to cell pathology which is counteracted by over-expressing α-syn. In line with this, an increased expression of α-syn protects against oxidative damage produced by dopamine. Remarkably, when the lack of α-syn is combined with a depletion of β- and γ- synucleins, alterations in brain structure and function occur. This review tries to balance the evidence showing a beneficial effect with the bulk of data reporting a detri-mental effect of endogenous α-syn. The specific role of α-syn as a chaperone protein is discussed to ex-plain such a dual effect. PMID:29150919

A Focus on the Beneficial Effects of Alpha Synuclein and a Re-Appraisal of Synucleinopathies.

PubMed

Ryskalin, Larisa; Busceti, Carla L; Limanaqi, Fiona; Biagioni, Francesca; Gambardella, Stefano; Fornai, Francesco

2018-01-01

Alpha synuclein (α-syn) belongs to a class of proteins which are commonly considered to play a detrimental role in neuronal survival. This assumption is based on the occurrence of a severe neuronal degeneration in patients carrying a multiplication of the α-syn gene (SNCA) and in a variety of experimental models, where overexpression of α-syn leads to cell death and neurological impairment. In these conditions, a higher amount of normally structured α-syn produces a damage, which is even worse compared with that produced by α-syn owning an abnormal structure (as occurring following point gene mutations). In line with this, knocking out the expression of α-syn is reported to protect from specific neurotoxins such as 1-methyl, 4-phenyl 1,2,3,6-tetrahydropyridine (MPTP). In the present review we briefly discuss these well-known detrimental effects but we focus on findings showing that, in specific conditions α-syn is beneficial for cell survival. This occurs during methamphetamine intoxication which is counteracted by endogenous α-syn. Similarly, the dysfunction of the chaperone cysteine-string protein- alpha leads to cell pathology which is counteracted by over-expressing α-syn. In line with this, an increased expression of α-syn protects against oxidative damage produced by dopamine. Remarkably, when the lack of α-syn is combined with a depletion of β- and γ- synucleins, alterations in brain structure and function occur. This review tries to balance the evidence showing a beneficial effect with the bulk of data reporting a detrimental effect of endogenous α-syn. The specific role of α-syn as a chaperone protein is discussed to explain such a dual effect. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Neuroprotective and nootropic drug noopept rescues α-synuclein amyloid cytotoxicity.

PubMed

Jia, Xueen; Gharibyan, Anna L; Öhman, Anders; Liu, Yonggang; Olofsson, Anders; Morozova-Roche, Ludmilla A

2011-12-16

Parkinson's disease is a common neurodegenerative disorder characterized by α-synuclein (α-Syn)-containing Lewy body formation and selective loss of dopaminergic neurons in the substantia nigra. We have demonstrated the modulating effect of noopept, a novel proline-containing dipeptide drug with nootropic and neuroprotective properties, on α-Syn oligomerization and fibrillation by using thioflavin T fluorescence, far-UV CD, and atomic force microscopy techniques. Noopept does not bind to a sterically specific site in the α-Syn molecule as revealed by heteronuclear two-dimensional NMR analysis, but due to hydrophobic interactions with toxic amyloid oligomers, it prompts their rapid sequestration into larger fibrillar amyloid aggregates. Consequently, this process rescues the cytotoxic effect of amyloid oligomers on neuroblastoma SH-SY5Y cells as demonstrated by using cell viability assays and fluorescent staining of apoptotic and necrotic cells and by assessing the level of intracellular oxidative stress. The mitigating effect of noopept against amyloid oligomeric cytotoxicity may offer additional benefits to the already well-established therapeutic functions of this new pharmaceutical. Copyright © 2011 Elsevier Ltd. All rights reserved.

Far-Infrared Spectroscopy of Syn-Vinyl Alcohol

NASA Astrophysics Data System (ADS)

Raston, Paul; Bunn, Hayley

2016-06-01

Vinyl alcohol has been extensively studied in both the microwave and mid-IR spectral regions, where 9 out of 15 vibrational modes have been identified. Here we present the first far-IR spectrum of vinyl alcohol, collected below 700 wn at the Australian Synchrotron. The high resolution (0.001 wn) spectrum reveals the νb{11} and νb{15} fundamentals of syn-vinyl alcohol at 489 wn and 407 wn, in addition to two hot bands of the νb{15} mode at 369 wn and 323 wn. High J transitions in the R-branch of the νb{15} band were found to be perturbed by an a-axis Coriolis interaction with the nearby νb{11} state. The νb{15} torsional mode of syn-vinyl alcohol was fit using a Watson's A-reduced Hamiltonian to yield rotational, centrifugal distortion, and Coriolis coupling parameters. S. Saito, Chem. Phys. Lett. 42, 3 (1976) M. Rodler et al., J. Am. Chem. Soc. 106, 4029 (1948) Y. Koga et al., J. Mol. Spec. 145, 315 (1991) D-L. Joo et al., J. Mol. Spec. 197, 68 (1999)

New systematic assignments in Gonyleptoidea (Arachnida, Opiliones, Laniatores)

PubMed Central

Pinto-da-Rocha, Ricardo; Benedetti, Alípio Rezende; de Vasconcelos, Eduardo Gomes; Hara, Marcos Ryotaro

2012-01-01

Abstract As part of an ongoing revision of the family Gonyleptidae, we have identified many species that are synonyms of previously described species or misplaced in this family. This article summarizes these findings, adding previously unavailable information or correcting imprecise observations to justify the presented taxonomic changes. The following new familial or subfamilial assignments are proposed: Nemastygnus Roewer, 1929 and Taulisa Roewer, 1956 are transferred to Agoristenidae, Agoristeninae; Napostygnus Roewer, 1929 to Cranaidae; Ceropachylinus peruvianus Roewer, 1956 and Pirunipygus Roewer, 1936 are transferred to Gonyleptidae, Ampycinae; Gyndesops Roewer, 1943, Haversia Roewer, 1913 and Oxapampeus Roewer, 1963 are transferred to Gonyleptidae, Pachylinae. The following generic synonymies are proposed for the family Gonyleptidae: Acanthogonyleptes Mello-Leitão, 1922 = Centroleptes Roewer, 1943; Acrographinotus Roewer, 1929 = Unduavius Roewer, 1929; Gonyleptes Kirby, 1819 = Collonychium Bertkau, 1880; Mischonyx Bertkau, 1880 = Eugonyleptes Roewer, 1913 and Gonazula Roewer, 1930; Parampheres Roewer, 1913 = Metapachyloides Roewer, 1917; Pseudopucrolia Roewer, 1912 = Meteusarcus Roewer, 1913; Haversia Roewer, 1913 = Hoggellula Roewer, 1930. The following specific synonymies are proposed for the family Gonyleptidae: Acanthogonyleptes singularis (Mello-Leitão, 1935) = Centroleptes flavus Roewer, 1943, syn. n.; Geraeocormobius sylvarum Holmberg, 1887 = Discocyrtus serrifemur Roewer, 1943, syn. n.; Gonyleptellus bimaculatus (Sørensen, 1884) = Gonyleptes cancellatus Roewer,1917, syn. n.; Gonyleptes atrus Mello-Leitão, 1923 = Weyhia brieni Giltay, 1928, syn. n.; Gonyleptes fragilis Mello-Leitão, 1923 = Gonyleptes banana Kury, 2003, syn. n.; Gonyleptes horridus Kirby, 1819 = Collonychium bicuspidatum Bertkau, 1880, syn. n., Gonyleptes borgmeyeri Mello-Leitão, 1932, syn. n., Gonyleptes curvicornis Mello-Leitão, 1932, syn. n., Metagonyleptes hamatus Roewer, 1913, syn. n. and Paragonyleptes simoni Roewer, 1930, syn. n.; Gonyleptes pustulatus Sørensen, 1884 = Gonyleptes guttatus Roewer, 1917, syn. n.; Haversia defensa (Butler, 1876) = Sadocus vallentini Hogg, 1913, syn. n.; Liogonyleptoides minensis (Piza, 1946) = Currala bahiensis Soares, 1972, syn. n.; Megapachylus grandis Roewer, 1913 = Metapachyloides almeidai Soares & Soares, 1946, syn. n.; Mischonyx cuspidatus (Roewer, 1913) = Gonazula gibbosa Roewer, 1930 syn. n.; Mischonyx scaber (Kirby, 1819) = Xundarava holacantha Mello-Leitão, 1927, syn. n.; Parampheres tibialis Roewer, 1917 = Metapachyloides rugosus Roewer, 1917, syn. n.; Parapachyloides uncinatus (Sørensen, 1879) = Goyazella armata Mello-Leitão, 1931, syn. n.; Pseudopucrolia mutica (Perty, 1833) = Meteusarcus armatus Roewer, 1913, syn. n. The following new combinations are proposed: Acrographinotus ornatus (Roewer, 1929), comb. n. (ex Unduavius); Gonyleptellus bimaculatus (Sørensen, 1884),comb. n. (ex Gonyleptes);Gonyleptes perlatus (Mello-Leitão, 1935), comb. n. (exMoojenia);Mischonyx scaber (Kirby, 1819), comb. n. (ex Gonyleptes); and Neopachyloides peruvianus (Roewer, 1956), comb. n. (ex Ceropachylus). The following species of Gonyleptidae, Gonyleptinae are revalidated: Gonyleptes atrus Mello-Leitão, 1923 and Gonyleptes curvicornis (Roewer, 1913). PMID:22707905

ABSINTH: A new continuum solvation model for simulations of polypeptides in aqueous solutions

PubMed Central

Vitalis, Andreas; Pappu, Rohit V.

2009-01-01

A new implicit solvation model for use in Monte Carlo simulations of polypeptides is introduced. The model is termed ABSINTH for self-Assembly of Biomolecules Studied by an Implicit, Novel, and Tunable Hamiltonian. It is designed primarily for simulating conformational equilibria and oligomerization reactions of intrinsically disordered proteins in aqueous solutions. The paradigm for ABSINTH is conceptually similar to the EEF1 model of Lazaridis and Karplus (Proteins: Struct. Func. Genet., 1999, 35: 133-152). In ABSINTH, the transfer of a polypeptide solute from the gas phase into a continuum solvent is the sum of a direct mean field interaction (DMFI), and a term to model the screening of polar interactions. Polypeptide solutes are decomposed into a set of distinct solvation groups. The DMFI is a sum of contributions from each of the solvation groups, which are analogs of model compounds. Continuum-mediated screening of electrostatic interactions is achieved using a framework similar to the one used for the DMFI. Promising results are shown for a set of test cases. These include the calculation of NMR coupling constants for short peptides, the assessment of the thermal stability of two small proteins, reversible folding of both an alpha-helix and a beta-hairpin forming peptide, and the polymeric properties of intrinsically disordered polyglutamine peptides of varying lengths. The tests reveal that the computational expense for simulations with the ABSINTH implicit solvation model increase by a factor that is in the range of 2.5-5.0 with respect to gas-phase calculations. PMID:18506808

α-Synuclein Mutation Inhibits Endocytosis at Mammalian Central Nerve Terminals.

PubMed

Xu, Jianhua; Wu, Xin-Sheng; Sheng, Jiansong; Zhang, Zhen; Yue, Hai-Yuan; Sun, Lixin; Sgobio, Carmelo; Lin, Xian; Peng, Shiyong; Jin, Yinghui; Gan, Lin; Cai, Huaibin; Wu, Ling-Gang

2016-04-20

α-Synuclein (α-syn) missense and multiplication mutations have been suggested to cause neurodegenerative diseases, including Parkinson's disease (PD) and dementia with Lewy bodies. Before causing the progressive neuronal loss, α-syn mutations impair exocytosis, which may contribute to eventual neurodegeneration. To understand how α-syn mutations impair exocytosis, we developed a mouse model that selectively expressed PD-related human α-syn A53T (h-α-synA53T) mutation at the calyx of Held terminals, where release mechanisms can be dissected with a patch-clamping technique. With capacitance measurement of endocytosis, we reported that h-α-synA53T, either expressed transgenically or dialyzed in the short term in calyces, inhibited two of the most common forms of endocytosis, the slow and rapid vesicle endocytosis at mammalian central synapses. The expression of h-α-synA53Tin calyces also inhibited vesicle replenishment to the readily releasable pool. These findings may help to understand how α-syn mutations impair neurotransmission before neurodegeneration. α-Synuclein (α-syn) missense or multiplication mutations may cause neurodegenerative diseases, such as Parkinson's disease and dementia with Lewy bodies. The initial impact of α-syn mutations before neuronal loss is impairment of exocytosis, which may contribute to eventual neurodegeneration. The mechanism underlying impairment of exocytosis is poorly understood. Here we report that an α-syn mutant, the human α-syn A53T, inhibited two of the most commonly observed forms of endocytosis, slow and rapid endocytosis, at a mammalian central synapse. We also found that α-syn A53T inhibited vesicle replenishment to the readily releasable pool. These results may contribute to accounting for the widely observed early synaptic impairment caused by α-syn mutations in the progression toward neurodegeneration. Copyright © 2016 the authors 0270-6474/16/364408-07$15.00/0.

Redox reactions of the α-synuclein-Cu(2+) complex and their effects on neuronal cell viability.

PubMed

Wang, Chengshan; Liu, Lin; Zhang, Lin; Peng, Yong; Zhou, Feimeng

2010-09-21

α-Synuclein (α-syn), a presynaptic protein believed to play an important role in neuropathology in Parkinson's disease (PD), is known to bind Cu(2+). Cu(2+) has been shown to accelerate the aggregation of α-syn to form various toxic aggregates in vitro. Copper is also a redox-active metal whose complexes with amyloidogenic proteins/peptides have been linked to oxidative stress in major neurodegenerative diseases. In this work, the formation of the Cu(2+) complex with α-syn or with an N-terminal peptide, α-syn(1-19), was confirmed with electrospray-mass spectrometry (ES-MS). The redox potentials of the Cu(2+) complex with α-syn (α-syn-Cu(2+)) and α-syn(1-19) were determined to be 0.018 and 0.053 V, respectively. Furthermore, the Cu(2+) center(s) can be readily reduced to Cu(+), and possible reactions of α-syn-Cu(2+) with cellular species (e.g., O(2), ascorbic acid, and dopamine) were investigated. The occurrence of a redox reaction can be rationalized by comparing the redox potential of the α-syn-Cu(2+) complex to that of the specific cellular species. For example, ascorbic acid can directly reduce α-syn-Cu(2+) to α-syn-Cu(+), setting up a redox cycle in which O(2) is reduced to H(2)O(2) and cellular redox species is continuously exhausted. In addition, the H(2)O(2) generated was demonstrated to reduce viability of the neuroblastoma SY-HY5Y cells. Although our results ruled out the direct oxidation of dopamine by α-syn-Cu(2+), the H(2)O(2) generated in the presence of α-syn-Cu(2+) can oxidize dopamine. Our results suggest that oxidative stress is at least partially responsible for the loss of dopaminergic cells in PD brain and reveal the multifaceted role of the α-syn-Cu(2+) complex in oxidative stress associated with PD symptoms.

The nuclear accumulation of alpha-synuclein is mediated by importin alpha and promotes neurotoxicity by accelerating the cell cycle.

PubMed

Ma, Kai-Li; Song, Lian-Kun; Yuan, Yu-He; Zhang, Ying; Han, Ning; Gao, Kai; Chen, Nai-Hong

2014-07-01

α-Synuclein (α-syn), a 14 kDa pre-synaptic protein, is widely involved in the Parkinson's disease (PD) pathogenesis. Recent studies have shown that the nuclear accumulation of α-syn might have a toxic effect. The main purpose of the present study was to explore which amino acid residues in α-syn are associated with its nuclear accumulation, the molecule(s) mediated the nuclear import of α-syn, and the role of α-syn accumulated in the nucleus. It has been noted that the nuclear import of α-syn may be mediated by importin α and that both the amino acid residues 1-60 and 103-140 of α-syn were indispensable for its nuclear import. After imported into the nucleus, the accumulated α-syn played a toxic role in both the PC12 cells and the C57 mice. Furthermore, α-syn-nuclear localization signal-injected mice showed behavioral symptoms associated with PD. Further studies performed in vitro showed that the toxicity of α-syn in the nucleus might be due to an interference of the cell cycle. Thus, it can be concluded that α-syn can accumulate in nucleus, which is mediated by importin α, and promote neurotoxicity by accelerating the cell cycle. Copyright © 2013 Elsevier Ltd. All rights reserved.

Stereospecific synthesis of syn-α-oximinoamides by a three-component reaction of isocyanides, syn-chlorooximes, and carboxylic acids.

PubMed

Pirali, Tracey; Mossetti, Riccardo; Galli, Simona; Tron, Gian Cesare

2011-07-15

A stereospecific multicomponent reaction among isocyanides, syn-chlorooximes, and carboxylic acids provides an efficient synthesis of biologically relevant syn-α-oximinoamides. © 2011 American Chemical Society

Categorizing Biases in High-Confidence High-Throughput Protein-Protein Interaction Data Sets

DTIC Science & Technology

2011-01-01

may partially explain why we did not observe any of the interactions between RNA polymerase II compo- nents in any of the Y2H set (11). Methodological...DNA. Fig. 5 shows that RNA syn- thesis complexes formed a highly interconnected cluster, in- cluding RNA polymerases I, II , and III, Transcription...factor complexes II F (TFIIF) and III C (TFIIIC), which were connected via direct protein-protein interactions with many other func- tional complexes. Fig

CSF tau and β-amyloid predict cerebral synucleinopathy in autopsied Lewy body disorders.

PubMed

Irwin, David J; Xie, Sharon X; Coughlin, David; Nevler, Naomi; Akhtar, Rizwan S; McMillan, Corey T; Lee, Edward B; Wolk, David A; Weintraub, Daniel; Chen-Plotkin, Alice; Duda, John E; Spindler, Meredith; Siderowf, Andrew; Hurtig, Howard I; Shaw, Leslie M; Grossman, Murray; Trojanowski, John Q

2018-03-20

To test the association of antemortem CSF biomarkers with postmortem pathology in Lewy body disorders (LBD). Patients with autopsy-confirmed LBD (n = 24) and autopsy-confirmed Alzheimer disease (AD) (n = 23) and cognitively normal (n = 36) controls were studied. In LBD, neuropathologic criteria defined Lewy body α-synuclein (SYN) stages with medium/high AD copathology (SYN + AD = 10) and low/no AD copathology (SYN - AD = 14). Ordinal pathology scores for tau, β-amyloid (Aβ), and SYN pathology were averaged across 7 cortical regions to obtain a global cerebral score for each pathology. CSF total tau (t-tau), phosphorylated tau at threonine 181 , and Aβ 1-42 levels were compared between LBD and control groups and correlated with global cerebral pathology scores in LBD with linear regression. Diagnostic accuracy for postmortem categorization of LBD into SYN + AD vs SYN - AD or neocortical vs brainstem/limbic SYN stage was tested with receiver operating curves. SYN + AD had higher CSF t-tau (mean difference 27.0 ± 8.6 pg/mL) and lower Aβ 1-42 (mean difference -84.0 ± 22.9 g/mL) compared to SYN - AD ( p < 0.01, both). Increasing global cerebral tau and plaque scores were associated with higher CSF t-tau ( R 2 = 0.15-0.16, p < 0.05, both) and lower Aβ 1-42 ( R 2 = 0.43-0.49, p < 0.001, both), while increasing cerebral SYN scores were associated with lower CSF Aβ 1-42 ( R 2 = 0.31, p < 0.001) and higher CSF t-tau/Aβ 1-42 ratio ( R 2 = 0.27, p = 0.01). CSF t-tau/Aβ 1-42 ratio had 100% specificity and 90% sensitivity for SYN + AD, and CSF Aβ 1-42 had 77% specificity and 82% sensitivity for neocortical SYN stage. Higher antemortem CSF t-tau/Aβ 1-42 and lower Aβ 1-42 levels are predictive of increasing cerebral AD and SYN pathology. These biomarkers may identify patients with LBD vulnerable to cortical SYN pathology who may benefit from both SYN and AD-targeted disease-modifying therapies. © 2018 American Academy of Neurology.

Generic revision of the ant subfamily Dorylinae (Hymenoptera, Formicidae)

PubMed Central

Borowiec, Marek L.

2016-01-01

Abstract The generic classification of the ant subfamily Dorylinae is revised, with the aim of facilitating identification of easily-diagnosable monophyletic genera. The new classification is based on recent molecular phylogenetic evidence and a critical reappraisal of doryline morphology. New keys and diagnoses based on workers and males are provided, along with reviews of natural history and phylogenetic relationships, distribution maps, and a list of valid species for each lineage. Twenty-eight genera (27 extant and 1 extinct) are recognized within the subfamily, an increase from 20 in the previous classification scheme. Species classified in the polyphyletic Cerapachys and Sphinctomyrmex prior to this publication are here distributed among 9 and 3 different genera, respectively. Amyrmex and Asphinctanilloides are synonymized under Leptanilloides and the currently recognized subgenera are synonymized for Dorylus. No tribal classification is proposed for the subfamily, but several apparently monophyletic genus-groups are discussed. Valid generic names recognized here include: Acanthostichus (= Ctenopyga), Aenictogiton, Aenictus (= Paraenictus, Typhlatta), Cerapachys (= Ceratopachys), Cheliomyrmex, Chrysapace gen. rev., Cylindromyrmex (= Holcoponera, Hypocylindromyrmex, Metacylindromyrmex), Dorylus (= Alaopone syn. n., Anomma syn. n., Cosmaecetes, Dichthadia syn. n., Rhogmus syn. n., Shuckardia, Sphecomyrmex, Sphegomyrmex, Typhlopone syn. n.), Eburopone gen. n., Eciton (= Camptognatha, Holopone, Mayromyrmex), Eusphinctus gen. rev., Labidus (= Nycteresia, Pseudodichthadia), Leptanilloides (= Amyrmex syn. n., Asphinctanilloides syn. n.), Lioponera gen. rev. (= Neophyracaces syn. n., Phyracaces syn. n.), Lividopone, Neivamyrmex (= Acamatus, Woitkowskia), Neocerapachys gen. n., Nomamyrmex, Ooceraea gen. rev. (= Cysias syn. n.), Parasyscia gen. rev., †Procerapachys, Simopone, Sphinctomyrmex, Syscia gen. rev., Tanipone, Vicinopone, Yunodorylus gen. rev., Zasphinctus gen. rev. (= Aethiopopone syn. n., Nothosphinctus syn. n.). PMID:27559303

A pH- and temperature-responsive bioresorbable injectable hydrogel based on polypeptide block copolymers for the sustained delivery of proteins in vivo.

PubMed

Turabee, Md Hasan; Thambi, Thavasyappan; Duong, Huu Thuy Trang; Jeong, Ji Hoon; Lee, Doo Sung

2018-02-27

Sustained delivery of protein therapeutics is limited owing to the fragile nature of proteins. Despite its great potential, delivery of proteins without any loss of bioactivity remains a challenge in the use of protein therapeutics in the clinic. To surmount this shortcoming, we report a pH- and temperature-responsive in situ-forming injectable hydrogel based on comb-type polypeptide block copolymers for the controlled delivery of proteins. Polypeptide block copolymers, composed of hydrophilic polyethylene glycol (PEG), temperature-responsive poly(γ-benzyl-l-glutamate) (PBLG), and pH-responsive oligo(sulfamethazine) (OSM), exhibit pH- and temperature-induced sol-to-gel transition behavior in aqueous solutions. Polypeptide block copolymers were synthesized by combining N-carboxyanhydride-based ring-opening polymerization and post-functionalization of the chain-end using N-hydroxy succinimide ester activated OSM. The physical properties of polypeptide-based hydrogels were tuned by varying the composition of temperature- and pH-responsive PBLG and OSM in block copolymers. Polypeptide block copolymers were non-toxic to human embryonic kidney cells at high concentrations (2000 μg mL -1 ). Subcutaneous administration of polypeptide block copolymer sols formed viscoelastic gel instantly at the back of Sprague-Dawley (SD) rats. The in vivo gels exhibited sustained degradation and were found to be bioresorbable in 6 weeks without any noticeable inflammation at the injection site. Anionic characteristics of hydrogels allow efficient loading of a cationic model protein, lysozyme, through electrostatic interaction. Lysozyme-loaded polypeptide block copolymer sols readily formed a viscoelastic gel in vivo and sustained lysozyme release for at least a week. Overall, the results demonstrate an elegant approach to control the release of certain charged proteins and open a myriad of therapeutic possibilities in protein therapeutics.

α-Synuclein transfer between neurons and astrocytes indicates that astrocytes play a role in degradation rather than in spreading.

PubMed

Loria, Frida; Vargas, Jessica Y; Bousset, Luc; Syan, Sylvie; Salles, Audrey; Melki, Ronald; Zurzolo, Chiara

2017-11-01

Recent evidence suggests that disease progression in Parkinson's disease (PD) could occur by the spreading of α-synuclein (α-syn) aggregates between neurons. Here we studied the role of astrocytes in the intercellular transfer and fate of α-syn fibrils, using in vitro and ex vivo models. α-Syn fibrils can be transferred to neighboring cells; however, the transfer efficiency changes depending on the cell types. We found that α-syn is efficiently transferred from astrocytes to astrocytes and from neurons to astrocytes, but less efficiently from astrocytes to neurons. Interestingly, α-syn puncta are mainly found inside the lysosomal compartments of the recipient cells. However, differently from neurons, astrocytes are able to efficiently degrade fibrillar α-syn, suggesting an active role for these cells in clearing α-syn deposits. Astrocytes co-cultured with organotypic brain slices are able to take up α-syn fibrils from the slices. Altogether our data support a role for astrocytes in trapping and clearing α-syn pathological deposits in PD.

Structural and functional properties of prefibrillar α-synuclein oligomers.

PubMed

Pieri, Laura; Madiona, Karine; Melki, Ronald

2016-04-14

The deposition of fibrillar alpha-synuclein (α-syn) within inclusions (Lewy bodies and Lewy neurites) in neurons and glial cells is a hallmark of synucleinopathies. α-syn populates a variety of assemblies ranging from prefibrillar oligomeric species to fibrils whose specific contribution to neurodegeneration is still unclear. Here, we compare the specific structural and biological properties of distinct soluble prefibrillar α-syn oligomers formed either spontaneously or in the presence of dopamine and glutaraldehyde. We show that both on-fibrillar assembly pathway and distinct dopamine-mediated and glutaraldehyde-cross-linked α-syn oligomers are only slightly effective in perturbing cell membrane integrity and inducing cytotoxicity, while mature fibrils exhibit the highest toxicity. In contrast to low-molecular weight and unstable oligomers, large stable α-syn oligomers seed the aggregation of soluble α-syn within reporter cells although to a lesser extent than mature α-syn fibrils. These oligomers appear elongated in shape. Our findings suggest that α-syn oligomers represent a continuum of species ranging from unstable low molecular weight particles to mature fibrils via stable elongated oligomers composed of more than 15 α-syn monomers that possess seeding capacity.

α-Synuclein stimulation of monoamine oxidase-B and legumain protease mediates the pathology of Parkinson's disease.

PubMed

Kang, Seong Su; Ahn, Eun Hee; Zhang, Zhentao; Liu, Xia; Manfredsson, Fredric P; Sandoval, Ivette M; Dhakal, Susov; Iuvone, P Michael; Cao, Xuebing; Ye, Keqiang

2018-06-15

Dopaminergic neurodegeneration in Parkinson's disease (PD) is associated with abnormal dopamine metabolism by MAO-B (monoamine oxidase-B) and intracellular α-Synuclein (α-Syn) aggregates, called the Lewy body. However, the molecular relationship between α-Syn and MAO-B remains unclear. Here, we show that α-Syn directly binds to MAO-B and stimulates its enzymatic activity, which triggers AEP (asparagine endopeptidase; legumain) activation and subsequent α-Syn cleavage at N103, leading to dopaminergic neurodegeneration. Interestingly, the dopamine metabolite, DOPAL, strongly activates AEP, and the N103 fragment of α-Syn binds and activates MAO-B. Accordingly, overexpression of AEP in SNCA transgenic mice elicits α-Syn N103 cleavage and accelerates PD pathogenesis, and inhibition of MAO-B by Rasagiline diminishes α-Syn-mediated PD pathology and motor dysfunction. Moreover, virally mediated expression of α-Syn N103 induces PD pathogenesis in wild-type, but not MAO-B-null mice. Our findings thus support that AEP-mediated cleavage of α-Syn at N103 is required for the association and activation of MAO-B, mediating PD pathogenesis. © 2018 The Authors.

Opposed Effects of Dityrosine Formation in Soluble and Aggregated α-Synuclein on Fibril Growth.

PubMed

Wördehoff, Michael M; Shaykhalishahi, Hamed; Groß, Luca; Gremer, Lothar; Stoldt, Matthias; Buell, Alexander K; Willbold, Dieter; Hoyer, Wolfgang

2017-10-13

Parkinson's disease is the second most common neurodegenerative disease. It is characterized by aggregation of the protein α-synuclein (α-syn) in Lewy bodies, mitochondrial dysfunction, and increased oxidative stress in the substantia nigra. Oxidative stress leads to several modifications of biomolecules including dityrosine (DiY) crosslinking in proteins, which has recently been detected in α-syn in Lewy bodies from Parkinson's disease patients. Here we report that α-syn is highly susceptible to ultraviolet-induced DiY formation. We investigated DiY formation of α-syn and nine tyrosine-to-alanine mutants and monitored its effect on α-syn fibril formation in vitro. Ultraviolet irradiation of intrinsically disordered α-syn generates DiY-modified monomers and dimers, which inhibit fibril formation of unmodified α-syn by interfering with fibril elongation. The inhibition depends on both the DiY group and its integration into α-syn. When preformed α-syn fibrils are crosslinked by DiY formation, they gain increased resistance to denaturation. DiY-stabilized α-syn fibrils retain their high seeding efficiency even after being exposed to denaturant concentrations that completely depolymerize non-crosslinked seeds. Oxidative stress-associated DiY crosslinking of α-syn therefore entails two opposing effects: (i) inhibition of aggregation by DiY-modified monomers and dimers, and (ii) stabilization of fibrillar aggregates against potential degradation mechanisms, which can lead to promotion of aggregation, especially in the presence of secondary nucleation. Copyright © 2017 The Author(s). Published by Elsevier Ltd.. All rights reserved.

Coenzyme Q supplementation or over-expression of the yeast Coq8 putative kinase stabilizes multi-subunit Coq polypeptide complexes in yeast coq null mutants.

PubMed

He, Cuiwen H; Xie, Letian X; Allan, Christopher M; Tran, Uyenphuong C; Clarke, Catherine F

2014-04-04

Coenzyme Q biosynthesis in yeast requires a multi-subunit Coq polypeptide complex. Deletion of any one of the COQ genes leads to respiratory deficiency and decreased levels of the Coq4, Coq6, Coq7, and Coq9 polypeptides, suggesting that their association in a high molecular mass complex is required for stability. Over-expression of the putative Coq8 kinase in certain coq null mutants restores steady-state levels of the sensitive Coq polypeptides and promotes the synthesis of late-stage Q-intermediates. Here we show that over-expression of Coq8 in yeast coq null mutants profoundly affects the association of several of the Coq polypeptides in high molecular mass complexes, as assayed by separation of digitonin extracts of mitochondria by two-dimensional blue-native/SDS PAGE. The Coq4 polypeptide persists at high molecular mass with over-expression of Coq8 in coq3, coq5, coq6, coq7, coq9, and coq10 mutants, indicating that Coq4 is a central organizer of the Coq complex. Supplementation with exogenous Q6 increased the steady-state levels of Coq4, Coq7, and Coq9, and several other mitochondrial polypeptides in select coq null mutants, and also promoted the formation of late-stage Q-intermediates. Q supplementation may stabilize this complex by interacting with one or more of the Coq polypeptides. The stabilizing effects of exogenously added Q6 or over-expression of Coq8 depend on Coq1 and Coq2 production of a polyisoprenyl intermediate. Based on the observed interdependence of the Coq polypeptides, the effect of exogenous Q6, and the requirement for an endogenously produced polyisoprenyl intermediate, we propose a new model for the Q-biosynthetic complex, termed the CoQ-synthome. Copyright © 2014 Elsevier B.V. All rights reserved.

Coenzyme Q supplementation or over-expression of the yeast Coq8 putative kinase stabilizes multi-subunit Coq polypeptide complexes in yeast coq null mutants*

PubMed Central

He, Cuiwen H.; Xie, Letian X.; Allan, Christopher M.; Tran, UyenPhuong C.; Clarke, Catherine F.

2014-01-01

Coenzyme Q biosynthesis in yeast requires a multi-subunit Coq polypeptide complex. Deletion of any one of the COQ genes leads to respiratory deficiency and decreased levels of the Coq4, Coq6, Coq7, and Coq9 polypeptides, suggesting that their association in a high molecular mass complex is required for stability. Over-expression of the putative Coq8 kinase in certain coq null mutants restores steady-state levels of the sensitive Coq polypeptides and promotes the synthesis of late-stage Q-intermediates. Here we show that over-expression of Coq8 in yeast coq null mutants profoundly affects the association of several of the Coq polypeptides in high molecular mass complexes, as assayed by separation of digitonin extracts of mitochondria by two-dimensional blue-native/SDS PAGE. The Coq4 polypeptide persists at high molecular mass with over-expression of Coq8 in coq3, coq5, coq6, coq7, coq9, and coq10 mutants, indicating that Coq4 is a central organizer of the Coq complex. Supplementation with exogenous Q6 increased the steady-state levels of Coq4, Coq7, Coq9, and several other mitochondrial polypeptides in select coq null mutants, and also promoted the formation of late-stage Q-intermediates. Q supplementation may stabilize this complex by interacting with one or more of the Coq polypeptides. The stabilizing effects of exogenously added Q6 or over-expression of Coq8 depend on Coq1 and Coq2 production of a polyisoprenyl intermediate. Based on the observed interdependence of the Coq polypeptides, the effect of exogenous Q6, and the requirement for an endogenously produced polyisoprenyl intermediate, we propose a new model for the Q-biosynthetic complex, termed the CoQ-synthome. PMID:24406904

The regulation of catalase activity by PPAR γ is affected by α-synuclein

PubMed Central

Yakunin, Eugenia; Kisos, Haya; Kulik, Willem; Grigoletto, Jessica; Wanders, Ronald J A; Sharon, Ronit

2014-01-01

Objective While evidence for oxidative injury is frequently detected in brains of humans affected by Parkinson's disease (PD) and in relevant animal models, there is uncertainty regarding its cause. We tested the potential role of catalase in the oxidative injury that characterizes PD. Methods Utilizing brains of A53T α-Syn and ntg mice, and cultured cells, we analyzed catalase activity and expression, and performed biochemical analyses of peroxisomal metabolites. Results Lower catalase expression and lower activity levels were detected in A53T α-Syn brains and α-Syn-expressing cells. The effect on catalase activity was independent of disease progression, represented by mouse age and α-Syn mutation, suggesting a potential physiological function for α-Syn. Notably, catalase activity and expression were unaffected in brains of mice modeling Alzheimer's disease. Moreover, we found that α-Syn expression downregulate the peroxisome proliferator-activated receptor (PPAR)γ, which controls catalase transcription. Importantly, activation of either PPARγ2, PPARα or retinoic X receptor eliminated the inhibiting effect of α-Syn on catalase activity. In addition, activation of these nuclear receptors enhanced the accumulation of soluble α-Syn oligomers, resulting in a positive association between the degree of soluble α-Syn oligomers and catalase activity. Of note, a comprehensive biochemical analysis of specific peroxisomal metabolites indicated no signs of dysfunction in specific peroxisomal activities in brains of A53T α-Syn mice. Interpretation Our results suggest that α-Syn expression may interfere with the complex and overlapping network of nuclear receptors transcription activation. In result, catalase activity is affected through mechanisms involved in the regulation of soluble α-Syn oligomers. PMID:25356396

Fluorescence probe of polypeptide conformational dynamics in gas phase and in solution

NASA Astrophysics Data System (ADS)

Iavarone, Anthony T.; Meinen, Jan; Schulze, Susanne; Parks, Joel H.

2006-07-01

Fluorescence measurements of polypeptides derivatized with the fluorescent dye BODIPY TMR have been used to probe the polypeptide conformational dynamics as a function of temperature and charge state. Measurements of (BODIPY TMR)-[Pro]n-Arg-Trp and (BODIPY TMR)-[Gly-Ser]m-Arg-Trp have been performed for charge states 1+ and 2+ of n = 4 and 10 and m = 2 and 5. The 2+ charge states of both of these polypeptides exhibit similar temperature dependences for equal chain lengths (n = 4, m = 2 and n = 10, m = 5) and suggest conformations dominated by Coulomb repulsion. In the absence of such Coulomb repulsion, the 1+ charge state conformations appear to be characterized by the flexibility of the polypeptide chain for which [Gly-Ser]m > [Pro]n. Comparisons of these gas phase polypeptide measurements with corresponding measurements in solution provide a direct measure of the effects of solvent on the conformational dynamics. The change in fluorescence as a function of temperature in the gas phase is two orders of magnitude greater than that in solution, a dramatic result we attribute to the restrictions on intramolecular dynamics imposed by diffusion-limited kinetics and the lack of shielding by solvent. Measurements were also made of unsolvated Pron peptides without the tryptophan (Trp) residue to isolate the interaction of the fluorescent dye with charges.

Revision of the fungus-farming ant genus Sericomyrmex Mayr (Hymenoptera, Formicidae, Myrmicinae)

PubMed Central

Ješovnik, Ana; Schultz, Ted R.

2017-01-01

Abstract The genus Sericomyrmex Mayr (Formicidae: Myrmicinae: Attini) is a Neotropical group of fungus-farming ants known for its problematic taxonomy, caused by low morphological variability across the species, vague and old species descriptions, and an outdated and incomplete key published in 1916. Recent molecular studies revealed that Sericomyrmex is the product of a rapid recent radiation, with a divergence date of 4.3 million years ago. Here we present a comprehensive taxonomic revision of the genus Sericomyrmex based on morphology and a recently published molecular phylogeny. We discuss and illustrate morphological characters for Sericomyrmex workers, males, queens, and larvae. We report 18 standard morphological measurements and 5 indices for 529 workers, 50 queens, and 39 males, which we employ in morphometric analyses. The revised genus Sericomyrmex comprises eleven species, including three new species, here described as S. maravalhas sp. n., S. radioheadi sp. n., and S. saramama sp. n. We also redescribe S. amabilis Wheeler, S. bondari Borgmeier, S. lutzi Wheeler, S. mayri Forel, S. opacus Mayr, S. parvulus Forel, S. saussurei Emery, and S. scrobifer Forel. The number of recognized species (11) is lower than the previously recognized 19 species and 3 subspecies. The following species and subspecies are synonymized: under S. opacus [=S. aztecus Forel syn. n., S. zacapanus Wheeler syn. n., and S. diego Forel syn. n.]; under S. bondari [=S. beniensis Weber syn. n.]; under S. mayri [=S. luederwaldti Santschi syn. n., S. moreirai Santschi syn. n., S. harekulli Weber syn. n., S. harekulli arawakensis Weber syn. n., S. urichi Forel syn. n.]; under S. saussurei [=S. burchelli Forel syn. n., S. impexus Wheeler syn. n., S. urichi maracas Weber syn. n.]; and under S. parvulus [=S. myersi Weber syn. n.]. We provide a key to Sericomyrmex species for the worker caste and information on the geographic distributions of all species. PMID:28769657

Silencing Alpha Synuclein in Mature Nigral Neurons Results in Rapid Neuroinflammation and Subsequent Toxicity

PubMed Central

Benskey, Matthew J.; Sellnow, Rhyomi C.; Sandoval, Ivette M.; Sortwell, Caryl E.; Lipton, Jack W.; Manfredsson, Fredric P.

2018-01-01

Human studies and preclinical models of Parkinson’s disease implicate the involvement of both the innate and adaptive immune systems in disease progression. Further, pro-inflammatory markers are highly enriched near neurons containing pathological forms of alpha synuclein (α-syn), and α-syn overexpression recapitulates neuroinflammatory changes in models of Parkinson’s disease. These data suggest that α-syn may initiate a pathological inflammatory response, however the mechanism by which α-syn initiates neuroinflammation is poorly understood. Silencing endogenous α-syn results in a similar pattern of nigral degeneration observed following α-syn overexpression. Here we aimed to test the hypothesis that loss of α-syn function within nigrostriatal neurons results in neuronal dysfunction, which subsequently stimulates neuroinflammation. Adeno-associated virus (AAV) expressing an short hairpin RNA (shRNA) targeting endogenous α-syn was unilaterally injected into the substantia nigra pars compacta (SNc) of adult rats, after which nigrostriatal pathology and indices of neuroinflammation were examined at 7, 10, 14 and 21 days post-surgery. Removing endogenous α-syn from nigrostriatal neurons resulted in a rapid up-regulation of the major histocompatibility complex class 1 (MHC-1) within transduced nigral neurons. Nigral MHC-1 expression occurred prior to any overt cell death and coincided with the recruitment of reactive microglia and T-cells to affected neurons. Following the induction of neuroinflammation, α-syn knockdown resulted in a 50% loss of nigrostriatal neurons in the SNc and a corresponding loss of nigrostriatal terminals and dopamine (DA) concentrations within the striatum. Expression of a control shRNA did not elicit any pathological changes. Silencing α-syn within glutamatergic neurons of the cerebellum did not elicit inflammation or cell death, suggesting that toxicity initiated by α-syn silencing is specific to DA neurons. These data provide evidence that loss of α-syn function within nigrostriatal neurons initiates a neuronal-mediated neuroinflammatory cascade, involving both the innate and adaptive immune systems, which ultimately results in the death of affected neurons. PMID:29497361

Passive immunization reduces behavioral and neuropathological deficits in an alpha-synuclein transgenic model of Lewy body disease.

PubMed

Masliah, Eliezer; Rockenstein, Edward; Mante, Michael; Crews, Leslie; Spencer, Brian; Adame, Anthony; Patrick, Christina; Trejo, Margarita; Ubhi, Kiren; Rohn, Troy T; Mueller-Steiner, Sarah; Seubert, Peter; Barbour, Robin; McConlogue, Lisa; Buttini, Manuel; Games, Dora; Schenk, Dale

2011-04-29

Dementia with Lewy bodies (DLB) and Parkinson's Disease (PD) are common causes of motor and cognitive deficits and are associated with the abnormal accumulation of alpha-synuclein (α-syn). This study investigated whether passive immunization with a novel monoclonal α-syn antibody (9E4) against the C-terminus (CT) of α-syn was able to cross into the CNS and ameliorate the deficits associated with α-syn accumulation. In this study we demonstrate that 9E4 was effective at reducing behavioral deficits in the water maze, moreover, immunization with 9E4 reduced the accumulation of calpain-cleaved α-syn in axons and synapses and the associated neurodegenerative deficits. In vivo studies demonstrated that 9E4 traffics into the CNS, binds to cells that display α-syn accumulation and promotes α-syn clearance via the lysosomal pathway. These results suggest that passive immunization with monoclonal antibodies against the CT of α-syn may be of therapeutic relevance in patients with PD and DLB.

Hydration and conformational mechanics of single, end-tethered elastin-like polypeptides.

PubMed

Valiaev, Alexei; Lim, Dong Woo; Schmidler, Scott; Clark, Robert L; Chilkoti, Ashutosh; Zauscher, Stefan

2008-08-20

We investigated the effect of temperature, ionic strength, solvent polarity, and type of guest residue on the force-extension behavior of single, end-tethered elastin-like polypeptides (ELPs), using single molecule force spectroscopy (SMFS). ELPs are stimulus-responsive polypeptides that contain repeats of the five amino acids Val-Pro-Gly-Xaa-Gly (VPGXG), where Xaa is a guest residue that can be any amino acid with the exception of proline. We fitted the force-extension data with a freely jointed chain (FJC) model which allowed us to resolve small differences in the effective Kuhn segment length distributions that largely arise from differences in the hydrophobic hydration behavior of ELP. Our results agree qualitatively with predictions from recent molecular dynamics simulations and demonstrate that hydrophobic hydration modulates the molecular elasticity for ELPs. Furthermore, our results show that SMFS, when combined with our approach for data analysis, can be used to study the subtleties of polypeptide-water interactions and thus provides a basis for the study of hydrophobic hydration in intrinsically unstructured biomacromolecules.

Structural and functional properties of prefibrillar α-synuclein oligomers

PubMed Central

Pieri, Laura; Madiona, Karine; Melki, Ronald

2016-01-01

The deposition of fibrillar alpha-synuclein (α-syn) within inclusions (Lewy bodies and Lewy neurites) in neurons and glial cells is a hallmark of synucleinopathies. α-syn populates a variety of assemblies ranging from prefibrillar oligomeric species to fibrils whose specific contribution to neurodegeneration is still unclear. Here, we compare the specific structural and biological properties of distinct soluble prefibrillar α-syn oligomers formed either spontaneously or in the presence of dopamine and glutaraldehyde. We show that both on-fibrillar assembly pathway and distinct dopamine-mediated and glutaraldehyde-cross-linked α-syn oligomers are only slightly effective in perturbing cell membrane integrity and inducing cytotoxicity, while mature fibrils exhibit the highest toxicity. In contrast to low-molecular weight and unstable oligomers, large stable α-syn oligomers seed the aggregation of soluble α-syn within reporter cells although to a lesser extent than mature α-syn fibrils. These oligomers appear elongated in shape. Our findings suggest that α-syn oligomers represent a continuum of species ranging from unstable low molecular weight particles to mature fibrils via stable elongated oligomers composed of more than 15 α-syn monomers that possess seeding capacity. PMID:27075649

Registration and Release of Syn1RR tall fescue

USDA-ARS?s Scientific Manuscript database

The Agricultural Research Service of the United States DepaRRment of Agriculture announces the release of the new tall fescue [Festuca arundinacea (syn., Lolium arundinaceum Darbyshire; Schedonorus phoenix (Scop.) Holub)] cultivar Syn1RR. Syn1RR is a rust tolerant tall fescue cultivar that exhibits...

Sodium butyrate rescues dopaminergic cells from alpha-synuclein-induced transcriptional deregulation and DNA damage.

PubMed

Paiva, Isabel; Pinho, Raquel; Pavlou, Maria Angeliki; Hennion, Magali; Wales, Pauline; Schütz, Anna-Lena; Rajput, Ashish; Szego, Éva M; Kerimoglu, Cemil; Gerhardt, Ellen; Rego, Ana Cristina; Fischer, André; Bonn, Stefan; Outeiro, Tiago F

2017-06-15

Alpha-synuclein (aSyn) is considered a major culprit in Parkinson's disease (PD) pathophysiology. However, the precise molecular function of the protein remains elusive. Recent evidence suggests that aSyn may play a role on transcription regulation, possibly by modulating the acetylation status of histones. Our study aimed at evaluating the impact of wild-type (WT) and mutant A30P aSyn on gene expression, in a dopaminergic neuronal cell model, and decipher potential mechanisms underlying aSyn-mediated transcriptional deregulation. We performed gene expression analysis using RNA-sequencing in Lund Human Mesencephalic (LUHMES) cells expressing endogenous (control) or increased levels of WT or A30P aSyn. Compared to control cells, cells expressing both aSyn variants exhibited robust changes in the expression of several genes, including downregulation of major genes involved in DNA repair. WT aSyn, unlike A30P aSyn, promoted DNA damage and increased levels of phosphorylated p53. In dopaminergic neuronal cells, increased aSyn expression led to reduced levels of acetylated histone 3. Importantly, treatment with sodium butyrate, a histone deacetylase inhibitor (HDACi), rescued WT aSyn-induced DNA damage, possibly via upregulation of genes involved in DNA repair. Overall, our findings provide novel and compelling insight into the mechanisms associated with aSyn neurotoxicity in dopaminergic cells, which could be ameliorated with an HDACi. Future studies will be crucial to further validate these findings and to define novel possible targets for intervention in PD. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Energy transport in the three coupled α-polypeptide chains of collagen molecule with long-range interactions effect

NASA Astrophysics Data System (ADS)

Mvogo, Alain; Ben-Bolie, G. H.; Kofané, T. C.

2015-06-01

The dynamics of three coupled α-polypeptide chains of a collagen molecule is investigated with the influence of power-law long-range exciton-exciton interactions. The continuum limit of the discrete equations reveal that the collagen dynamics is governed by a set of three coupled nonlinear Schrödinger equations, whose dispersive coefficient depends on the LRI parameter r. We construct the analytic symmetric and asymmetric (antisymmetric) soliton solutions, which match with the structural features of collagen related with the acupuncture channels. These solutions are used as initial conditions for the numerical simulations of the discrete equations, which reveal a coherent transport of energy in the molecule for r > 3. The results also indicate that the width of the solitons is a decreasing function of r, which help to stabilize the solitons propagating in the molecule. To confirm further the efficiency of energy transport in the molecule, the modulational instability of the system is performed and the numerical simulations show that the energy can flow from one polypeptide chain to another in the form of nonlinear waves.

Leptotrombidium (Acari: Trombiculidae) of the World.

PubMed

Stekolnikov, Alexandr A

2013-01-01

The chigger mite genus Leptotrombidium Nagayo, Miyagawa, Mitamura and Imamura, 1916 is reviewed using literature data. For 340 larval species brief diagnoses, synonymy, data on type hosts and type localities are provided. The genus is divided into species-groups based on morphological evidence enabling easier establishment of group-membership of un-known specimens in the future. Some species groups are supported by a hierarchical cluster analysis with multiscale boot-strap resampling applied to a matrix including 335 species and geographic morphotypes and 19 standard quantitative characters. Six new species from mammalian hosts are described: L. aenigmami sp. nov., L. abramovi sp. nov., L. tikhon-ovi sp. nov., L. bochkovi sp. nov., L. laoense sp. nov., and L. megaloti sp. nov. from Laos. Seven names created by Ver-cammen-Grandjean and Langston (1976) for infrasubspecific entities are applied to species with the same descriptions: Leptotrombidium tenompaki sp. nov., L. kinabalui sp. nov., L. megabodense sp. nov., L. minului sp. nov., L. ului sp. nov., L. megalangati sp. nov., and L. saigoni sp. nov. A new replacement name is proposed: L. ushi nom. nov. pro L. hsui Wu, Yang and Li, 1999 (praeocc. Yu, Yang and Gong, 1986). Nineteen new synonyms and 7 new combinations are proposed: Leptotrombidium (= Hsuella Wang, Li and Shi, 1989, syn. nov.; = Leptotrombidium (Monosigmum) Wen, 2001, syn. nov.), L. deliense (Walch, 1922) (= L. deliense sinense Wen and Chen, 1984, syn. nov.; = L. deliense microsetosa Zhao, Tang and Mo, 1986, syn. nov.), L. sialkotense Vercammen-Grandjean and Langston, 1976 (= L. jishoum Wen, Li, Zhang and Liao, 1988, syn. nov.), L. imphalum Vercammen-Grandjean and Langston, 1976 (= L. imphalum sabahense Vercam-men-Grandjean and Langston, 1976, syn. nov.; = L. chiangraiensis Tanskul and Linthicum, 1997, syn. nov.), L. wenense Wu, Wen, Yang and Wu, 1982 (= L. kaohuense Li, Wang and Chen, 1997, syn. nov.), L. longimedian Brown, 1992 (= L. mindanensis Brown, 1992, syn. nov.), L. silvaticum Hushcha and Schluger, 1967 (= L. pakistanum Vercammen-Grandjean and Langston, 1976, syn. nov.), L. cricethrionis Wen, Sun and Sun, 1984 (= L. rusticum Yu, Yang and Gong, 1986, syn. nov.), L. intermedium (Nagayo, Mitamura and Tamiya, 1920) (= Trombicula (L.) daisen Kumada and Sasa, 1953, syn. nov.; = Trombicula hiranumai Kanda, 1942, syn. nov.), L. fletcheri (Womersley and Heaslip, 1943) (= L. fletcheri fran-colini Wen and Xiang, 1984b, syn. nov.), L. apertum Kudryashova, 1979 (= L. sorosi Kharadov, 1995, syn. nov.; = L. tolaicus Kharadov, 2000, syn. nov.), L. turdicola Vercammen-Grandjean and Langston, 1976 (= L. muntiaci Xiang and Wen, 1984d, syn. nov.; = L. suense Wen, 1984g, syn. nov.), L. paradux Vercammen-Grandjean and Langston, 1976 (= L. montanum Stekolnikov, 2004, syn. nov.), L. hubeiense (Wang, Li and Shi, 1989) comb. nov. from Hsuella, L. dunqingi (Liu, Xiang and Ma, 2003) comb. nov. from Hsuella, L. nainae (Kharadov, 1990) comb. nov. from Montivagum, L. mon-golicum (Kudryashova, 1988) comb. nov. from Montivagum, L. kunitzkyi (Kudryashova, 1988) comb. nov. from Monti-vagum, L. alaicum (Kharadov, 1994) comb. nov. from Montivagum, and Lorillatum nudisensillum (Yu, Gong and Tao, 1981) comb. nov. from Leptotrombidium. A key to Leptotrombidium species is provided.

Effects of Protein Structure on Iron–Polypeptide Vibrational Dynamic Coupling in Cytochrome c

DOE PAGES

Galinato, Mary Grace I.; Bowman, Sarah E. J.; Kleingardner, Jesse G.; ...

2014-12-22

Cytochrome c (Cyt c) has a heme covalently bound to the polypeptide via a Cys-X-X-Cys-His (CXXCH) linker that is located in the interface region for protein–protein interactions. To determine whether the polypeptide matrix influences iron vibrational dynamics, nuclear resonance vibrational spectroscopy (NRVS) measurements were performed on 57Fe-labeled ferric Hydrogenobacter thermophilus cytochrome c-552, and variants M13V, M13V/K22M, and A7F, which have structural modifications that alter the composition or environment of the CXXCH pentapeptide loop. Simulations of the NRVS data indicate that the 150–325 cm –1 region is dominated by N His–Fe–S Met axial ligand and polypeptide motions, while the 325–400 cmmore » –1 region shows dominant contributions from ν(Fe–N Pyr) (Pyr = pyrrole) and other heme-based modes. Diagnostic spectral signatures that directly relate to structural features of the heme active site are identified using a quantum chemistry-centered normal coordinate analysis (QCC-NCA). In particular, spectral features that directly correlate with CXXCH loop stiffness, the strength of the Fe–His interaction, and the degree of heme distortion are identified. Cumulative results from our investigation suggest that compared to the wild type (wt), variants M13V and M13V/K22M have a more rigid CXXCH pentapeptide segment, a stronger Fe–N His interaction, and a more ruffled heme. Conversely, the A7F variant has a more planar heme and a weaker Fe–NHis bond. These results are correlated to the observed changes in reduction potential between wt protein and the variants studied here. Lastly, we discuss the implications of these results for Cyt c biogenesis and electron transfer.« less

Effects of Protein Structure on Iron–Polypeptide Vibrational Dynamic Coupling in Cytochrome c

PubMed Central

2015-01-01

Cytochrome c (Cyt c) has a heme covalently bound to the polypeptide via a Cys-X-X-Cys-His (CXXCH) linker that is located in the interface region for protein–protein interactions. To determine whether the polypeptide matrix influences iron vibrational dynamics, nuclear resonance vibrational spectroscopy (NRVS) measurements were performed on 57Fe-labeled ferric Hydrogenobacter thermophilus cytochrome c-552, and variants M13V, M13V/K22M, and A7F, which have structural modifications that alter the composition or environment of the CXXCH pentapeptide loop. Simulations of the NRVS data indicate that the 150–325 cm–1 region is dominated by NHis–Fe–SMet axial ligand and polypeptide motions, while the 325–400 cm–1 region shows dominant contributions from ν(Fe–NPyr) (Pyr = pyrrole) and other heme-based modes. Diagnostic spectral signatures that directly relate to structural features of the heme active site are identified using a quantum chemistry-centered normal coordinate analysis (QCC-NCA). In particular, spectral features that directly correlate with CXXCH loop stiffness, the strength of the Fe–His interaction, and the degree of heme distortion are identified. Cumulative results from our investigation suggest that compared to the wild type (wt), variants M13V and M13V/K22M have a more rigid CXXCH pentapeptide segment, a stronger Fe–NHis interaction, and a more ruffled heme. Conversely, the A7F variant has a more planar heme and a weaker Fe–NHis bond. These results are correlated to the observed changes in reduction potential between wt protein and the variants studied here. Implications of these results for Cyt c biogenesis and electron transfer are also discussed. PMID:25531247

Synthetic biology: a utilitarian perspective.

PubMed

Smith, Kevin

2013-10-01

I examine the positive and negative features of synthetic biology ('SynBio') from a utilitarian ethical perspective. The potential beneficial outcomes from SynBio in the context of medicine are substantial; however it is not presently possible to predict precise outcomes due to the nascent state of the field. Potential negative outcomes from SynBio also exist, including iatrogenesis and bioterrorism; however it is not yet possible to quantify these risks. I argue that the application of a 'precautionary' approach to SynBio is ethically fraught, as is the notion that SynBio-associated knowledge ought to be restricted. I conclude that utilitarians ought to support a broadly laissez-faire stance in respect of SynBio. © 2013 John Wiley & Sons Ltd.

Studies on the structure of sphingomyelinase. Amino acid composition and heterogeneity on isoelectric focusing.

PubMed Central

Jones, C S; Shankaran, P; Davidson, D J; Poulos, A; Callahan, J W

1983-01-01

Sphingomyelinase, purified to apparent homogeneity from human placenta, is an acidic protein, as judged from its amino acid composition and by isoelectric focusing of the carboxymethylated protein. The amino acid composition is characterized by an approximately equal content of hydrophobic and polar amino acid residues. The reduced-alkylated polypeptides were separated into two groups. Most of the polypeptides were heterogeneous with pI values of 4.4-5.0, but an additional more minor component was observed at pI 5.4. Liquid isoelectric focusing resolved the purified enzyme into a single major component (pI 4.7-4.8), a minor component (pI 5.0-5.4) and a plateau region of activity (pI 6-7). On thin-layer isoelectric focusing, the protein profile obtained from each of these regions was the same. In addition, the substrate specificity, Km values and effect of inhibitory substances were identical. We conclude that sphingomyelinase is an acidic, microheterogeneous protein that likely exists as a holopolymer of a single major polypeptide chain. the heterogeneity of the intact protein on isoelectric focusing appears to reflect this microheterogeneity, which is influenced by a tendency to associate with itself and with detergents such as Triton X-100. Images Fig. 1. Fig. 2. Fig. 4. PMID:6303305

Aqueous cholesteric liquid crystals using uncharged rodlike polypeptides. Polypeptide vesicles by conformation-specific assembly. Ordered chiral macroporous hybrid silica-polypeptide composites

NASA Astrophysics Data System (ADS)

Bellomo, Enrico Giuseppe

2005-07-01

Aqueous cholesteric liquid crystals using uncharged rodlike polypeptides . The aqueous, lyotropic liquid-crystalline phase behavior of an alpha helical polypeptide, has been studied using optical microscopy and X-ray scattering. Solutions of optically pure polypeptide were found to form cholesteric liquid crystals at volume fractions that decreased with increasing average chain length. At very high volume fractions, the formation of a hexagonal mesophase was observed. The pitch of the cholesteric phase could be varied by a mixture of enantiomeric samples, where the pitch increased as the mixture approached equimolar. The cholesteric phases could be untwisted, using either magnetic field or shear flow, into nematic phases, which relaxed into cholesterics upon removal of field or shear. We have found that the phase diagram of this polypeptide in aqueous solution parallels that of poly(gamma-benzyl glutamate) in organic solvents, thus providing a useful system for liquid-crystal applications requiring water as solvent. Polypeptide vesicles by conformation-specific assembly. We have found that block copolymers composed of polypeptide segments provide significant advantages in controlling both the function and supramolecular structure of bioinspired self-assemblies. Incorporation of the stable chain conformations found in proteins into block copolymers was found to provide an additional element of control, beyond amphiphilicity and composition that defines self-assembled architecture. The abundance of functionality present in amino acids, and the ease by which they can be incorporated into these materials, also provides a powerful mechanism to impart block copolypeptides with function. This combination of structure and function work synergistically to enable significant advantages in the preparation of therapeutic agents as well as provide insight into design of self-assemblies beginning to approach the complexity of natural structures such as virus capsids. Ordered chiral macroporous hybrid silica-polypeptide composites. The mineralization of organic templates has been investigated as an effective way to control the size and structure of inorganic frameworks. Hybrid structures incorporating polypeptide with silica have been prepared and characterized using X-ray scattering, TGA, SEM and TEM. The results support the interaction between silica and polymer to form ordered chiral macroporous structures that can be easily controlled by polymer molecular weight and volume fraction.

Genome wide analysis of the transition to pathogenic lifestyles in Magnaporthales fungi

USDA-ARS?s Scientific Manuscript database

The rice blast fungus Pyricularia oryzae (syn. Magnaporthe oryzae, Magnaporthe grisea), a member of the order Magnaporthales in the class Sordariomycetes, is an important plant pathogen and a model species for studying pathogen infection and plant-fungal interaction. In this study, we generated geno...

Large α-synuclein oligomers inhibit neuronal SNARE-mediated vesicle docking

PubMed Central

Choi, Bong-Kyu; Choi, Mal-Gi; Kim, Jae-Yeol; Yang, Yoosoo; Lai, Ying; Kweon, Dae-Hyuk; Lee, Nam Ki; Shin, Yeon-Kyun

2013-01-01

Parkinson disease and dementia with Lewy bodies are featured with the formation of Lewy bodies composed mostly of α-synuclein (α-Syn) in the brain. Although evidence indicates that the large oligomeric or protofibril forms of α-Syn are neurotoxic agents, the detailed mechanisms of the toxic functions of the oligomers remain unclear. Here, we show that large α-Syn oligomers efficiently inhibit neuronal SNARE-mediated vesicle lipid mixing. Large α-Syn oligomers preferentially bind to the N-terminal domain of a vesicular SNARE protein, synaptobrevin-2, which blocks SNARE-mediated lipid mixing by preventing SNARE complex formation. In sharp contrast, the α-Syn monomer has a negligible effect on lipid mixing even with a 30-fold excess compared with the case of large α-Syn oligomers. Thus, the results suggest that large α-Syn oligomers function as inhibitors of dopamine release, which thus provides a clue, at the molecular level, to their neurotoxicity. PMID:23431141

Drug Targets from Genetics: Alpha-Synuclein

PubMed Central

Danzer, Karin M.; McLean, Pamela J.

2012-01-01

One of the critical issues in Parkinson disease (PD) research is the identity of the specific toxic, pathogenic moiety. In PD, mutations in alpha-synuclein (αsyn) or multiplication of the SNCA gene encoding αsyn, result in a phenotype of cellular inclusions, cell death, and brain dysfunction. While the historical point of view has been that the macroscopic aggregates containing αsyn are the toxic species, in the last several years evidence has emerged that suggests instead that smaller soluble species - likely oligomers containing misfolded αsyn - are actually the toxic moiety and that the fibrillar inclusions may even be a cellular detoxification pathway and less harmful. If soluble misfolded species of αsyn are the toxic moieties, then cellular mechanisms that degrade misfolded αsyn would be neuroprotective and a rational target for drug development. In this review we will discuss the fundamental mechanisms underlying αsyn toxicity including oligomer formation, oxidative stress, and degradation pathways and consider rational therapeutic strategies that may have the potential to prevent or halt αsyn induced pathogenesis in PD. PMID:21838671

Passive Immunization Reduces Behavioral and Neuropathological Deficits in an Alpha-Synuclein Transgenic Model of Lewy Body Disease

PubMed Central

Masliah, Eliezer; Rockenstein, Edward; Mante, Michael; Crews, Leslie; Spencer, Brian; Adame, Anthony; Patrick, Christina; Trejo, Margarita; Ubhi, Kiren; Rohn, Troy T.; Mueller-Steiner, Sarah; Seubert, Peter; Barbour, Robin; McConlogue, Lisa; Buttini, Manuel; Games, Dora; Schenk, Dale

2011-01-01

Dementia with Lewy bodies (DLB) and Parkinson's Disease (PD) are common causes of motor and cognitive deficits and are associated with the abnormal accumulation of alpha-synuclein (α-syn). This study investigated whether passive immunization with a novel monoclonal α-syn antibody (9E4) against the C-terminus (CT) of α-syn was able to cross into the CNS and ameliorate the deficits associated with α-syn accumulation. In this study we demonstrate that 9E4 was effective at reducing behavioral deficits in the water maze, moreover, immunization with 9E4 reduced the accumulation of calpain-cleaved α-syn in axons and synapses and the associated neurodegenerative deficits. In vivo studies demonstrated that 9E4 traffics into the CNS, binds to cells that display α-syn accumulation and promotes α-syn clearance via the lysosomal pathway. These results suggest that passive immunization with monoclonal antibodies against the CT of α-syn may be of therapeutic relevance in patients with PD and DLB. PMID:21559417

Cellular response of human neuroblastoma cells to α-synuclein fibrils, the main constituent of Lewy bodies.

PubMed

Pieri, Laura; Chafey, Philippe; Le Gall, Morgane; Clary, Guilhem; Melki, Ronald; Redeker, Virginie

2016-01-01

α-Synuclein (α-Syn) fibrils are the main constituent of Lewy bodies and a neuropathological hallmark of Parkinson's disease (PD). The propagation of α-Syn assemblies from cell to cell suggests that they are involved in PD progression. We previously showed that α-Syn fibrils are toxic because of their ability to bind and permeabilize cell membranes. Here, we document the cellular response in terms of proteome changes of SH-SY5Y cells exposed to exogenous α-Syn fibrils. We compare the proteomes of cells of neuronal origin exposed or not either to oligomeric or fibrillar α-Syn using two dimensional differential in-gel electrophoresis (2D-DIGE) and mass spectrometry. Only α-Syn fibrils induce significant changes in the proteome of SH-SY5Y cells. In addition to proteins associated to apoptosis and toxicity, or proteins previously linked to neurodegenerative diseases, we report an overexpression of proteins involved in intracellular vesicle trafficking. We also report a remarkable increase in fibrillar α-Syn heterogeneity, mainly due to C-terminal truncations. Our results show that cells of neuronal origin adapt their proteome to exogenous α-Syn fibrils and actively modify those assemblies. Cells of neuronal origin adapt their proteome to exogenous toxic α-Syn fibrils and actively modify those assemblies. Our results bring insights into the cellular response and clearance events the cells implement to face the propagation of α-Syn assemblies associated to pathology.

Birth, life, and demise of the Andean-syn-collisional Gissar arc: Late Paleozoic tectono-magmatic-metamorphic evolution of the southwestern Tian Shan, Tajikistan

NASA Astrophysics Data System (ADS)

Worthington, James R.; Kapp, Paul; Minaev, Vladislav; Chapman, James B.; Mazdab, Frank K.; Ducea, Mihai N.; Oimahmadov, Ilhomjon; Gadoev, Mustafo

2017-10-01

The amalgamation of the Central Asian Orogenic Belt in the southwestern Tian Shan in Tajikistan is represented by tectono-magmatic-metamorphic processes that accompanied late Paleozoic ocean closure and collision between the Karakum-Tarim and Kazakh-Kyrgyz terranes. Integrated U-Pb geochronology, thermobarometry, pseudosection modeling, and Hf geochemistry constrain the timing and petro-tectonic nature of these processes. The Gissar batholith and the Garm massif represent an eastward, along-strike increase in paleodepth from upper-batholith ( 21-7 km) to arc-root ( 36-19 km) levels of the Andean-syn-collisional Gissar arc, which developed from 323-288 Ma in two stages: (i) Andean, I-type granitoid magmatism from 323-306 Ma due to northward subduction of the Gissar back-arc ocean basin under the Gissar microcontinent, which was immediately followed by (ii) syn-collisional, I-S-type granitoid magmatism in the Gissar batholith and the Garm massif from 304-288 Ma due to northward subduction/underthrusting of Karakum marginal-continental crust under the Gissar microcontinent. A rapid isotopic pull-up from 288-286 Ma signals the onset of juvenile, alkaline-syenitic, post-collisional magmatism by 280 Ma, which was driven by delamination of the Gissar arclogite root and consequent convective asthenospheric upwelling. Whereas M-HT/LP prograde metamorphism in the Garm massif (650-750°C/6-7 kbar) from 310-288 Ma was associated with subduction-magma inundation and crustal thickening, HT/LP heating and decompression to peak-metamorphic temperatures ( 800-820°C/6-4 kbar) at 288 ± 6 Ma was driven by the transmission of a post-collisional, mantle-derived heat wave through the Garm-massif crust.

Nonclinical Safety Assessment of SYN-004: An Oral β-lactamase for the Protection of the Gut Microbiome From Disruption by Biliary-Excreted, Intravenously Administered Antibiotics.

PubMed

Kokai-Kun, John F; Bristol, J Andrew; Setser, John; Schlosser, Michael

2016-05-01

SYN-004 is a first in class, recombinant β-lactamase that degrades β-lactam antibiotics and has been formulated to be administered orally to patients receiving intravenous β-lactam antibiotics including cephalosporins. SYN-004 is intended to degrade unmetabolized antibiotics excreted into the intestines and thus has the potential to protect the gut microbiome from disruption by these antibiotics. Protection of the gut microbiome is expected to protect against opportunistic enteric infections such as Clostridium difficile infection as well as antibiotic-associated diarrhea. In order to demonstrate that oral SYN-004 is safe for human clinical trials, 2 Good Laboratory Practice-compliant toxicity studies were conducted in Beagle dogs. In both studies, SYN-004 was administered orally 3 times per day up to the maximum tolerated dose of the formulation. In the first study, doses of SYN-004 administered over 28 days were safe and well tolerated in dogs with the no-observed-adverse-effect level at the high dose of 57 mg/kg/day. Systemic absorption of SYN-004 was minimal and sporadic and showed no accumulation during the study. In the second study, doses up to 57 mg/kg/day were administered to dogs in combination with an intravenous dose of ceftriaxone (300 mg/kg) given once per day for 14 days. Coadministration of oral SYN-004 with intravenous ceftriaxone was safe and well tolerated, with SYN-004 having no noticeable effect on the plasma pharmacokinetics of ceftriaxone. These preclinical studies demonstrate that SYN-004 is well tolerated and, when coadministered with ceftriaxone, does not interfere with its systemic pharmacokinetics. These data supported advancing SYN-004 into human clinical trials. © The Author(s) 2015.

A sensitive assay reveals structural requirements for α-synuclein fibril growth

PubMed Central

Tsai, Christina; Bagchi, Devika P.; Engel, Laura A.; Sarezky, Jonathan; Kotzbauer, Paul T.

2017-01-01

The accumulation of α-synuclein (α-syn) fibrils in neuronal inclusions is the defining pathological process in Parkinson's disease (PD). A pathogenic role for α-syn fibril accumulation is supported by the identification of dominantly inherited α-syn (SNCA) gene mutations in rare cases of familial PD. Fibril formation involves a spontaneous nucleation event in which soluble α-syn monomers associate to form seeds, followed by fibril growth during which monomeric α-syn molecules sequentially associate with existing seeds. To better investigate this process, we developed sensitive assays that use the fluorescein arsenical dye FlAsH (fluorescein arsenical hairpin binder) to detect soluble oligomers and mature fibrils formed from recombinant α-syn protein containing an N-terminal bicysteine tag (C2-α-syn). Using seed growth by monomer association (SeGMA) assays to measure fibril growth over 3 h in the presence of C2-α-syn monomer, we observed that some familial PD-associated α-syn mutations (i.e. H50Q and A53T) greatly increased growth rates, whereas others (E46K, A30P, and G51D) decreased growth rates. Experiments with wild-type seeds extended by mutant monomer and vice versa revealed that single-amino acid differences between seed and monomer proteins consistently decreased growth rates. These results demonstrate that α-syn monomer association during fibril growth is a highly ordered process that can be disrupted by misalignment of individual amino acids and that only a subset of familial-PD mutations causes fibril accumulation through increased fibril growth rates. The SeGMA assays reported herein can be utilized to further elucidate structural requirements of α-syn fibril growth and to identify growth inhibitors as a potential therapeutic approach in PD. PMID:28373279

Mutual exacerbation of peroxisome proliferator-activated receptor γ coactivator 1α deregulation and α-synuclein oligomerization.

PubMed

Eschbach, Judith; von Einem, Björn; Müller, Kathrin; Bayer, Hanna; Scheffold, Annika; Morrison, Bradley E; Rudolph, K Lenhard; Thal, Dietmar R; Witting, Anke; Weydt, Patrick; Otto, Markus; Fauler, Michael; Liss, Birgit; McLean, Pamela J; Spada, Albert R La; Ludolph, Albert C; Weishaupt, Jochen H; Danzer, Karin M

2015-01-01

Aggregation of α-synuclein (α-syn) and α-syn cytotoxicity are hallmarks of sporadic and familial Parkinson disease (PD), with accumulating evidence that prefibrillar oligomers and protofibrils are the pathogenic species in PD and related synucleinopathies. Peroxisome proliferator-activated receptor γ coactivator 1α (PGC-1α), a key regulator of mitochondrial biogenesis and cellular energy metabolism, has recently been associated with the pathophysiology of PD. Despite extensive effort on studying the function of PGC-1α in mitochondria, no studies have addressed whether PGC-1α directly influences oligomerization of α-syn or whether α-syn oligomers impact PGC-1α expression. We tested whether pharmacological or genetic activation of PGC-1α or PGC-11α knockdown could modulate the oligomerization of α-syn in vitro by using an α-syn -fragment complementation assay. In this study, we found that both PGC-1α reference gene (RG-PGC-1α) and the central nervous system (CNS)-specific PGC-1α (CNS-PGC-1α) are downregulated in human PD brain, in A30P α-syn transgenic animals, and in a cell culture model for α-syn oligomerization. Importantly, downregulation of both RG-PGC-1α and CNS-PGC-1α in cell culture or neurons from RG-PGC-1α-deficient mice leads to a strong induction of α-syn oligomerization and toxicity. In contrast, pharmacological activation or genetic overexpression of RG-PGC-1α reduced α-syn oligomerization and rescued α-syn-mediated toxicity. Based on our results, we propose that PGC-1α downregulation and α-syn oligomerization form a vicious circle, thereby influencing and/or potentiating each other. Our data indicate that restoration of PGC-1α is a promising approach for development of effective drugs for the treatment of PD and related synucleinopathies. © 2014 American Neurological Association.

Syntaxin-4 mediates exocytosis of pre-docked and newcomer insulin granules underlying biphasic glucose-stimulated insulin secretion in human pancreatic beta cells.

PubMed

Xie, Li; Zhu, Dan; Dolai, Subhankar; Liang, Tao; Qin, Tairan; Kang, Youhou; Xie, Huanli; Huang, Ya-Chi; Gaisano, Herbert Y

2015-06-01

Of the four exocytotic syntaxins (Syns), much is now known about the role of Syn-1A (pre-docked secretory granules [SGs]) and Syn-3 (newcomer SGs) in insulin exocytosis. Some work was reported on Syn-4's role in biphasic glucose-stimulated insulin secretion (GSIS), but its precise role in insulin SG exocytosis remains unclear. In this paper we examine this role in human beta cells. Endogenous function of Syn-4 in human islets was assessed by knocking down its expression with lentiviral single hairpin RNA (lenti-shRNA)-RFP. Biphasic GSIS was determined by islet perifusion assay. Single-cell analysis of exocytosis of red fluorescent protein (RFP)-positive beta cells (exhibiting near-total depletion of Syn-4) was by patch clamp capacitance measurements (Cm) and total internal reflection fluorescence microscopy (TIRFM), the latter to further assess single SG behaviour. Co-immunoprecipitations were conducted on INS-1 cells to assess exocytotic complexes. Syn-4 knockdown (KD) of 77% in human islets caused a concomitant reduction in cognate Munc18c expression (46%) without affecting expression of other exocytotic proteins; this resulted in reduction of GSIS in the first phase (by 42%) and the second phase (by 40%). Cm of RFP-tagged Syn-4-KD beta cells showed severe inhibition in the readily releasable pool (by 71%) and mobilisation from reserve pools (by 63%). TIRFM showed that Syn-4-KD-induced inhibition of first-phase GSIS was attributed to reduction in exocytosis of both pre-docked and newcomer SGs (which undergo minimal residence or docking time at the plasma membrane before fusion). Second-phase inhibition was attributed to reduction in newcomer SGs. Stx-4 co-immunoprecipitated Munc18c, VAMP2 and VAMP8, suggesting that these exocytotic complexes may be involved in exocytosis of pre-docked and newcomer SGs. Syn-4 is involved in distinct molecular machineries that influence exocytosis of both pre-docked and newcomer SGs in a manner functionally redundant to Syn-1A and Syn-3, respectively; this underlies Syn-4's role in mediating portions of first-phase and second-phase GSIS.

C-Terminal Tyrosine Residue Modifications Modulate the Protective Phosphorylation of Serine 129 of α-Synuclein in a Yeast Model of Parkinson's Disease

PubMed Central

Lázaro, Diana F.; Pinho, Raquel; Valerius, Oliver; Outeiro, Tiago F.; Braus, Gerhard H.

2016-01-01

Parkinson´s disease (PD) is characterized by the presence of proteinaceous inclusions called Lewy bodies that are mainly composed of α-synuclein (αSyn). Elevated levels of oxidative or nitrative stresses have been implicated in αSyn related toxicity. Phosphorylation of αSyn on serine 129 (S129) modulates autophagic clearance of inclusions and is prominently found in Lewy bodies. The neighboring tyrosine residues Y125, Y133 and Y136 are phosphorylation and nitration sites. Using a yeast model of PD, we found that Y133 is required for protective S129 phosphorylation and for S129-independent proteasome clearance. αSyn can be nitrated and form stable covalent dimers originating from covalent crosslinking of two tyrosine residues. Nitrated tyrosine residues, but not di-tyrosine-crosslinked dimers, contributed to αSyn cytotoxicity and aggregation. Analysis of tyrosine residues involved in nitration and crosslinking revealed that the C-terminus, rather than the N-terminus of αSyn, is modified by nitration and di-tyrosine formation. The nitration level of wild-type αSyn was higher compared to that of A30P mutant that is non-toxic in yeast. A30P formed more dimers than wild-type αSyn, suggesting that dimer formation represents a cellular detoxification pathway in yeast. Deletion of the yeast flavohemoglobin gene YHB1 resulted in an increase of cellular nitrative stress and cytotoxicity leading to enhanced aggregation of A30P αSyn. Yhb1 protected yeast from A30P-induced mitochondrial fragmentation and peroxynitrite-induced nitrative stress. Strikingly, overexpression of neuroglobin, the human homolog of YHB1, protected against αSyn inclusion formation in mammalian cells. In total, our data suggest that C-terminal Y133 plays a major role in αSyn aggregate clearance by supporting the protective S129 phosphorylation for autophagy and by promoting proteasome clearance. C-terminal tyrosine nitration increases pathogenicity and can only be partially detoxified by αSyn di-tyrosine dimers. Our findings uncover a complex interplay between S129 phosphorylation and C-terminal tyrosine modifications of αSyn that likely participates in PD pathology. PMID:27341336

C-Terminal Tyrosine Residue Modifications Modulate the Protective Phosphorylation of Serine 129 of α-Synuclein in a Yeast Model of Parkinson's Disease.

PubMed

Kleinknecht, Alexandra; Popova, Blagovesta; Lázaro, Diana F; Pinho, Raquel; Valerius, Oliver; Outeiro, Tiago F; Braus, Gerhard H

2016-06-01

Parkinson´s disease (PD) is characterized by the presence of proteinaceous inclusions called Lewy bodies that are mainly composed of α-synuclein (αSyn). Elevated levels of oxidative or nitrative stresses have been implicated in αSyn related toxicity. Phosphorylation of αSyn on serine 129 (S129) modulates autophagic clearance of inclusions and is prominently found in Lewy bodies. The neighboring tyrosine residues Y125, Y133 and Y136 are phosphorylation and nitration sites. Using a yeast model of PD, we found that Y133 is required for protective S129 phosphorylation and for S129-independent proteasome clearance. αSyn can be nitrated and form stable covalent dimers originating from covalent crosslinking of two tyrosine residues. Nitrated tyrosine residues, but not di-tyrosine-crosslinked dimers, contributed to αSyn cytotoxicity and aggregation. Analysis of tyrosine residues involved in nitration and crosslinking revealed that the C-terminus, rather than the N-terminus of αSyn, is modified by nitration and di-tyrosine formation. The nitration level of wild-type αSyn was higher compared to that of A30P mutant that is non-toxic in yeast. A30P formed more dimers than wild-type αSyn, suggesting that dimer formation represents a cellular detoxification pathway in yeast. Deletion of the yeast flavohemoglobin gene YHB1 resulted in an increase of cellular nitrative stress and cytotoxicity leading to enhanced aggregation of A30P αSyn. Yhb1 protected yeast from A30P-induced mitochondrial fragmentation and peroxynitrite-induced nitrative stress. Strikingly, overexpression of neuroglobin, the human homolog of YHB1, protected against αSyn inclusion formation in mammalian cells. In total, our data suggest that C-terminal Y133 plays a major role in αSyn aggregate clearance by supporting the protective S129 phosphorylation for autophagy and by promoting proteasome clearance. C-terminal tyrosine nitration increases pathogenicity and can only be partially detoxified by αSyn di-tyrosine dimers. Our findings uncover a complex interplay between S129 phosphorylation and C-terminal tyrosine modifications of αSyn that likely participates in PD pathology.

The contribution of alpha synuclein to neuronal survival and function - Implications for Parkinson's disease.

PubMed

Benskey, Matthew J; Perez, Ruth G; Manfredsson, Fredric P

2016-05-01

The aggregation of alpha synuclein (α-syn) is a neuropathological feature that defines a spectrum of disorders collectively termed synucleinopathies, and of these, Parkinson's disease (PD) is arguably the best characterized. Aggregated α-syn is the primary component of Lewy bodies, the defining pathological feature of PD, while mutations or multiplications in the α-syn gene result in familial PD. The high correlation between α-syn burden and PD has led to the hypothesis that α-syn aggregation produces toxicity through a gain-of-function mechanism. However, α-syn has been implicated to function in a diverse range of essential cellular processes such as the regulation of neurotransmission and response to cellular stress. As such, an alternative hypothesis with equal explanatory power is that the aggregation of α-syn results in toxicity because of a toxic loss of necessary α-syn function, following sequestration of functional forms α-syn into insoluble protein aggregates. Within this review, we will provide an overview of the literature linking α-syn to PD and the knowledge gained from current α-syn-based animal models of PD. We will then interpret these data from the viewpoint of the α-syn loss-of-function hypothesis and provide a potential mechanistic model by which loss of α-syn function could result in at least some of the neurodegeneration observed in PD. By providing an alternative perspective on the etiopathogenesis of PD and synucleinopathies, this may reveal alternative avenues of research in order to identify potential novel therapeutic targets for disease modifying strategies. The correlation between α-synuclein burden and Parkinson's disease pathology has led to the hypothesis that α-synuclein aggregation produces toxicity through a gain-of-function mechanism. However, in this review, we discuss data supporting the alternative hypothesis that the aggregation of α-synuclein results in toxicity because of loss of necessary α-synuclein function at the presynaptic terminal, following sequestration of functional forms of α-synuclein into aggregates. © 2016 The Authors. Journal of Neurochemistry published by John Wiley & Sons Ltd on behalf of International Society for Neurochemistry.

Mutations in the C-terminal fragment of DnaK affecting peptide binding.

PubMed Central

Burkholder, W F; Zhao, X; Zhu, X; Hendrickson, W A; Gragerov, A; Gottesman, M E

1996-01-01

Escherichia coli DnaK acts as a molecular chaperone through its ATP-regulated binding and release of polypeptide substrates. Overexpressing a C-terminal fragment (CTF) of DnaK (Gly-384 to Lys-638) containing the polypeptide substrate binding domain is lethal in wild-type E. coli. This dominant-negative phenotype may result from the nonproductive binding of CTF to cellular polypeptide targets of DnaK. Mutations affecting DnaK substrate binding were identified by selecting noncytotoxic CTF mutants followed by in vitro screening. The clustering of such mutations in the three-dimensional structure of CTF suggests the model that loops L1,2 and L4,5 form a rigid core structure critical for interactions with substrate. Images Fig. 1 Fig. 2 Fig. 3 PMID:8855230

Conformational analysis of cellobiose by electronic structure theories.

PubMed

French, Alfred D; Johnson, Glenn P; Cramer, Christopher J; Csonka, Gábor I

2012-03-01

Adiabatic Φ/ψ maps for cellobiose were prepared with B3LYP density functional theory. A mixed basis set was used for minimization, followed with 6-31+G(d) single-point calculations, with and without SMD continuum solvation. Different arrangements of the exocyclic groups (38 starting geometries) were considered for each Φ/ψ point. The vacuum calculations agreed with earlier computational and experimental results on the preferred gas phase conformation (anti-Φ(H), syn-ψ(H)), and the results from the solvated calculations were consistent with the (syn Φ(H)/ψ(H) conformations from condensed phases (crystals or solutions). Results from related studies were compared, and there is substantial dependence on the solvation model as well as arrangements of exocyclic groups. New stabilizing interactions were revealed by Atoms-In-Molecules theory. Published by Elsevier Ltd.

Identification of immunogenic polypeptides from a Mycoplasma hyopneumoniae genome library by phage display.

PubMed

Kügler, Jonas; Nieswandt, Simone; Gerlach, Gerald F; Meens, Jochen; Schirrmann, Thomas; Hust, Michael

2008-09-01

The identification of immunogenic polypeptides of pathogens is helpful for the development of diagnostic assays and therapeutic applications like vaccines. Routinely, these proteins are identified by two-dimensional polyacrylamide gel electrophoresis and Western blot using convalescent serum, followed by mass spectrometry. This technology, however, is limited, because low or differentially expressed proteins, e.g. dependent on pathogen-host interaction, cannot be identified. In this work, we developed and improved a M13 genomic phage display-based method for the selection of immunogenic polypeptides of Mycoplasma hyopneumoniae, a pathogen causing porcine enzootic pneumonia. The fragmented genome of M. hyopneumoniae was cloned into a phage display vector, and the genomic library was packaged using the helperphage Hyperphage to enrich open reading frames (ORFs). Afterwards, the phage display library was screened by panning using convalescent serum. The analysis of individual phage clones resulted in the identification of five genes encoding immunogenic proteins, only two of which had been previously identified and described as immunogenic. This M13 genomic phage display, directly combining ORF enrichment and the presentation of the corresponding polypeptide on the phage surface, complements proteome-based methods for the identification of immunogenic polypeptides and is particularly well suited for the use in mycoplasma species.

Studies of protein aggregation in A53T α-synuclein transgenic, Tg2576 transgenic, and P246L presenilin-1 knock-in cross bred mice.

PubMed

Emmer, Kristel L; Covy, Jason P; Giasson, Benoit I

2012-01-24

Synucleinopathies are a group of neurodegenerative disorders, including Parkinson disease, associated with neuronal amyloid inclusions comprised of the presynaptic protein α-synuclein (α-syn); however the biological events that initiate and lead to the formation of these inclusions are still poorly understood. There is mounting evidence that intracellular α-syn aggregation may proceed via a seeding mechanism and could spread between neurons through a prion-like mechanism that may involve other amyloidogenic proteins. Several lines of evidence suggest that Aβ peptides and/or extracellular Aβ deposits may directly or indirectly promote intracellular α-syn aggregation. To assess the effects of Aβ peptides and extracellular Aβ deposits on α-syn aggregate formation, transgenic mice (line M83) expressing A53T human α-syn that are sensitive to developing α-syn pathological inclusions were cross bred to Tg2576 transgenic mice that generated elevated levels of Aβ peptides and develop abundant Aβ plaques. In addition these mice were bred to mice with the P264L presenilin-1 knock-in mutation that further promotes Aβ plaque formation. These mice demonstrated the expected formation of Aβ plaques; however despite the accumulation of hyperphosphorylated α-syn dystrophic neurites within or surrounding Aβ plaques, no additional α-syn pathologies were observed. These studies show that Aβ amyloid deposits can cause the local aggregation of α-syn, but these did not lead to more extensive α-syn pathology. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

Best Practices for Generating and Using Alpha-Synuclein Pre-Formed Fibrils to Model Parkinson’s Disease in Rodents

PubMed Central

Polinski, Nicole K.; Volpicelli-Daley, Laura A.; Sortwell, Caryl E.; Luk, Kelvin C.; Cremades, Nunilo; Gottler, Lindsey M.; Froula, Jessica; Duffy, Megan F.; Lee, Virginia M.Y.; Martinez, Terina N.; Dave, Kuldip D.

2018-01-01

Parkinson’s disease (PD) is the second most common neurodegenerative disease, affecting approximately one-percent of the population over the age of sixty. Although many animal models have been developed to study this disease, each model presents its own advantages and caveats. A unique model has arisen to study the role of alpha-synuclein (aSyn) in the pathogenesis of PD. This model involves the conversion of recombinant monomeric aSyn protein to a fibrillar form—the aSyn pre-formed fibril (aSyn PFF)—which is then injected into the brain or introduced to the media in culture. Although many groups have successfully adopted and replicated the aSyn PFF model, issues with generating consistent pathology have been reported by investigators. To improve the replicability of this model and diminish these issues, The Michael J. Fox Foundation for Parkinson’s Research (MJFF) has enlisted the help of field leaders who performed key experiments to establish the aSyn PFF model to provide the research community with guidelines and practical tips for improving the robustness and success of this model. Specifically, we identify key pitfalls and suggestions for avoiding these mistakes as they relate to generating the aSyn PFFs from monomeric protein, validating the formation of pathogenic aSyn PFFs, and using the aSyn PFFs in vivo or in vitro to model PD. With this additional information, adoption and use of the aSyn PFF model should present fewer challenges, resulting in a robust and widely available model of PD. PMID:29400668

Synapsins Are Downstream Players of the BDNF-Mediated Axonal Growth.

PubMed

Marte, Antonella; Messa, Mirko; Benfenati, Fabio; Onofri, Franco

2017-01-01

Synapsins (Syns) are synaptic vesicle-associated phosphoproteins involved in neuronal development and neurotransmitter release. While Syns are implicated in the regulation of brain-derived neurotrophic factor (BDNF)-induced neurotransmitter release, their role in the BDNF developmental effects has not been fully elucidated. By using primary cortical neurons from Syn I knockout (KO) and Syn I/II/III KO mice, we studied the effects of BDNF and nerve growth factor (NGF) on axonal growth. While NGF had similar effects in all genotypes, BDNF induced significant differences in Syn KO axonal outgrowth compared to wild type (WT), an effect that was rescued by the re-expression of Syn I. Moreover, the significant increase of axonal branching induced by BDNF in WT neurons was not detectable in Syn KO neurons. The expression analysis of BDNF receptors in Syn KO neurons revealed a significant decrease of the full length TrkB receptor and an increase in the levels of the truncated TrkB.t1 isoform and p75 NTR associated with a marked reduction of the BDNF-induced MAPK/Erk activation. By using the Trk inhibitor K252a, we demonstrated that these differences in BDNF effects were dependent on a TrkB/p75 NTR imbalance. The data indicate that Syn I plays a pivotal role in the BDNF signal transduction during axonal growth.

Environmental and genetic factors support the dissociation between α-synuclein aggregation and toxicity.

PubMed

Villar-Piqué, Anna; Lopes da Fonseca, Tomás; Sant'Anna, Ricardo; Szegö, Éva Mónika; Fonseca-Ornelas, Luis; Pinho, Raquel; Carija, Anita; Gerhardt, Ellen; Masaracchia, Caterina; Abad Gonzalez, Enrique; Rossetti, Giulia; Carloni, Paolo; Fernández, Claudio O; Foguel, Debora; Milosevic, Ira; Zweckstetter, Markus; Ventura, Salvador; Outeiro, Tiago Fleming

2016-10-18

Synucleinopathies are a group of progressive disorders characterized by the abnormal aggregation and accumulation of α-synuclein (aSyn), an abundant neuronal protein that can adopt different conformations and biological properties. Recently, aSyn pathology was shown to spread between neurons in a prion-like manner. Proteins like aSyn that exhibit self-propagating capacity appear to be able to adopt different stable conformational states, known as protein strains, which can be modulated both by environmental and by protein-intrinsic factors. Here, we analyzed these factors and found that the unique combination of the neurodegeneration-related metal copper and the pathological H50Q aSyn mutation induces a significant alteration in the aggregation properties of aSyn. We compared the aggregation of WT and H50Q aSyn with and without copper, and assessed the effects of the resultant protein species when applied to primary neuronal cultures. The presence of copper induces the formation of structurally different and less-damaging aSyn aggregates. Interestingly, these aggregates exhibit a stronger capacity to induce aSyn inclusion formation in recipient cells, which demonstrates that the structural features of aSyn species determine their effect in neuronal cells and supports a lack of correlation between toxicity and inclusion formation. In total, our study provides strong support in favor of the hypothesis that protein aggregation is not a primary cause of cytotoxicity.

Environmental and genetic factors support the dissociation between α-synuclein aggregation and toxicity

PubMed Central

Villar-Piqué, Anna; Lopes da Fonseca, Tomás; Sant’Anna, Ricardo; Szegö, Éva Mónika; Fonseca-Ornelas, Luis; Pinho, Raquel; Carija, Anita; Gerhardt, Ellen; Masaracchia, Caterina; Abad Gonzalez, Enrique; Rossetti, Giulia; Carloni, Paolo; Fernández, Claudio O.; Foguel, Debora; Milosevic, Ira; Zweckstetter, Markus; Ventura, Salvador; Outeiro, Tiago Fleming

2016-01-01

Synucleinopathies are a group of progressive disorders characterized by the abnormal aggregation and accumulation of α-synuclein (aSyn), an abundant neuronal protein that can adopt different conformations and biological properties. Recently, aSyn pathology was shown to spread between neurons in a prion-like manner. Proteins like aSyn that exhibit self-propagating capacity appear to be able to adopt different stable conformational states, known as protein strains, which can be modulated both by environmental and by protein-intrinsic factors. Here, we analyzed these factors and found that the unique combination of the neurodegeneration-related metal copper and the pathological H50Q aSyn mutation induces a significant alteration in the aggregation properties of aSyn. We compared the aggregation of WT and H50Q aSyn with and without copper, and assessed the effects of the resultant protein species when applied to primary neuronal cultures. The presence of copper induces the formation of structurally different and less-damaging aSyn aggregates. Interestingly, these aggregates exhibit a stronger capacity to induce aSyn inclusion formation in recipient cells, which demonstrates that the structural features of aSyn species determine their effect in neuronal cells and supports a lack of correlation between toxicity and inclusion formation. In total, our study provides strong support in favor of the hypothesis that protein aggregation is not a primary cause of cytotoxicity. PMID:27708160

Novel immunolocalization of alpha-synuclein in human muscle of inclusion-body myositis, regenerating and necrotic muscle fibers, and at neuromuscular junctions.

PubMed

Askanas, V; Engel, W K; Alvarez, R B; McFerrin, J; Broccolini, A

2000-07-01

Alpha-synuclein (alpha-syn) is an important component of neuronal and glial inclusions in brains of patients with several neurodegenerative disorders. Sporadic inclusion-body myositis (s-IBM) is the most common progressive muscle disease of older patients. Its muscle phenotype shows several similarities with Alzheimer disease brain. A distinct feature of s-IBM pathology is specific vacuolar degeneration of muscle fibers characterized by intracellular amyloid inclusions formed by both amyloid-beta (Abeta) and paired-helical filaments composed of phosphorylated tau. We immunostained alpha-syn in muscle biopsies of s-IBM, disease-control, and normal patients. Approximately 60% of Abeta-positive vacuolated muscle fibers (VMF) contained well-defined inclusions immunoreactive with antibodies against alpha-syn. In those fibers. alpha-syn co-localized with Abeta, both by light microscopy, and ultrastructurally. Paired-helical filaments did not contain alpha-syn immunoreactivity. In all muscle biopsies, alpha-syn was strongly immunoreactive at the postsynaptic region of the neuromuscular junctions. alpha-syn immunoreactivity also occurred diffusely in regenerating and necrotic muscle fibers. In cultured human muscle fibers, alpha-syn and its mRNA were expressed by immunocytochemistry, immunoblots, and Northern blots. Our study provides the first demonstration that alpha-syn participates in normal and pathologic processes of human muscle. Therefore. its function is not exclusive to the brain and neurodegenerative diseases.

1,3-syn-Diaxial Repulsion of Typical Protecting Groups Used in Carbohydrate Chemistry in 3-O-Substituted Derivatives of Isopropyl d-Idopyranosides.

PubMed

Komarova, Bozhena S; Gerbst, Alexey G; Finogenova, Anastasiia M; Dmitrenok, Andrey S; Tsvetkov, Yury E; Nifantiev, Nikolay E

2017-09-01

The strength of 1,3-syn-diaxial repulsion was evaluated for main types of protecting groups (alkyl, silyl, and acyl) usually used in carbohydrate chemistry. As molecular probes for this study, derivatives of isopropyl 2-O-benzyl-4,6-O-benzylidene-α-d-idopyranoside bearing allyl, acetyl, and tert-butyldiphenylsilyl (TBDPS) protecting groups at O-3 were prepared from p-methoxyphenyl d-galactopyranoside. The equilibrium between O S 2 and 4 C 1 conformations in these compounds was investigated using 3 J H,H and 3 J C,H coupling constants that were determined from 1D 1 H NMR and 2D J-resolved HMBC spectra in various solvents. The analysis of the corresponding coupling constants calculated using DFT/B3LYP/pcJ-1 approximation applied to conformations optimized at DFT/B3LYP/6-311++G** level supported the investigation. Proportions of conformers in the equilibrium revealed the highest repulsion between the 3-allyloxy group and the isopropoxy aglycon and its dependence on the solvent polarity. Differences in the conformational behavior of 3-O-allyl and 3-O-acetyl-α-d-idopyranoside derivatives complied with the notion that higher electron density on O-3 increased 1,3-syn-diaxial repulsion. 3-O-TBDPS derivative existed mainly in 4 C 1 conformation. The attenuation of the 1,3-syn-diaxial repulsive interaction indicates that TBDPS has stereoelectronic properties that may have significance in context of fixing unnatural pyranoside conformation with the help of silyl groups but have been disregarded until now.

Sequestration of synaptic proteins by alpha-synuclein aggregates leading to neurotoxicity is inhibited by small peptide

PubMed Central

Choi, Mal-Gi; Kim, Mi Jin; Kim, Do-Geun; Yu, Ri; Jang, You-Na

2018-01-01

α-Synuclein (α-syn) is a major component of Lewy bodies found in synucleinopathies including Parkinson’s disease (PD) and Dementia with Lewy Bodies (DLB). Under the pathological conditions, α-syn tends to generate a diverse form of aggregates showing toxicity to neuronal cells and able to transmit across cells. However, mechanisms by which α-syn aggregates affect cytotoxicity in neurons have not been fully elucidated. Here we report that α-syn aggregates preferentially sequester specific synaptic proteins such as vesicle-associated membrane protein 2 (VAMP2) and synaptosomal-associated protein 25 (SNAP25) through direct binding which is resistant to SDS. The sequestration effect of α-syn aggregates was shown in a cell-free system, cultured primary neurons, and PD mouse model. Furthermore, we identified a specific blocking peptide derived from VAMP2 which partially inhibited the sequestration by α-syn aggregates and contributed to reduced neurotoxicity. These results provide a mechanism of neurotoxicity mediated by α-syn aggregates and suggest that the blocking peptide interfering with the pathological role of α-syn aggregates could be useful for designing a potential therapeutic drug for the treatment of PD. PMID:29608598

MicroRNA-7 facilitates the degradation of alpha-synuclein and its aggregates by promoting autophagy.

PubMed

Choi, Doo Chul; Yoo, Myungsik; Kabaria, Savan; Junn, Eunsung

2018-05-05

Alpha-Synuclein (α-Syn) is an important protein in the pathogenesis of Parkinson disease (PD) as it accumulates as fibrillar inclusions in affected brain regions including dopaminergic neurons in the substantia nigra. Elevated levels of α-Syn seem to be crucial in mediating its toxicity. Thus, detailed information regarding the regulatory mechanism of α-Syn expression in several layers such as transcription, post-transcription and post-translation is needed in order to devise therapeutic interventions for PD. Previously, we reported that expression of α-Syn is repressed by microRNA-7 (miR-7) through its effect on the 3'-untranslated region (UTR) of α-Syn mRNA. Here, we show that miR-7 also accelerates the clearance of α-Syn and its aggregates by promoting autophagy in differentiated ReNcell VM cells. Further, miR-7 facilitates the degradation of pre-formed fibrils of α-Syn transported from outside the cells. This additional mechanism for reducing α-Syn levels show miR-7 to be an important molecular target for PD and other alpha-synucleinopathies. Copyright © 2018 Elsevier B.V. All rights reserved.

α-Synuclein interferes with the ESCRT-III complex contributing to the pathogenesis of Lewy body disease.

PubMed

Spencer, Brian; Kim, Changyoun; Gonzalez, Tania; Bisquertt, Alejandro; Patrick, Christina; Rockenstein, Edward; Adame, Anthony; Lee, Seung-Jae; Desplats, Paula; Masliah, Eliezer

2016-03-15

α-Synuclein (α-syn) has been implicated in neurological disorders with parkinsonism, including Parkinson's disease and Dementia with Lewy body. Recent studies have shown α-syn oligomers released from neurons can propagate from cell-to-cell in a prion-like fashion exacerbating neurodegeneration. In this study, we examined the role of the endosomal sorting complex required for transport (ESCRT) pathway on the propagation of α-syn. α-syn, which is transported via the ESCRT pathway through multivesicular bodies for degradation, can also target the degradation of the ESCRT protein-charged multivesicular body protein (CHMP2B), thus generating a roadblock of endocytosed α-syn. Disruption of the ESCRT transport system also resulted in increased exocytosis of α-syn thus potentially increasing cell-to-cell propagation of synuclein. Conversely, delivery of a lentiviral vector overexpressing CHMP2B rescued the neurodegeneration in α-syn transgenic mice. Better understanding of the mechanisms of intracellular trafficking of α-syn might be important for understanding the pathogenesis and developing new treatments for synucleinopathies. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Metabolic syndrome does not detect metabolic risk in African men living in the U.S.

PubMed

Ukegbu, Ugochi J; Castillo, Darleen C; Knight, Michael G; Ricks, Madia; Miller, Bernard V; Onumah, Barbara M; Sumner, Anne E

2011-10-01

Metabolic risk and metabolic syndrome (MetSyn) prevalence were compared in Africans who immigrated to the U.S. and African Americans. If MetSyn were an effective predictor of cardiometabolic risk, then the group with a worse metabolic risk profile would have a higher rate of MetSyn. Cross-sectional analyses were performed on 95 men (39 Africans, 56 African Americans, age 38 ± 6 years [mean ± SD]). Glucose tolerance was determined by oral glucose tolerance test, visceral adipose tissue (VAT) was determined by computerized tomography, and MetSyn was determined by the presence of three of five factors: central obesity, hypertriglyceridemia, low levels of HDL cholesterol, hypertension, and fasting hyperglycemia. MetSyn prevalence was similar in Africans and African Americans (10 vs. 13%, P = 0.74), but hypertension, glycemia (fasting and 2-h glucose), and VAT were higher in Africans. African immigrants have a worse metabolic profile than African Americans but a similar prevalence of MetSyn. Therefore, MetSyn may underpredict metabolic risk in Africans.

Novel animal model defines genetic contributions for neuron-to-neuron transfer of α-synuclein.

PubMed

Tyson, Trevor; Senchuk, Megan; Cooper, Jason F; George, Sonia; Van Raamsdonk, Jeremy M; Brundin, Patrik

2017-08-08

Cell-to-cell spreading of misfolded α-synuclein (α-syn) is suggested to contribute to the progression of neuropathology in Parkinson's disease (PD). Compelling evidence supports the hypothesis that misfolded α-syn transmits from neuron-to-neuron and seeds aggregation of the protein in the recipient cells. Furthermore, α-syn frequently appears to propagate in the brains of PD patients following a stereotypic pattern consistent with progressive spreading along anatomical pathways. We have generated a C. elegans model that mirrors this progression and allows us to monitor α-syn neuron-to-neuron transmission in a live animal over its lifespan. We found that modulation of autophagy or exo/endocytosis, affects α-syn transfer. Furthermore, we demonstrate that silencing C. elegans orthologs of PD-related genes also increases the accumulation of α-syn. This novel worm model is ideal for screening molecules and genes to identify those that modulate prion-like spreading of α-syn in order to target novel strategies for disease modification in PD and other synucleinopathies.

On the identity of some weevil species described by Johann Christian Fabricius (1745-1808) in the Museum of Zoology of Copenhagen (Coleoptera, Cucujoidea, Curculionoidea, Tenebrionoidea).

PubMed

Alonso-Zarazaga, Miguel A

2014-01-01

The types of thirty-two nominal weevil species described by Johann Christian Fabricius are reviewed and lecto- and paralectotypes are designated for twenty-two of them. A neotype is designated for Curculiosticticus Fabricius, 1777. Protapionvaripes (Germar, 1817) is declared a nomen protectum over Curculioflavipes Fabricius, 1775. Based on a study of syntypes, Rhinomacercurculioides Fabricius, 1781 is confirmed as a member of Mycterus (Mycteridae), Bruchusundatus Fabricius, 1787 is tentatively transferred to Erotylidae, Curculiofulvirostris Fabricius, 1787 and Anthribusroboris Fabricius, 1798 are confirmed as members of Salpingus (Salpingidae), and Brachyceruscristatus Fabricius, 1798 is transferred to Tenebrionidae. Based on lectotype designation, Curculiocaninus Fabricius, 1792 is confirmed as a synonym of Sitonalineatus (Linnaeus, 1758) and Curculioinnocuus Fabricius, 1802 as a synonym of Cneorhinusbarcelonicus (Herbst, 1797). Bruchusrufipes Fabricius, 1792 is not considered an available species name, but a later use of Bruchusrufipes Olivier, 1790. Cossonusincisus Pascoe, 1885 is reinstated as valid from synonymy under Cossonusilligeri Champion, 1909 and Cossonusvulneratus Illiger, 1805 from synonymy under Cossonuscanaliculatus (Fabricius, 1792) (a primary homonym of Curculiocanaliculatus Olivier, 1791). Cossonuscanaliculatus Fabricius, 1802 is a secondary homonym of the former and is replaced with Cossonusincisus. Salpingusfulvirostris (Fabricius, 1787) is reinstated as valid from synonymy under Salpingusplanirostris (Fabricius, 1787), a primary homonym of Curculioplanirostris Piller & Mitterpacher, 1783. The following new combinations are proposed: Brachysomuserinaceus (Fabricius, 1802) (from Curculio), Bronchusferus (Gyllenhal, 1840) (from Hipporhinus), Bronchusglandifer (Fabricius, 1792) (from Curculio), Bronchusnivosus (Sparrman, 1785) (from Curculio), Bronchussparrmani (Gyllenhal, 1833) (from Hipporhinus), Coelocephalapionatrirostre (Fabricius, 1802) (from Attelabus), Nerthopssticticus (Fabricius, 1777) (from Curculio), Piezotracheluscrotalariae (Fabricius, 1802) (from Attelabus), and Poropterusgranulatus (Fabricius, 1802) (from Curculio). The junior homonym Brachycerusuva Fabricius, 1792 (non Sparrman, 1785) is replaced by Brachycerusfabricii nom. n. The following new synonymies are established: Brachycerusobesus (Fabricius, 1775) = Curculioscalaris Fabricius, 1777, syn. n., Brachydereslusitanicus (Fabricius, 1781) = Curculiomoratus Fabricius, 1798, syn. n., Brachypera (Brachypera) crinita (Boheman, 1834) = Curculiostriatus Fabricius, 1787, syn. n., Brachysomuserinaceus (Fabricius, 1802) = Brachysomusvillosulus (Germar, 1824), syn. n., Bronchusabruptecostatus (Gyllenhal, 1833) = Curculiospectrum Fabricius, 1802, syn. n., Bronchusnivosus (Sparrman, 1785) = Curculiorecurvus Fabricius, 1802, syn. n., Camptorhinustibialis (Sparrman, 1785) = Rhynchaenusalienatus Fabricius, 1802, syn. n., Coelocephalapionatrirostre (Fabricius, 1802) = Coelocephalapionluteirostre (Gerstäcker, 1854), syn. n., Cyrtoderescristatus (DeGeer, 1778) (Tenebrionidae) = Brachyceruscristatus Fabricius, 1798, syn. n., Desmidophorushebes (Fabricius, 1781) = Curculiotuberculatus Fabricius, 1792, syn. n., Donussalviae (Schrank, 1789) = Curculiodenticornis Fabricius, 1798, syn. n., Exomiasholosericeus (Fabricius, 1802) = Exomiaschevrolati (Boheman, 1842), syn. n., Nerthopssticticus (Fabricius, 1777) = Nerthopsguttatus (Olivier, 1807), syn. n., Phyllobiusoblongus (Linnaeus, 1758) = Curculiomali Fabricius, 1782, syn. n., and Rhinocyllusconicus (Froelich, 1792) = Bruchuspunctatus Fabricius, 1798, syn. n. Bronchussynthesys sp. n. is described to represent the concept of Hipporhinusspectrum sensu Marshall, 1904, a misidentification.

Excitatory Synaptic Drive and Feedforward Inhibition in the Hippocampal CA3 Circuit Are Regulated by SynCAM 1.

PubMed

Park, Kellie A; Ribic, Adema; Laage Gaupp, Fabian M; Coman, Daniel; Huang, Yuegao; Dulla, Chris G; Hyder, Fahmeed; Biederer, Thomas

2016-07-13

Select adhesion proteins control the development of synapses and modulate their structural and functional properties. Despite these important roles, the extent to which different synapse-organizing mechanisms act across brain regions to establish connectivity and regulate network properties is incompletely understood. Further, their functional roles in different neuronal populations remain to be defined. Here, we applied diffusion tensor imaging (DTI), a modality of magnetic resonance imaging (MRI), to map connectivity changes in knock-out (KO) mice lacking the synaptogenic cell adhesion protein SynCAM 1. This identified reduced fractional anisotropy in the hippocampal CA3 area in absence of SynCAM 1. In agreement, mossy fiber refinement in CA3 was impaired in SynCAM 1 KO mice. Mossy fibers make excitatory inputs onto postsynaptic specializations of CA3 pyramidal neurons termed thorny excrescences and these structures were smaller in the absence of SynCAM 1. However, the most prevalent targets of mossy fibers are GABAergic interneurons and SynCAM 1 loss unexpectedly reduced the number of excitatory terminals onto parvalbumin (PV)-positive interneurons in CA3. SynCAM 1 KO mice additionally exhibited lower postsynaptic GluA1 expression in these PV-positive interneurons. These synaptic imbalances in SynCAM 1 KO mice resulted in CA3 disinhibition, in agreement with reduced feedforward inhibition in this network in the absence of SynCAM 1-dependent excitatory drive onto interneurons. In turn, mice lacking SynCAM 1 were impaired in memory tasks involving CA3. Our results support that SynCAM 1 modulates excitatory mossy fiber inputs onto both interneurons and principal neurons in the hippocampal CA3 area to balance network excitability. This study advances our understanding of synapse-organizing mechanisms on two levels. First, the data support that synaptogenic proteins guide connectivity and can function in distinct brain regions even if they are expressed broadly. Second, the results demonstrate that a synaptogenic process that controls excitatory inputs to both pyramidal neurons and interneurons can balance excitation and inhibition. Specifically, the study reveals that hippocampal CA3 connectivity is modulated by the synapse-organizing adhesion protein SynCAM 1 and identifies a novel, SynCAM 1-dependent mechanism that controls excitatory inputs onto parvalbumin-positive interneurons. This enables SynCAM 1 to regulate feedforward inhibition and set network excitability. Further, we show that diffusion tensor imaging is sensitive to these cellular refinements affecting neuronal connectivity. Copyright © 2016 the authors 0270-6474/16/367465-12$15.00/0.

Cotranslational structure acquisition of nascent polypeptides monitored by NMR spectroscopy.

PubMed

Eichmann, Cédric; Preissler, Steffen; Riek, Roland; Deuerling, Elke

2010-05-18

The folding of proteins in living cells may start during their synthesis when the polypeptides emerge gradually at the ribosomal exit tunnel. However, our current understanding of cotranslational folding processes at the atomic level is limited. We employed NMR spectroscopy to monitor the conformation of the SH3 domain from alpha-spectrin at sequential stages of elongation via in vivo ribosome-arrested (15)N,(13)C-labeled nascent polypeptides. These nascent chains exposed either the entire SH3 domain or C-terminally truncated segments thereof, thus providing snapshots of the translation process. We show that nascent SH3 polypeptides remain unstructured during elongation but fold into a compact, native-like beta-sheet assembly when the entire sequence information is available. Moreover, the ribosome neither imposes major conformational constraints nor significantly interacts with exposed unfolded nascent SH3 domain moieties. Our data provide evidence for a domainwise folding of the SH3 domain on ribosomes without significant population of folding intermediates. The domain follows a thermodynamically favorable pathway in which sequential folding units are stabilized, thus avoiding kinetic traps during the process of cotranslational folding.

Protein- protein interaction detection system using fluorescent protein microdomains

DOEpatents

Waldo, Geoffrey S.; Cabantous, Stephanie

2010-02-23

The invention provides a protein labeling and interaction detection system based on engineered fragments of fluorescent and chromophoric proteins that require fused interacting polypeptides to drive the association of the fragments, and further are soluble and stable, and do not change the solubility of polypeptides to which they are fused. In one embodiment, a test protein X is fused to a sixteen amino acid fragment of GFP (.beta.-strand 10, amino acids 198-214), engineered to not perturb fusion protein solubility. A second test protein Y is fused to a sixteen amino acid fragment of GFP (.beta.-strand 11, amino acids 215-230), engineered to not perturb fusion protein solubility. When X and Y interact, they bring the GFP strands into proximity, and are detected by complementation with a third GFP fragment consisting of GFP amino acids 1-198 (strands 1-9). When GFP strands 10 and 11 are held together by interaction of protein X and Y, they spontaneous association with GFP strands 1-9, resulting in structural complementation, folding, and concomitant GFP fluorescence.

Study of the influence of the bridge on the magnetic coupling in cobalt(II) complexes.

PubMed

Fabelo, Oscar; Cañadillas-Delgado, Laura; Pasán, Jorge; Delgado, Fernando S; Lloret, Francesc; Cano, Joan; Julve, Miguel; Ruiz-Pérez, Catalina

2009-12-07

Two new cobalt(II) complexes of formula [Co(2)(bta)(H(2)O)(6)](n) x 2nH(2)O (1) and [Co(phda)(H(2)O)](n) x nH(2)O (2) [H(4)bta = 1,2,4,5-benzenetetracarboxylic acid, H(2)phda = 1,4-phenylenediacetic acid] have been characterized by single crystal X-ray diffraction. Compound 1 is a one-dimensional compound where the bta(4-) ligand acts as 2-fold connector between the cobalt(II) ions through two carboxylate groups in para-conformation. Triply bridged dicobalt(II) units occur within each chain, a water molecule, a carboxylate group in the syn-syn conformation, and an oxo-carboxylate with the mu(2)O(1);kappa(2)O(1),O(2) coordination mode acting as bridges. Compound 2 is a three-dimensional compound, where the phda(2-) group acts as a bridge through its two carboxylate groups, one of them adopting the mu-O,O' coordination mode in the syn-syn conformation and the other exhibiting the single mu(2)-O'' bridging mode. As in 1, chains of cobalt(II) ions occur in 2 with a water molecule, a syn-syn carboxylate group, and an oxo-carboxylate constitute the triply intrachain bridging skeleton. Each chain is linked to other four ones through the phda(2-) ligand, giving rise to the three-dimensional structure. The values of the intrachain cobalt-cobalt separation are 3.1691(4) (1) and 3.11499(2) A (2) whereas those across the phenyl ring of the extended bta(4-) (1) and phda(2-) (2) groups are 10.1120(6) and 11.4805(69 A, respectively. The magnetic properties of 1 and 2 have been investigated in the temperature range 1.9-300 K, and their analysis has revealed the occurrence of moderate intrachain ferromagnetic couplings [J = +5.4 (1) and +2.16 cm(-1) (2), J being the isotropic magnetic coupling parameter], the magnetic coupling through the extended bta(4-) and phda(2-) with cobalt-cobalt separations larger than 10 A being negligible. The nature and magnitude of the magnetic interactions between the high-spin cobalt(II) ions in 1 and 2 are compared to those of related systems and discussed as a function of the complementarity-countercomplementarity effects of the triple bridges.

Phosphorylation of Synaptic GTPase-activating Protein (synGAP) by Ca2+/Calmodulin-dependent Protein Kinase II (CaMKII) and Cyclin-dependent Kinase 5 (CDK5) Alters the Ratio of Its GAP Activity toward Ras and Rap GTPases*

PubMed Central

Walkup, Ward G.; Washburn, Lorraine; Sweredoski, Michael J.; Carlisle, Holly J.; Graham, Robert L.; Hess, Sonja; Kennedy, Mary B.

2015-01-01

synGAP is a neuron-specific Ras and Rap GTPase-activating protein (GAP) found in high concentrations in the postsynaptic density (PSD) fraction from the mammalian forebrain. We have previously shown that, in situ in the PSD fraction or in recombinant form in Sf9 cell membranes, synGAP is phosphorylated by Ca2+/calmodulin-dependent protein kinase II (CaMKII), another prominent component of the PSD. Here, we show that recombinant synGAP (r-synGAP), lacking 102 residues at the N terminus, can be purified in soluble form and is phosphorylated by cyclin-dependent kinase 5 (CDK5) as well as by CaMKII. Phosphorylation of r-synGAP by CaMKII increases its HRas GAP activity by 25% and its Rap1 GAP activity by 76%. Conversely, phosphorylation by CDK5 increases r-synGAP's HRas GAP activity by 98% and its Rap1 GAP activity by 20%. Thus, phosphorylation by both kinases increases synGAP activity; CaMKII shifts the relative GAP activity toward inactivation of Rap1, and CDK5 shifts the relative activity toward inactivation of HRas. GAP activity toward Rap2 is not altered by phosphorylation by either kinase. CDK5 phosphorylates synGAP primarily at two sites, Ser-773 and Ser-802. Phosphorylation at Ser-773 inhibits r-synGAP activity, and phosphorylation at Ser-802 increases it. However, the net effect of concurrent phosphorylation of both sites, Ser-773 and Ser-802, is an increase in GAP activity. synGAP is phosphorylated at Ser-773 and Ser-802 in the PSD fraction, and its phosphorylation by CDK5 and CaMKII is differentially regulated by activation of NMDA-type glutamate receptors in cultured neurons. PMID:25533468

Does the tautomeric status of the adenine bases change upon the dissociation of the A*·A(syn) Topal-Fresco DNA mismatch? A combined QM and QTAIM atomistic insight.

PubMed

Brovarets', Ol'ha O; Zhurakivsky, Roman O; Hovorun, Dmytro M

2014-02-28

We have scrupulously explored the tautomerisation mechanism via the double proton transfer of the A*·A(syn) Topal-Fresco base mispair (C(s) symmetry), formed by the imino and amino tautomers of the adenine DNA base in the anti- and syn-conformations, respectively, bridging quantum-mechanical calculations with Bader's quantum theory of atoms in molecules. It was found that the A*·A(syn) ↔ A·A*(syn) tautomerisation is the asynchronous concerted process. It was established that the A*·A(syn) DNA mismatch is stabilized by the N6H···N6 (6.35) and N1H···N7 (6.17) hydrogen (H) bonds, whereas the A·A*(syn) base mispair (Cs) by the N6H···N6 (8.82) and N7H···N1 (9.78) H-bonds and the C8H···HC2 HH-bond (0.30 kcal mol(-1)). Using the sweeps of the energies of the intermolecular H-bonds, it was observed that the N6H···N6 and N1H···N7/N7H···N1 H-bonds are anti-cooperative and mutually weaken each other in the A*·A(syn) and A·A*(syn) mispairs. It was revealed that the A·A*(syn) DNA mismatch is a dynamically unstable structure with a short lifetime of 1.12 × 10(-13) s and any of its 6 low-frequency intermolecular vibrations can develop during this period of time. This observation makes it impossible to change the tautomeric status of the A bases upon the dissociation of the A*·A(syn) base mispair into the monomers during DNA replication.

Inoculation of α-synuclein preformed fibrils into the mouse gastrointestinal tract induces Lewy body-like aggregates in the brainstem via the vagus nerve.

PubMed

Uemura, Norihito; Yagi, Hisashi; Uemura, Maiko T; Hatanaka, Yusuke; Yamakado, Hodaka; Takahashi, Ryosuke

2018-05-11

Intraneuronal α-synuclein (α-Syn) aggregates known as Lewy bodies (LBs) and the loss of dopaminergic neurons in the substantia nigra pars compacta (SNpc) are the pathological hallmarks of Parkinson's disease (PD). Braak's hypothesis based on autopsy studies suggests that Lewy pathology initially occurs in the enteric nervous system (ENS) and then travels retrogradely to the dorsal motor nucleus of the vagus nerve (dmX), proceeding from there in a caudo-rostral direction. Recent evidence that α-Syn aggregates propagate between interconnected neurons supports this hypothesis. However, there is no direct evidence demonstrating this transmission from the ENS to the dmX and then to the SNpc. We inoculated α-Syn preformed fibrils (PFFs) or phosphate-buffered saline (PBS) into the mouse gastric wall and analyzed the progression of the pathology. The mice inoculated with α-Syn PFFs, but not with PBS, developed phosphorylated α-Syn (p-α-Syn)-positive LB-like aggregates in the dmX at 45 days postinoculation. This aggregate formation was completely abolished when vagotomy was performed prior to inoculation of α-Syn PFFs, suggesting that the aggregates in the dmX were retrogradely induced via the vagus nerve. Unexpectedly, the number of neurons containing p-α-Syn-positive aggregates in the dmX decreased over time, and no further caudo-rostral propagation beyond the dmX was observed up to 12 months postinoculation. P-α-Syn-positive aggregates were also present in the myenteric plexus at 12 months postinoculation. However, unlike in patients with PD, there was no cell-type specificity in neurons containing those aggregates in this model. These results indicate that α-Syn PFF inoculation into the mouse gastrointestinal tract can induce α-Syn pathology resembling that of very early PD, but other factors are apparently required if further progression of PD pathology is to be replicated in this animal model.

Method for altering antibody light chain interactions

DOEpatents

Stevens, Fred J.; Stevens, Priscilla Wilkins; Raffen, Rosemarie; Schiffer, Marianne

2002-01-01

A method for recombinant antibody subunit dimerization including modifying at least one codon of a nucleic acid sequence to replace an amino acid occurring naturally in the antibody with a charged amino acid at a position in the interface segment of the light polypeptide variable region, the charged amino acid having a first polarity; and modifying at least one codon of the nucleic acid sequence to replace an amino acid occurring naturally in the antibody with a charged amino acid at a position in an interface segment of the heavy polypeptide variable region corresponding to a position in the light polypeptide variable region, the charged amino acid having a second polarity opposite the first polarity. Nucleic acid sequences which code for novel light chain proteins, the latter of which are used in conjunction with the inventive method, are also provided.

Isolation and characterization of spinach photosystem II membrane-associated catalase and polyphenol oxidase.

PubMed

Sheptovitsky, Y G; Brudvig, G W

1996-12-17

Photosystem II (PSII) membranes exhibit catalase and polyphenol oxidase (PPO) activities. Mild heat treatment of PSII membranes for 90 min at 30 degrees C releases most of these enzyme activities into the supernatant, accompanied by a 7-fold activation of PPO. In contrast, mild heat treatment of thylakoid membranes does not release significant amounts of either activity, indicating that both enzymes are bound to the luminal surface of the thylakoid membrane. The heat-released PSII membrane-associated catalase and PPO have been purified and characterized. Catalase activity was correlated with a 63 kDa polypeptide which was purified by batch adsorption to anion-exchange beads followed by gel filtration. The PSII membrane-associated catalase is unstable in solution, probably due to irreversible aggregation. The enzyme was characterized in terms of molecular and subunit size, amino-acid composition, UV-visible absorption, heme content, pH optimum, inhibitor sensitivity, and K(m) value for H2O2. Its properties indicate that the PSII membrane-associated catalase is a luminal thylakoid membrane-bound heme enzyme that has not been identified previously. The residual catalase activity of PSII membranes after mild heat treatment is irreversibly inhibited with 3-amino-1,2,4-triazole, a specific inhibitor of heme catalases, without inhibition of O2-evolution activity. This result indicates that little, if any, of the catalase activity from PSII membranes in the dark is catalyzed by the O2-evolving center of PSII. PPO activity was correlated with a 48 kDa polypeptide. However, the 48 kDa polypeptide and another heat-released polypeptide of 72 kDa have the same N-terminal sequence, which is also identical to that of a known 64 kDa protein [Hind, G., Marshak, D. R., & Coughlan, S. J. (1995) Biochemistry 34, 8157-8164]. During heat treatment of PSII membranes and further manipulations it was found that the 72 kDa polypeptide was largely converted into the 48 kDa polypeptide. Thus, the 72 kDa polypeptide appears to be a latent precursor of the active 48 kDa PPO. The PSII membrane-associated PPO was purified by anion-exchange chromatography and was characterized in terms of substrate specificity, pH optimum, inhibitor sensitivity and native molecular weight. The heat-released PPO appears to be identical to the enzyme previously isolated from spinach thylakoid membranes [Golbeck, J. H., & Cammarata, K. V. (1981) Plant Physiol. 67, 977-984].

Preclinical development of a vaccine against oligomeric alpha-synuclein based on virus-like particles.

PubMed

Doucet, Marika; El-Turabi, Aadil; Zabel, Franziska; Hunn, Benjamin H M; Bengoa-Vergniory, Nora; Cioroch, Milena; Ramm, Mauricio; Smith, Amy M; Gomes, Ariane Cruz; Cabral de Miranda, Gustavo; Wade-Martins, Richard; Bachmann, Martin F

2017-01-01

Parkinson's disease (PD) is a progressive and currently incurable neurological disorder characterised by the loss of midbrain dopaminergic neurons and the accumulation of aggregated alpha-synuclein (a-syn). Oligomeric a-syn is proposed to play a central role in spreading protein aggregation in the brain with associated cellular toxicity contributing to a progressive neurological decline. For this reason, a-syn oligomers have attracted interest as therapeutic targets for neurodegenerative conditions such as PD and other alpha-synucleinopathies. In addition to strategies using small molecules, neutralisation of the toxic oligomers by antibodies represents an attractive and highly specific strategy for reducing disease progression. Emerging active immunisation approaches using vaccines are already being trialled to induce such antibodies. Here we propose a novel vaccine based on the RNA bacteriophage (Qbeta) virus-like particle conjugated with short peptides of human a-syn. High titres of antibodies were successfully and safely generated in wild-type and human a-syn over-expressing (SNCA-OVX) transgenic mice following vaccination. Antibodies from vaccine candidates targeting the C-terminal regions of a-syn were able to recognise Lewy bodies, the hallmark aggregates in human PD brains. Furthermore, antibodies specifically targeted oligomeric and aggregated a-syn as they exhibited 100 times greater affinity for oligomeric species over monomer a-syn proteins in solution. In the SNCA-OVX transgenic mice used, vaccination was, however, unable to confer significant changes to oligomeric a-syn bioburden. Similarly, there was no discernible effect of vaccine treatment on behavioural phenotype as compared to control groups. Thus, antibodies specific for oligomeric a-syn induced by vaccination were unable to treat symptoms of PD in this particular mouse model.

Preclinical development of a vaccine against oligomeric alpha-synuclein based on virus-like particles

PubMed Central

Zabel, Franziska; Hunn, Benjamin H.M.; Bengoa-Vergniory, Nora; Cioroch, Milena; Ramm, Mauricio; Smith, Amy M.; Gomes, Ariane Cruz; Cabral de Miranda, Gustavo; Wade-Martins, Richard; Bachmann, Martin F.

2017-01-01

Parkinson's disease (PD) is a progressive and currently incurable neurological disorder characterised by the loss of midbrain dopaminergic neurons and the accumulation of aggregated alpha-synuclein (a-syn). Oligomeric a-syn is proposed to play a central role in spreading protein aggregation in the brain with associated cellular toxicity contributing to a progressive neurological decline. For this reason, a-syn oligomers have attracted interest as therapeutic targets for neurodegenerative conditions such as PD and other alpha-synucleinopathies. In addition to strategies using small molecules, neutralisation of the toxic oligomers by antibodies represents an attractive and highly specific strategy for reducing disease progression. Emerging active immunisation approaches using vaccines are already being trialled to induce such antibodies. Here we propose a novel vaccine based on the RNA bacteriophage (Qbeta) virus-like particle conjugated with short peptides of human a-syn. High titres of antibodies were successfully and safely generated in wild-type and human a-syn over-expressing (SNCA-OVX) transgenic mice following vaccination. Antibodies from vaccine candidates targeting the C-terminal regions of a-syn were able to recognise Lewy bodies, the hallmark aggregates in human PD brains. Furthermore, antibodies specifically targeted oligomeric and aggregated a-syn as they exhibited 100 times greater affinity for oligomeric species over monomer a-syn proteins in solution. In the SNCA-OVX transgenic mice used, vaccination was, however, unable to confer significant changes to oligomeric a-syn bioburden. Similarly, there was no discernible effect of vaccine treatment on behavioural phenotype as compared to control groups. Thus, antibodies specific for oligomeric a-syn induced by vaccination were unable to treat symptoms of PD in this particular mouse model. PMID:28797124

Annotated type catalogue of the Chrysididae (Insecta, Hymenoptera) deposited in the collection of Radoszkowski in the Polish Academy of Sciences, Kraków

PubMed Central

Rosa, Paolo; Wiśniowski, Bogdan; Xu, Zai-fu

2015-01-01

Abstract A critical and annotated catalogue of 183 types of Hymenoptera Chrysididae belonging to 124 taxa housed in the Radoszkowski collection is given. Radoszkowski type material from other institutes has also been checked. Six lectotypes are designated in Kraków (ISEA-PAN): Chrysis acceptabilis Radoszkowski, 1891; Chrysis persica Radoczkowsky, 1881; Chrysis daphnis Mocsáry, 1889; Chrysis lagodechii Radoszkowski, 1889; Chrysis remota Mocsáry, 1889 and Chrysis vagans Radoszkowski, 1877. The lectotype of Brugmoia pellucida Radoszkowski, 1877 is designated in Moscow (MMU). Four new combinations are proposed: Philoctetes araraticus (Radoszkowski, 1890), comb. n.; Pseudomalus hypocrita (du Buysson, 1893), comb. n.; Chrysis eldari (Radoszkowski, 1893), comb. n.; and Chrysura mlokosewitzi (Radoszkowski, 1889), comb. n.. Ten new synonyms are given: Chrysis auropunctata Mocsáry, 1889, syn. n. of Chrysis angolensis Radoszkovsky, 1881; Chrysis chrysochlora Mocsáry, 1889, syn. n. and Chrysis viridans Radoszkowski, 1891, syn. n. of Chrysis keriensis Radoszkowski, 1887; Chrysis angustifrons var. ignicollis Trautmann, 1926, syn. n. of Chrysis eldari (Radoszkowski, 1893); Chrysis maracandensis var. simulatrix Radoszkowski, 1891, syn. n. of Chrysis maracandensis Radoszkowski, 1877; Chrysis pulchra Radoszkovsky, 1880, syn. n. of Spinolia dallatorreana (Mocsáry, 1896); Chrysis rubricollis du Buysson, 1900, syn. n. of Chrysis eldari (Radoszkowski, 1893); Chrysis subcoerulea Radoszkowski, 1891, syn. n. of Chrysis chlorochrysa Mocsáry, 1889; Chrysis therates Mocsáry, 1889, syn. n. of Chrysis principalis Smith, 1874; and Notozus komarowi Radoszkowski, 1893, syn. n. of Elampus obesus (Mocsáry, 1890). One species is revaluated: Chrysis chalcochrysa Mocsáry, 1887. Chrysis kizilkumiana Rosa is the new name for Chrysis uljanini Radoszkowski & Mocsáry, 1889 nec Radoszkowski, 1877. Pictures of seventy-seven type specimens are given. PMID:25829848

Serine 129 phosphorylation of membrane-associated α-synuclein modulates dopamine transporter function in a G protein–coupled receptor kinase–dependent manner

PubMed Central

Hara, Susumu; Arawaka, Shigeki; Sato, Hiroyasu; Machiya, Youhei; Cui, Can; Sasaki, Asuka; Koyama, Shingo; Kato, Takeo

2013-01-01

Most α-synuclein (α-syn) deposited in Lewy bodies, the pathological hallmark of Parkinson disease (PD), is phosphorylated at Ser-129. However, the physiological and pathological roles of this modification are unclear. Here we investigate the effects of Ser-129 phosphorylation on dopamine (DA) uptake in dopaminergic SH-SY5Y cells expressing α-syn. Subcellular fractionation of small interfering RNA (siRNA)–treated cells shows that G protein–coupled receptor kinase 3 (GRK3), GRK5, GRK6, and casein kinase 2 (CK2) contribute to Ser-129 phosphorylation of membrane-associated α-syn, whereas cytosolic α-syn is phosphorylated exclusively by CK2. Expression of wild-type α-syn increases DA uptake, and this effect is diminished by introducing the S129A mutation into α-syn. However, wild-type and S129A α-syn equally increase the cell surface expression of dopamine transporter (DAT) in SH-SY5Y cells and nonneuronal HEK293 cells. In addition, siRNA-mediated knockdown of GRK5 or GRK6 significantly attenuates DA uptake without altering DAT cell surface expression, whereas knockdown of CK2 has no effect on uptake. Taken together, our results demonstrate that membrane-associated α-syn enhances DA uptake capacity of DAT by GRKs-mediated Ser-129 phosphorylation, suggesting that α-syn modulates intracellular DA levels with no functional redundancy in Ser-129 phosphorylation between GRKs and CK2. PMID:23576548

The Potential Role of Toll-Like Receptor 4 in Mediating Dopaminergic Cell Loss and Alpha-Synuclein Expression in the Acute MPTP Mouse Model of Parkinson's Disease.

PubMed

Mariucci, Giuseppina; Pagiotti, Rita; Galli, Francesco; Romani, Luigina; Conte, Carmela

2018-04-01

Toll-like receptors (TLRs) may have a role in Parkinson's disease (PD). In this study, we aimed at investigating the dopaminergic cell loss and alpha-synuclein (α-SYN) expression in TLR4-deficient mice (TLR4 -/- ) acutely exposed to 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), a pharmacological PD model. TLR4 ablation restrained the number of dopaminergic neurons in the substantia nigra (SN), as assessed by tyrosine hydroxylase (TH) protein expression. Intriguingly, TLR4 -/- mice showed massive α-SYN protein accumulation in the midbrain along with high α-SYN mRNA levels in cerebral cortex, striatum, hippocampus, and cerebellum. Contrary to expectations, the high levels of α-SYN do not correlate with greater dopaminergic neuronal loss. The levels of nigral α-SYN protein in TLR4 -/- mice further, but not significantly, increased during MPTP treatment. Contrariwise, MPTP treatment significantly induced the mRNA expression of α-SYN in examined brain regions of WT and TLR4 -/- mice. Protein levels of GATA2, a transcription factor proposed to control α-SYN gene expression, did not change in TLR4 -/- mice at baseline and after MPTP treatment. These findings suggest a role for TLR4 in mediating dopaminergic cell loss and in the constitutive expression of brain α-SYN. However, further exploration is needed in order to establish the actual role of α-SYN in the relative absence of TLR4.

Aromatic interactions are not required for amyloid fibril formation by islet amyloid polypeptide but do influence the rate of fibril formation and fibril morphology.

PubMed

Marek, Peter; Abedini, Andisheh; Song, BenBen; Kanungo, Mandakini; Johnson, Megan E; Gupta, Ruchi; Zaman, Warda; Wong, Stanislaus S; Raleigh, Daniel P

2007-03-20

Amyloid formation has been implicated in a wide range of human diseases, and a diverse set of proteins is involved. There is considerable interest in elucidating the interactions which lead to amyloid formation and which contribute to amyloid fibril stability. Recent attention has been focused upon the potential role of aromatic-aromatic and aromatic-hydrophobic interactions in amyloid formation by short to midsized polypeptides. Here we examine whether aromatic residues are necessary for amyloid formation by islet amyloid polypeptide (IAPP). IAPP is responsible for the formation of islet amyloid in type II diabetes which is thought to play a role in the pathology of the disease. IAPP is 37 residues in length and contains three aromatic residues, Phe-15, Phe-23, and Tyr-37. Structural models of IAPP amyloid fibrils postulate that Tyr-37 is near one of the phenylalanine residues, and it is known that Tyr-37 interacts with one of the phenylalanines during fibrillization; however, it is not known if aromatic-aromatic or aromatic-hydrophobic interactions are absolutely required for amyloid formation. An F15L/F23L/Y37L triple mutant (IAPP-3XL) was prepared, and its ability to form amyloid was tested. CD, thioflavin binding assays, AFM, and TEM measurements all show that the triple leucine mutant readily forms amyloid fibrils. The substitutions do, however, decrease the rate of fibril formation and alter the tendency of fibrils to aggregate. Thus, while aromatic residues are not an absolute requirement for amyloid formation by IAPP, they do play a role in the fibril assembly process.

Mutational analysis of the ability of resveratrol to inhibit amyloid formation by islet amyloid polypeptide: critical evaluation of the importance of aromatic-inhibitor and histidine-inhibitor interactions.

PubMed

Tu, Ling-Hsien; Young, Lydia M; Wong, Amy G; Ashcroft, Alison E; Radford, Sheena E; Raleigh, Daniel P

2015-01-27

The process of amyloid formation by the normally soluble hormone islet amyloid polypeptide (IAPP) contributes to β-cell death in type 2 diabetes and in islet transplants. There are no clinically approved inhibitors of islet amyloidosis, and the mode of action of existing inhibitors is not well-understood. Resveratrol, a natural polyphenol, has been reported to inhibit amyloid formation by IAPP and by the Alzheimer's disease Aβ peptide. The mechanism of action of this compound is not known, nor is its mode of interaction with IAPP. In this study, we use a series of IAPP variants to examine possible interactions between resveratrol and IAPP. Fluorescence assays, transmission electron microscopy, and mass spectrometry demonstrate that resveratrol is much less effective as an inhibitor of IAPP amyloid formation than the polyphenol (-)-epigallocatechin 3-gallate (EGCG) and, unlike EGCG, does not significantly disaggregate preformed IAPP amyloid fibrils. Resveratrol is also shown to interfere with thioflavin-T assays. His-18 mutants, a truncation mutant, mutants of each of the aromatic residues, and mutants of Arg-11 of IAPP were examined. Mutation of His to Gln or Leu weakens the ability of resveratrol to inhibit amyloid formation by IAPP, as do mutations of Arg-11, Phe-15, or Tyr-37 to Leu, and truncation to form the variant Ac 8-37-IAPP, which removes the first seven residues to eliminate Lys-1 and the N-terminal amino group. In contrast, replacement of Phe-23 with Leu has a smaller effect. The data highlight Phe-15, His-18, and Tyr-37 as being important for IAPP-resveratrol interactions and are consistent with a potential role of the N-terminus and Arg-11 in polypeptide-resveratrol interactions.

Munc18-1 is a molecular chaperone for α-synuclein, controlling its self-replicating aggregation.

PubMed

Chai, Ye Jin; Sierecki, Emma; Tomatis, Vanesa M; Gormal, Rachel S; Giles, Nichole; Morrow, Isabel C; Xia, Di; Götz, Jürgen; Parton, Robert G; Collins, Brett M; Gambin, Yann; Meunier, Frédéric A

2016-09-12

Munc18-1 is a key component of the exocytic machinery that controls neurotransmitter release. Munc18-1 heterozygous mutations cause developmental defects and epileptic phenotypes, including infantile epileptic encephalopathy (EIEE), suggestive of a gain of pathological function. Here, we used single-molecule analysis, gene-edited cells, and neurons to demonstrate that Munc18-1 EIEE-causing mutants form large polymers that coaggregate wild-type Munc18-1 in vitro and in cells. Surprisingly, Munc18-1 EIEE mutants also form Lewy body-like structures that contain α-synuclein (α-Syn). We reveal that Munc18-1 binds α-Syn, and its EIEE mutants coaggregate α-Syn. Likewise, removal of endogenous Munc18-1 increases the aggregative propensity of α-Syn(WT) and that of the Parkinson's disease-causing α-Syn(A30P) mutant, an effect rescued by Munc18-1(WT) expression, indicative of chaperone activity. Coexpression of the α-Syn(A30P) mutant with Munc18-1 reduced the number of α-Syn(A30P) aggregates. Munc18-1 mutations and haploinsufficiency may therefore trigger a pathogenic gain of function through both the corruption of native Munc18-1 and a perturbed chaperone activity for α-Syn leading to aggregation-induced neurodegeneration. © 2016 Chai et al.

Lysosomal enzyme cathepsin B enhances the aggregate forming activity of exogenous α-synuclein fibrils.

PubMed

Tsujimura, Atsushi; Taguchi, Katsutoshi; Watanabe, Yoshihisa; Tatebe, Harutsugu; Tokuda, Takahiko; Mizuno, Toshiki; Tanaka, Masaki

2015-01-01

The formation of intracellular aggregates containing α-synuclein (α-Syn) is one of the key steps in the progression of Parkinson's disease and dementia with Lewy bodies. Recently, it was reported that pathological α-Syn fibrils can undergo cell-to-cell transmission and form Lewy body-like aggregates. However, little is known about how they form α-Syn aggregates from fibril seeds. Here, we developed an assay to study the process of aggregate formation using fluorescent protein-tagged α-Syn-expressing cells and examined the aggregate forming activity of exogenous α-Syn fibrils. α-Syn fibril-induced formation of intracellular aggregates was suppressed by a cathepsin B specific inhibitor, but not by a cathepsin D inhibitor. α-Syn fibrils pretreated with cathepsin B in vitro enhanced seeding activity in cells. Knockdown of cathepsin B also reduced fibril-induced aggregate formation. Moreover, using LAMP-1 immunocytochemistry and live-cell imaging, we observed that these aggregates initially occurred in the lysosome. They then rapidly grew larger and moved outside the boundary of the lysosome within one day. These results suggest that the lysosomal protease cathepsin B is involved in triggering intracellular aggregate formation by α-Syn fibrils. Copyright © 2015. Published by Elsevier Inc.

Novel syn intramolecular pathway in base-catalyzed 1,2-elimination reactions of beta-acetoxy esters.

PubMed

Mohrig, Jerry R; Carlson, Hans K; Coughlin, Jane M; Hofmeister, Gretchen E; McMartin, Lea A; Rowley, Elizabeth G; Trimmer, Elizabeth E; Wild, Andrew J; Schultz, Steve C

2007-02-02

As part of a comprehensive investigation of electronic effects on the stereochemistry of base-catalyzed 1,2-elimination reactions, we observed a new syn intramolecular pathway in the elimination of acetic acid from beta-acetoxy esters and thioesters. 1H and 2H NMR investigation of reactions using stereospecifically labeled tert-butyl (2R*,3R*)-3-acetoxy-2,3-2H2-butanoate (1) and its (2R*,3S*) diastereomer (2) shows that 23 +/- 2% syn elimination occurs. The elimination reactions were catalyzed with KOH or (CH3)4NOH in ethanol/water under rigorously non-ion-pairing conditions. By contrast, the more sterically hindered beta-trimethylacetoxy ester produces only 6 +/- 1% syn elimination. These data strongly support an intramolecular (Ei) syn path for elimination of acetic acid, most likely through the oxyanion produced by nucleophilic attack at the carbonyl carbon of the beta-acetoxy group. The analogous thioesters, S-tert-butyl (2R*,3R*)-3-acetoxy-2,3-2H2-butanethioate (3) and its (2R*,3S*) diastereomer (4), showed 18 +/- 2% syn elimination, whereas the beta-trimethylacetoxy substrate gave 5 +/- 1% syn elimination. The more acidic thioester substrates do not produce an increased amount of syn stereoselectivity even though their elimination reactions are at the E1cb interface.

Munc18-1 is a molecular chaperone for α-synuclein, controlling its self-replicating aggregation

PubMed Central

Giles, Nichole; Morrow, Isabel C.; Collins, Brett M.

2016-01-01

Munc18-1 is a key component of the exocytic machinery that controls neurotransmitter release. Munc18-1 heterozygous mutations cause developmental defects and epileptic phenotypes, including infantile epileptic encephalopathy (EIEE), suggestive of a gain of pathological function. Here, we used single-molecule analysis, gene-edited cells, and neurons to demonstrate that Munc18-1 EIEE-causing mutants form large polymers that coaggregate wild-type Munc18-1 in vitro and in cells. Surprisingly, Munc18-1 EIEE mutants also form Lewy body–like structures that contain α-synuclein (α-Syn). We reveal that Munc18-1 binds α-Syn, and its EIEE mutants coaggregate α-Syn. Likewise, removal of endogenous Munc18-1 increases the aggregative propensity of α-SynWT and that of the Parkinson’s disease–causing α-SynA30P mutant, an effect rescued by Munc18-1WT expression, indicative of chaperone activity. Coexpression of the α-SynA30P mutant with Munc18-1 reduced the number of α-SynA30P aggregates. Munc18-1 mutations and haploinsufficiency may therefore trigger a pathogenic gain of function through both the corruption of native Munc18-1 and a perturbed chaperone activity for α-Syn leading to aggregation-induced neurodegeneration. PMID:27597756

Metabolic Syndrome Does Not Detect Metabolic Risk in African Men Living in the U.S.

PubMed Central

Ukegbu, Ugochi J.; Castillo, Darleen C.; Knight, Michael G.; Ricks, Madia; Miller, Bernard V.; Onumah, Barbara M.; Sumner, Anne E.

2011-01-01

OBJECTIVE Metabolic risk and metabolic syndrome (MetSyn) prevalence were compared in Africans who immigrated to the U.S. and African Americans. If MetSyn were an effective predictor of cardiometabolic risk, then the group with a worse metabolic risk profile would have a higher rate of MetSyn. RESEARCH DESIGN AND METHODS Cross-sectional analyses were performed on 95 men (39 Africans, 56 African Americans, age 38 ± 6 years [mean ± SD]). Glucose tolerance was determined by oral glucose tolerance test, visceral adipose tissue (VAT) was determined by computerized tomography, and MetSyn was determined by the presence of three of five factors: central obesity, hypertriglyceridemia, low levels of HDL cholesterol, hypertension, and fasting hyperglycemia. RESULTS MetSyn prevalence was similar in Africans and African Americans (10 vs. 13%, P = 0.74), but hypertension, glycemia (fasting and 2-h glucose), and VAT were higher in Africans. CONCLUSIONS African immigrants have a worse metabolic profile than African Americans but a similar prevalence of MetSyn. Therefore, MetSyn may underpredict metabolic risk in Africans. PMID:21873563

Reversible thermal denaturation of a 60-kDa genetically engineered beta-sheet polypeptide.

PubMed

Lednev, Igor K; Ermolenkov, Vladimir V; Higashiya, Seiichiro; Popova, Ludmila A; Topilina, Natalya I; Welch, John T

2006-11-15

A de novo 687-amino-acid residue polypeptide with a regular 32-amino-acid repeat sequence, (GA)(3)GY(GA)(3)GE(GA)(3)GH(GA)(3)GK, forms large beta-sheet assemblages that exhibit remarkable folding properties and, as well, form fibrillar structures. This construct is an excellent tool to explore the details of beta-sheet formation yielding intimate folding information that is otherwise difficult to obtain and may inform folding studies of naturally occurring materials. The polypeptide assumes a fully folded antiparallel beta-sheet/turn structure at room temperature, and yet is completely and reversibly denatured at 125 degrees C, adopting a predominant polyproline II conformation. Deep ultraviolet Raman spectroscopy indicated that melting/refolding occurred without any spectroscopically distinct intermediates, yet the relaxation kinetics depend on the initial polypeptide state, as would be indicative of a non-two-state process. Thermal denaturation and refolding on cooling appeared to be monoexponential with characteristic times of approximately 1 and approximately 60 min, respectively, indicating no detectable formation of hairpin-type nuclei in the millisecond timescale that could be attributed to nonlocal "nonnative" interactions. The polypeptide folding dynamics agree with a general property of beta-sheet proteins, i.e., initial collapse precedes secondary structure formation. The observed folding is much faster than expected for a protein of this size and could be attributed to a less frustrated free-energy landscape funnel for folding. The polypeptide sequence suggests an important balance between the absence of strong nonnative contacts (salt bridges or hydrophobic collapse) and limited repulsion of charged side chains.

A new family of Ni4 and Ni6 aggregates from the self-assembly of [Ni2] building units: role of carboxylate and carbonate bridges.

PubMed

Pait, Moumita; Bauzá, Antonio; Frontera, Antonio; Colacio, Enrique; Ray, Debashis

2015-05-18

Carboxylato (R = (t)Bu and Et) and carbonato bridges have been utilized for nickel(II)-based aggregates [Ni4(μ-H2L)2(μ3-OH)2(μ1,3-O2CBu(t))2](NO3)2·H2O·2DMF (1·H2O·2DMF), Ni4(μ-(hy)HL)2(μ3-OMe)2(μ1,1-N3)2(μ1,3-O2CEt)2]·4H2O (2·4H2O), and Ni6(μ4-L)(μ3-L)2(μ6-CO3)(H2O)8](ClO4)·9H2O (3·9H2O). Building blocks [Ni2(μ-H2L)](3+), [Ni2(μ-(hy)HL)](3+), and [Ni2(μ-L)](+) originating from [Ni2(μ-H2L)](3+) have been trapped in these complexes. The complexes have been characterized by X-ray crystallography, magnetic measurements, and density functional theory (DFT) analysis. In 1, the magnetic interactions are transmitted through the μ3-phenoxido/μ3-hydroxido/syn-syn-(t)BuCO2(-), μ3-phenoxido/μ3- hydroxido, and double μ3-phenoxido/double μ3-hydroxido bridges with J = +11.4 cm(-1), J1 = -2.1 cm(-1), and J2 = -2.8 cm(-1), respectively. In 2, the interactions are ferromagnetic, with J1 = +27.5 cm(-1), J2 = +20.62 cm(-1), and J3 = +1.52 cm(-1) describing the magnetic couplings through the μ-phenoxidoo/μ3-methoxido, μ-azido/μ3-methoxido, and μ3-methoxido/μ3-methoxido exchange pathways, respectively. Complex 3 gives J1 = -3.30 cm(-1), J2 = +1.7 cm(-1), and J3 = -12.8 cm(-1) for exchange pathways mediated by μ-phenoxido/μ-carbonato, μ-alkoxido/μ-alkooxido/μ-syn-syn-carbonato, and the μ-phenoxido/μ-carbonato, respectively. Interestingly, 1 and 3 below 20 K and 35 K, respectively, show an abrupt increase of the χMT product to reach a magnetic-field-dependent maximum, which is associated with a slightly frequency-dependent out-of-phase alternating-current peak. DFT calculations have also been performed on 1-3 to explain the exchange interaction mechanisms and to support the magnitude and sign of the magnetic coupling constants between the Ni(II) ions.

Stereodivergent Synthesis of 1,3-Syn-Polyol Natural Product for Stereochemical-Based Structure Activity Relationship Studies

NASA Astrophysics Data System (ADS)

Zheng, Jiamin

The 1,3-syn-diol functionality is very common in many natural products. An important class containing this moiety are the 1,3-syn-polyol/pyranone natural products, which have been isolated from a variety of plant sources, and possess biological activities like plant growth inhibition as well as antifeedant, antifungal, antibacterial, and antitumor properties. The feature of this class is a 6-membered lactone where the lactoe oxygen is part of a 1,3-syn-diol motif. To pursue the 1,3-syn-polyol/pyranone natural products, an iterative hydration of polyene strategy was utilized to provide the 1,3- syn-diol functionality, and asymmetric synthetic strategies were explored to form the requisite stereochemistry. The versatility of the asymmetric approach was demonstrated in the synthesis of eupatorium pyranone and also in an ongoing project aimed at the synthesis of SIA7248. As an outgrowth of our work on the total syntheses of 1,3-syn -polyol natural products inspired a stereo-divergent synthesis of 1,3-syn-polyol natural products and their analogs for stereochemical-based structure-activity relationship (SSAR) studies. To identify the key structural factors important for the anticancer activity of the 1,3-syn-polyol/pyranones, a stereo-divergent 16-member library of pyranone/polyol congeners was designed, synthesized and tested with variations in both stereochemistry and numbers of polyol repeat units. Having access to stereochemical isomers of the biologically active natural products allowed us to design experiments that help illustrate their mechanisms of action.

Age- and brain region-dependent α-synuclein oligomerization is attributed to alterations in intrinsic enzymes regulating α-synuclein phosphorylation in aging monkey brains.

PubMed

Chen, Min; Yang, Weiwei; Li, Xin; Li, Xuran; Wang, Peng; Yue, Feng; Yang, Hui; Chan, Piu; Yu, Shun

2016-02-23

We previously reported that the levels of α-syn oligomers, which play pivotal pathogenic roles in age-related Parkinson's disease (PD) and dementia with Lewy bodies, increase heterogeneously in the aging brain. Here, we show that exogenous α-syn incubated with brain extracts from older cynomolgus monkeys and in Lewy body pathology (LBP)-susceptible brain regions (striatum and hippocampus) forms higher amounts of phosphorylated and oligomeric α-syn than that in extracts from younger monkeys and LBP-insusceptible brain regions (cerebellum and occipital cortex). The increased α-syn phosphorylation and oligomerization in the brain extracts from older monkeys and in LBP-susceptible brain regions were associated with higher levels of polo-like kinase 2 (PLK2), an enzyme promoting α-syn phosphorylation, and lower activity of protein phosphatase 2A (PP2A), an enzyme inhibiting α-syn phosphorylation, in these brain extracts. Further, the extent of the age- and brain-dependent increase in α-syn phosphorylation and oligomerization was reduced by inhibition of PLK2 and activation of PP2A. Inversely, phosphorylated α-syn oligomers reduced the activity of PP2A and showed potent cytotoxicity. In addition, the activity of GCase and the levels of ceramide, a product of GCase shown to activate PP2A, were lower in brain extracts from older monkeys and in LBP-susceptible brain regions. Our results suggest a role for altered intrinsic metabolic enzymes in age- and brain region-dependent α-syn oligomerization in aging brains.

The proinflammatory cytokines IL-1beta and TNF-alpha induce the expression of Synoviolin, an E3 ubiquitin ligase, in mouse synovial fibroblasts via the Erk1/2-ETS1 pathway.

PubMed

Gao, Beixue; Calhoun, Karen; Fang, Deyu

2006-01-01

The overgrowth of synovial tissues is critical in the pathogenesis of rheumatoid arthritis (RA). The expression of Synoviolin (SYN), an E3 ubiquitin ligase, is upregulated in arthritic synovial fibroblasts and is involved in the overgrowth of synovial cells during RA. However, the molecular mechanisms involved in the elevated SYN expression are not known. Here, we found that SYN expression is elevated in the synovial fibroblasts from mice with collagen-induced arthritis (CIA). The proinflammatory cytokines interleukin (IL)-1beta and tumor necrosis factor-alpha (TNF-alpha) induce SYN expression in mouse synovial fibroblasts. Cultivation of mouse synovial fibroblasts with IL-1beta activates mitogen-activated protein kinases, including extra-cellular signal-regulated kinase (Erk), JNK (c-Jun N-terminal kinase), and p38, while only Erk-specific inhibitor blocks IL-1beta-induced SYN expression. Expression of transcription factor ETS1 further enhances IL-1beta-induced SYN expression. The dominant negative ETS1 mutant lacking the transcription activation domain inhibits SYN expression in a dose-dependent manner. The activation of both Erk1/2 and ETS1 is increased in the CIA synovial fibroblasts. Inhibition of Erk activation reduces ETS1 phosphorylation and SYN expression. Our data indicate that the proinflammatory cytokines IL-1beta and TNF-alpha induce the overgrowth of synovial cells by upregulating SYN expression via the Erk1/-ETS1 pathway. These molecules or pathways could therefore be potential targets for the treatment of RA.

The proinflammatory cytokines IL-1β and TNF-α induce the expression of Synoviolin, an E3 ubiquitin ligase, in mouse synovial fibroblasts via the Erk1/2-ETS1 pathway

PubMed Central

Gao, Beixue; Calhoun, Karen; Fang, Deyu

2006-01-01

The overgrowth of synovial tissues is critical in the pathogenesis of rheumatoid arthritis (RA). The expression of Synoviolin (SYN), an E3 ubiquitin ligase, is upregulated in arthritic synovial fibroblasts and is involved in the overgrowth of synovial cells during RA. However, the molecular mechanisms involved in the elevated SYN expression are not known. Here, we found that SYN expression is elevated in the synovial fibroblasts from mice with collagen-induced arthritis (CIA). The proinflammatory cytokines interleukin (IL)-1β and tumor necrosis factor-α (TNF-α) induce SYN expression in mouse synovial fibroblasts. Cultivation of mouse synovial fibroblasts with IL-1β activates mitogen-activated protein kinases, including extra-cellular signal-regulated kinase (Erk), JNK (c-Jun N-terminal kinase), and p38, while only Erk-specific inhibitor blocks IL-1β-induced SYN expression. Expression of transcription factor ETS1 further enhances IL-1β-induced SYN expression. The dominant negative ETS1 mutant lacking the transcription activation domain inhibits SYN expression in a dose-dependent manner. The activation of both Erk1/2 and ETS1 is increased in the CIA synovial fibroblasts. Inhibition of Erk activation reduces ETS1 phosphorylation and SYN expression. Our data indicate that the proinflammatory cytokines IL-1β and TNF-α induce the overgrowth of synovial cells by upregulating SYN expression via the Erk1/-ETS1 pathway. These molecules or pathways could therefore be potential targets for the treatment of RA. PMID:17105652

Nomenclatural Studies Toward a World List of Diptera Genus-Group Names. Part IV: Charles Henry Tyler Townsend.

PubMed

Evenhuis, Neal L; Pont, Adrian C; Whitmore, Daniel

2015-06-25

The Diptera genus-group names of Charles Henry Tyler Townsend are reviewed and annotated. A total of 1506 available genus-group names in 12 families of Diptera are listed alphabetically for each name, giving author, year and page of original publication, originally included species, type species and method of fixation, current status of the name, family placement, and a list of any emendations of it that have been found in the literature. Remarks are given to clarify nomenclatural and/or taxonomic information. In addition, an index to all the species-group names of Diptera proposed by Townsend (1595, of which 1574 are available names) is given with bibliographic reference (year and page) to each original citation. An appendix with a full bibliography of almost 650 papers written by Townsend is presented with accurate dates of publication.        Two new replacement names are proposed for preoccupied genus-group names and both are named to honor our good friend and colleague, James E. O'Hara, for his decades of work on tachinids: Oharamyia Evenhuis, Pont & Whitmore, n. name, for Lindigia Townsend, 1931 [Tachinidae] (preoccupied by Karsten, 1858); Jimimyia Evenhuis, Pont & Whitmore, n. name, for Siphonopsis Townsend, 1916 [Tachinidae] (preoccupied by Agassiz, 1846).        Earlier dates of availability are found for the following: Eucnephalia Townsend, 1892 [Tachinidae]; Gabanimyia Townsend, 1914 [Tachinidae]; Incamyia Townsend, 1912 [Tachinidae]; Muscopteryx Townsend, 1892 [Tachinidae]; Philippolophosia Townsend, 1927 [Tachinidae]; Pseudokea Townsend, 1927 [Tachinidae].        Corrected or clarified included species and/or corrected or clarified type-species and methods of typification are given for: Alitophasia Townsend, 1934 [Tachinidae]; Almugmyia Townsend, 1911 [Tachinidae]; Arachnidomyia Townsend, 1934 [Sarcophagidae]; Austenina Townsend, 1921 [Glossinidae]; Austrohartigia Townsend, 1937 [Sarcophagidae]; Awatia Townsend, 1921 [Muscidae]; Azygobothria Townsend, 1911 [Tachinidae]; Brachymasicera Townsend, 1911 [Tachinidae]; Calocarcelia Townsend, 1927 [Tachinidae]; Cnephalodes Townsend, 1911 [Tachinidae]; Cyacyrtoneura Townsend, 1931 [Muscidae]; Cyrtoneuropsis Townsend, 1931 [Muscidae]; Cyrtosoma Brauer & Bergenstamm, 1893 [Tachinidae]; Epiphyllophila Townsend, 1927 [Tachinidae]; Eucalodexia Townsend, 1892 [Tachinidae]; Eumesembrina Townsend, 1908 [Muscidae]; Eumyobia Townsend, 1911 [Tachinidae]; Eusisyropa Townsend, 1908 [Tachinidae]; Gabanimyia Townsend, 1914 [Tachinidae]; Galactomyia Townsend, 1908 [Tachinidae]; Girschneria Townsend, 1919 [Tachinidae]; Gymnochaetopsis Townsend, 1914 [Tachinidae]; Himantostomopsis Townsend, 1921 [Tachinidae]; Incamyia Townsend, 1912 [Tachinidae]; Lithoexorista Townsend, 1921 [Tachinidae]; Muscopteryx Townsend, 1892 [Tachinidae]; Myocuphocera Townsend, 1931 [Tachinidae]; Myxexoristops Townsend, 1911 [Tachinidae]; Neojurinia Townsend, 1914 [Tachinidae]; Newsteadina Townsend, 1921 [Glossinidae]; Ommasicera Townsend, 1911 [Tachinidae]; Ophirion Townsend, 1911 [Tachinidae]; Ophiriodexia Townsend, 1911 [Tachinidae]; Ophiriosturmia Townsend, 1911 [Tachinidae]; Opsozelia Townsend, 1919 [Tachinidae]; Paleotachina Townsend, 1921 [Tachinidae]; Palexorista Townsend, 1921 [Tachinidae]; Phasiatacta Townsend, 1911 [Tachinidae]; Philippolophosia Townsend, 1927 [Tachinidae]; Phrissopolia Townsend, 1908 [Tachinidae]; Pseudokea Townsend, 1927 [Tachinidae]; Pygocalcager Townsend, 1935 [Tachinidae]; Trichobius Townsend, 1891 [Hippoboscidae]; Villeneuvia Townsend, 1921 [Tachinidae]; Zonoepalpus Townsend, 1927 [Tachinidae]; Zygosturmia Townsend, 1911 [Tachinidae].        The following names previously treated as available are shown to be unavailable.-Genera: Denatella Townsend, 1931, n. stat. [Calliphoridae]; Epseudocyptera Townsend, 1927, n. stat. [Tachinidae]; Eustomatodexia Townsend, 1892, n. stat. [Tachinidae].-Species: Epseudocyptera epalpata Townsend, 1927, n. stat. [Tachinidae]; Eustomatodexia insulensis Townsend, 1892, n. stat. [Tachinidae].       The following genus-group names, not listed in previous regional catalogs, are treated here: Arabisca Townsend, 1935 [Sarcophagidae]; Eupeleteria Townsend, 1908 [Tachinidae]; Macropatelloa Townsend, 1931 [Tachinidae]; Neohypostena Townsend, 1915 [Tachinidae]; Neometapodia Townsend, 1892 [Sarcophagidae]; Tricyclopsis Townsend, 1916 [Calliphoridae]; Trongia Townsend, 1916 [Calliphoridae].        Previous First Reviser actions for multiple original spellings that were overlooked by other workers are given for the following: Genus-group names-Microchaetona Townsend, 1919 [Tachinidae]; Neopodomyia Townsend, 1927 [Tachinidae]; Opsophytopsis Townsend, 1918 [Sarcophagidae]; Prohypotachina Townsend, 1933 [Tachinidae]; Rhinomyodes Townsend, 1933 [Tachinidae]; Servilliodes Townsend, 1926 [Tachinidae]; Tephromyiella Townsend, 1918 [Sarcophagidae]; Thelairochaetona Townsend, 1919 [Tachinidae]; Xanthopteromyia Townsend, 1926 [Tachinidae]. Species-group names-Brachybelvosia brasiliensis Townsend, 1927 [Tachinidae]; Neocraspedothrix nova Townsend, 1927 [Tachinidae].        The following nominal genera enter into new synonymies: Bathytheresia Townsend, 1915 under Billaea Robineau-Desvoidy, 1830, n. syn. [Tachinidae]; Brachycoma Brauer & Bergenstamm, 1889 under Brachicoma Rondani, 1856, n. syn. [Sarcophagidae]; Chaetolyga Brauer, 1880 under Carcelia Robineau-Desvoidy, 1830, n. syn. [Tachinidae]; Chaetoprosopa Marschall, 1873 under Choeteprosopa Macquart, 1851, n. syn. [Tachinidae]; Chlororhynchomyia Senior-White, Aubertin & Smart, 1940 under Metallea Wulp, 1880, n. syn. [Rhiniidae]; Chrysomyia Macquart, 1835 under Chrysomya Robineau-Desvoidy, 1830, n. syn. [Calliphoridae]; Echinomyia Fischer von Waldheim, 1808 under Echinomya Latreille, 1805, n. syn. [Tachinidae]; Euhypochaetopsis Townsend, 1928 under Campylocheta Rondani, 1859, n. syn. [Tachinidae]; Graphomyia Macquart, 1834 under Graphomya Robineau-Desvoidy, 1830, n. syn. [Muscidae]; Kurintjimyia Townsend, 1926 under Tachina Meigen, 1803, n. syn. [Tachinidae]; Labidigaster Macquart, 1844 under Labigastera Macquart, 1834, n. syn. [Tachinidae]; Mellanactia Guimarães, 1971 under Oxynops Townsend, 1912, n. syn. [Tachinidae]; Ochromia Townsend, 1935 under Bengalia Robineau-Desvoidy, 1830, n. syn. [Tachinidae]; Pachyrrhina Osten Sacken, 1881 under Nephrotoma Meigen, 1803, n. syn. [Tipulidae]; Procraspedothrix Townsend, 1932 under Phytomyptera Rondani, 1844, n. syn. [Tachinidae]; Pseudogymnosoma Townsend, 1918 under Neomyia Walker, 1859, n. syn. [Muscidae]; Pseudoservillia Townsend, 1916 under Tachina Meigen, 1803, n. syn. [Tachinidae]; Rhymosia Mik, 1886 under Rymosia Winnertz, 1863, n. syn. [Mycetophilidae]; Rhynchomyia Macquart, 1835 under Rhyncomya Robineau-Desvoidy, 1830, n. syn. [Rhiniidae]; Servillioides Townsend, 1926 under Tachina Meigen, 1803, n. syn. [Tachinidae]; Servilliopsis Townsend, 1916 under Tachina Meigen, 1803, n. syn. [Tachinidae]; Stephanostoma Cole, 1923 under Bercaea Robineau-Desvoidy, 1863, n. syn. [Sarcophagidae]; Stomatorhinia Townsend, 1935 under Stomorhina Rondani, 1861, n. syn. [Rhiniidae]; Toxorrhina Osten Sacken, 1869 under Toxorhina Loew, 1850, n. syn. [Limoniidae]; Trichoneura Townsend, 1935 under Stevenia Robineau-Desvoidy, 1830, n. syn. [Rhinophoridae]; Trichopticus Schnabl, 1889 under Thricops Rondani, 1856, n. syn. [Muscidae]; Tricyclopsis Townsend, 1916 under Paracalliphora Townsend, 1916, n. syn. [Calliphoridae].

Mapping the Emergence of Synthetic Biology

PubMed Central

2016-01-01

In this paper, we apply an original scientometric analyses to a corpus comprising synthetic biology (SynBio) publications in Thomson Reuters Web of Science to characterize the emergence of this new scientific field. Three results were drawn from this empirical investigation. First, despite the exponential growth of publications, the study of population level statistics (newcomers proportion, collaboration network structure) shows that SynBio has entered a stabilization process since 2010. Second, the mapping of textual and citational networks shows that SynBio is characterized by high heterogeneity and four different approaches: the central approach, where biobrick engineering is the most widespread; genome engineering; protocell creation; and metabolic engineering. We suggest that synthetic biology acts as an umbrella term allowing for the mobilization of resources, and also serves to relate scientific content and promises of applications. Third, we observed a strong intertwinement between epistemic and socio-economic dynamics. Measuring scientific production and impact and using structural analysis data, we identified a core set of mostly American scientists. Biographical analysis shows that these central and influential scientists act as “boundary spanners,” meaning that their importance to the field lies not only in their academic contributions, but also in their capacity to interact with other social spaces that are outside the academic sphere. PMID:27611324

SU-G-JeP2-06: Dosimetric and Workflow Evaluation of First Commercial Synthetic CT Software for Clinical Use in Pelvis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tyagi, N; Zhang, J; Happersett, L

2016-06-15

Purpose: evaluate a commercial synthetic CT (syn-CT) software for use in prostate radiotherapy Methods: Twenty prostate patients underwent CT and MR simulation scans in treatment position on a 3T Philips scanner. The MR protocol consisted of a T2w turbo spin-echo for soft tissue contrast, a 2D balanced-fast field echo (b-FFE) for fiducial identification, a dual-echo 3D FFE B0 map for distortion analysis and a 3D mDIXON FFE sequence to generate syn-CT. Two echoes are acquired during mDIXON scan, allowing water, fat, and in-phase images to be derived using the frequency shift of the fat and water protons. Tissues were classifiedmore » as: air, adipose, water, trabecular/spongy bone and compact/cortical bone and assigned specific bulk HU values. Bone structures are segmented based on a pelvis bone atlas. Accuracy of syn-CT for patient treatment planning was analyzed by transferring the original plan and structures from the CT to syn-CT via rigid registration and recalculating dose. In addition, new IMRT plans were generated on the syn-CT using structures contoured on MR and transferred to the syn-CT. Accuracy of fiducial-based localization at the treatment machine performed using syn-CT or DRRs generated from syn-CT was assessed by comparing to orthogonal kV radiographs or CBCT. Results: Dosimetric comparison between CT and syn-CT was within 0.5% for all structures. The de-novo optimized plans generated on the syn-CT met our institutional clinical objectives for target and normal structures. Patient-induced susceptibility distortion based on B0 maps was within 1mm and 0.4 mm in the body and prostate. The rectal and bladder outlines on the syn-CT were deemed sufficient for assessing rectal and bladder filling on the CBCT at the time of treatment. CBCT localization showed a median error of < ±1 mm in LR, AP and SI direction. Conclusion: MRI derived syn-CT can be used clinically in MR-alone planning and treatment process for prostate. Drs. Deasy, Hunt and Tyagi have Master research agreement with Philips healthcare.« less

Unique active site promotes error-free replication opposite an 8-oxo-guanine lesion by human DNA polymerase iota

PubMed Central

Kirouac, Kevin N.; Ling, Hong

2011-01-01

The 8-oxo-guanine (8-oxo-G) lesion is the most abundant and mutagenic oxidative DNA damage existing in the genome. Due to its dual coding nature, 8-oxo-G causes most DNA polymerases to misincorporate adenine. Human Y-family DNA polymerase iota (polι) preferentially incorporates the correct cytosine nucleotide opposite 8-oxo-G. This unique specificity may contribute to polι’s biological role in cellular protection against oxidative stress. However, the structural basis of this preferential cytosine incorporation is currently unknown. Here we present four crystal structures of polι in complex with DNA containing an 8-oxo-G lesion, paired with correct dCTP or incorrect dATP, dGTP, and dTTP nucleotides. An exceptionally narrow polι active site restricts the purine bases in a syn conformation, which prevents the dual coding properties of 8-oxo-G by inhibiting syn/anti conformational equilibrium. More importantly, the 8-oxo-G base in a syn conformation is not mutagenic in polι because its Hoogsteen edge does not form a stable base pair with dATP in the narrow active site. Instead, the syn 8-oxo-G template base forms the most stable replicating base pair with correct dCTP due to its small pyrimidine base size and enhanced hydrogen bonding with the Hoogsteen edge of 8-oxo-G. In combination with site directed mutagenesis, we show that Gln59 in the finger domain specifically interacts with the additional O8 atom of the lesion base, which influences nucleotide selection, enzymatic efficiency, and replication stalling at the lesion site. Our work provides the structural mechanism of high-fidelity 8-oxo-G replication by a human DNA polymerase. PMID:21300901

An intercomparison and validation of satellite-based surface radiative energy flux estimates over the Arctic

NASA Astrophysics Data System (ADS)

Riihelä, Aku; Key, Jeffrey R.; Meirink, Jan Fokke; Kuipers Munneke, Peter; Palo, Timo; Karlsson, Karl-Göran

2017-05-01

Accurate determination of radiative energy fluxes over the Arctic is of crucial importance for understanding atmosphere-surface interactions, melt and refreezing cycles of the snow and ice cover, and the role of the Arctic in the global energy budget. Satellite-based estimates can provide comprehensive spatiotemporal coverage, but the accuracy and comparability of the existing data sets must be ascertained to facilitate their use. Here we compare radiative flux estimates from Clouds and the Earth's Radiant Energy System (CERES) Synoptic 1-degree (SYN1deg)/Energy Balanced and Filled, Global Energy and Water Cycle Experiment (GEWEX) surface energy budget, and our own experimental FluxNet / Satellite Application Facility on Climate Monitoring cLoud, Albedo and RAdiation (CLARA) data against in situ observations over Arctic sea ice and the Greenland Ice Sheet during summer of 2007. In general, CERES SYN1deg flux estimates agree best with in situ measurements, although with two particular limitations: (1) over sea ice the upwelling shortwave flux in CERES SYN1deg appears to be underestimated because of an underestimated surface albedo and (2) the CERES SYN1deg upwelling longwave flux over sea ice saturates during midsummer. The Advanced Very High Resolution Radiometer-based GEWEX and FluxNet-CLARA flux estimates generally show a larger range in retrieval errors relative to CERES, with contrasting tendencies relative to each other. The largest source of retrieval error in the FluxNet-CLARA downwelling shortwave flux is shown to be an overestimated cloud optical thickness. The results illustrate that satellite-based flux estimates over the Arctic are not yet homogeneous and that further efforts are necessary to investigate the differences in the surface and cloud properties which lead to disagreements in flux retrievals.

RNA interference targeting α-synuclein attenuates methamphetamine-induced neurotoxicity in SH-SY5Y cells.

PubMed

Chen, Ling; Huang, Enping; Wang, Huijun; Qiu, Pingming; Liu, Chao

2013-07-12

The protein α-synuclein (α-syn) is abundant in neurons and has been claimed to play critical roles in the pathophysiology of Parkinson's disease. Overexpression of α-syn has been shown to be toxicity in methamphetamine (METH)-induced model in vivo and in vitro which has Parkinson's-like pathology. However, the exact mechanisms underlying toxicity of α-syn mediated METH-induced neuron remain unknown. In the present study, human dopaminergic-like neuroblastoma SH-SY5Y cells were used as METH-induced model in vitro. Cell viability was found to be dramatically increased after silencing α-syn expression followed by METH treatment compared with a-syn wild-type cells and the morphological damage to cells after METH treatment was abated through knockdown of α-syn expression in this model. The expression levels of tyrosine hydroxylase (TH), dopamine transporter (DAT) and vesicular monoamine transporter 2(VMAT-2) were significantly decreased and the activity/levels of reactive oxygen species (ROS), nitric oxide synthase (NOS) and nitrogen (NO) were notably increased after METH treatment. However, the changes of these expression levels were reversed in cells transfected with α-syn-shRNA. These results suggested that TH, DAT, VMAT-2, ROS and NOS maybe involved in α-syn mediated METH-induced neuronal toxicity. Copyright © 2013 Elsevier B.V. All rights reserved.

An alpha-synuclein MRM assay with diagnostic potential for Parkinson's disease and monitoring disease progression.

PubMed

Yang, Li; Stewart, Tessandra; Shi, Min; Pottiez, Gwenael; Dator, Romel; Wu, Rui; Aro, Patrick; Schuster, Robert J; Ginghina, Carmen; Pan, Catherine; Gao, Yuqian; Qian, Weijun; Zabetian, Cyrus P; Hu, Shu-Ching; Quinn, Joseph F; Zhang, Jing

2017-07-01

The alpha-synuclein (α-syn) level in human cerebrospinal fluid (CSF), as measured by immunoassays, is promising as a Parkinson's disease (PD) biomarker. However, the levels of total α-syn are inconsistent among studies with large cohorts and different measurement platforms. Total α-syn level also does not correlate with disease severity or progression. Here, the authors developed a highly sensitive MRM method to measure absolute CSF α-syn peptide concentrations without prior enrichment or fractionation, aiming to discover new candidate biomarkers. Six peptides covering 73% of protein sequence were reliably identified, and two were consistently quantified in cross-sectional and longitudinal cohorts. Absolute concentration of α-syn in human CSF was determined to be 2.1 ng/mL. A unique α-syn peptide, TVEGAGSIAAATGFVK (81-96), displayed excellent correlation with previous immunoassay results in two independent PD cohorts (p < 0.001), correlated with disease severity, and its changes significantly tracked the disease progression longitudinally. An MRM assay to quantify human CSF α-syn was developed and optimized. Sixty clinical samples from cross-sectional and longitudinal PD cohorts were analyzed with this approach. Although further larger scale validation is needed, the results suggest that α-syn peptide could serve as a promising biomarker in PD diagnosis and progression. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

An alpha-synuclein MRM assay with diagnostic potential for Parkinson's disease and monitoring disease progression

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yang, Li; Stewart, Tessandra; Shi, Min

Aim: The alpha-synuclein (α-syn) level in human cerebrospinal fluid (CSF), as measured by immunoassays, is promising as a Parkinson’s disease (PD) biomarker. However, the levels of total α-syn are inconsistent among studies with large cohorts and different measurement platforms. Total α-syn level also does not correlate with disease severity or progression. Here, we developed a highly sensitive Multiple Reaction Monitoring (MRM) method to measure absolute CSF α-syn peptide concentrations without prior enrichment or fractionation, aiming to discover new candidate biomarkers. Results: Six peptides covering 73% of protein sequence were reliably identified, and two were consistently quantified in cross-sectional and longitudinalmore » cohorts. Absolute concentration of α-syn in human CSF was determined to be 2.1ng/mL. A unique α-syn peptide, TVEGAGSIAAATGFVK (81-96), displayed excellent correlation with previous immunoassay results in two independent PD cohorts (p < 0.001), correlated with disease severity, and its changes significantly tracked the disease progression longitudinally. Conclusions: An MRM assay to quantify human CSF α-syn was developed and optimized. Sixty clinical samples from cross-sectional and longitudinal PD cohorts were analyzed with this approach. Although further larger-scale validation is needed, the results suggest that α-syn peptide could serve as a promising biomarker in PD diagnosis and progression.« less

Enrichment of anaerobic syngas-converting bacteria from thermophilic bioreactor sludge.

PubMed

Alves, Joana I; Stams, Alfons J M; Plugge, Caroline M; Alves, M Madalena; Sousa, Diana Z

2013-12-01

Thermophilic (55 °C) anaerobic microbial communities were enriched with a synthetic syngas mixture (composed of CO, H2 , and CO2 ) or with CO alone. Cultures T-Syn and T-CO were incubated and successively transferred with syngas (16 transfers) or CO (9 transfers), respectively, with increasing CO partial pressures from 0.09 to 0.88 bar. Culture T-Syn, after 4 successive transfers with syngas, was also incubated with CO and subsequently transferred (9 transfers) with solely this substrate - cultures T-Syn-CO. Incubation with syngas and CO caused a rapid decrease in the microbial diversity of the anaerobic consortium. T-Syn and T-Syn-CO showed identical microbial composition and were dominated by Desulfotomaculum and Caloribacterium species. Incubation initiated with CO resulted in the enrichment of bacteria from the genera Thermincola and Thermoanaerobacter. Methane was detected in the first two to three transfers of T-Syn, but production ceased afterward. Acetate was the main product formed by T-Syn and T-Syn-CO. Enriched T-CO cultures showed a two-phase conversion, in which H2 was formed first and then converted to acetate. This research provides insight into how thermophilic anaerobic communities develop using syngas/CO as sole energy and carbon source can be steered for specific end products and subsequent microbial synthesis of chemicals. © 2013 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

A checklist of Chinese crickets (Orthoptera: Gryllidea).

PubMed

He, Zhu-Qing

2018-01-07

A checklist of Chinese crickets, including Taiwan, is offered. Presently 331 species or subspecies have been reported including true crickets, scale crickets, ant crickets and mole crickets belonging to 6 families, 16 subfamilies and 83 genera. Modicogryllus (Modicogryllus) maculatus (Shiraki, 1930) is moved to Comidoblemmus as C. maculatus (Shiraki, 1930) comb. nov. Velarifictorus (Velarifictorus) koshunensis (Shiraki, 1911) is moved to Turanogryllus as T. koshunensis (Shiraki, 1911) comb. nov. Qingryllus Chen Zheng, 1995 syn. is the junior synonym of Goniogryllus Chopard, 1936. Loxoblemmus angulatus Bey-Bienko, 1956 syn. is the junior synonym of Loxoblemmus appendicularis Shiraki, 1930. Cophogryllus kuhlgatzi Karny, 1908 syn. is the junior synonym of Teleogryllus (Brachyteleogryllus) occipitalis occipitalis (Serville, 1838). Velarifictorus (Velarifictorus) aspersus borealis Gorochov, 1985 syn. is the junior synonym of Velarifictorus (Velarifictorus) aspersus aspersus (Walker, 1869). Modicogryllus (Modicogryllus) latefasciatus (Chopard, 1933) syn. is the junior synonym of Velarifictorus (Velarifictorus) micado (Saussure, 1877). Velarifictorus (Velarifictorus) ornatus caudatus (Shiraki, 1930) syn. is the junior synonym of Velarifictorus (Velarifictorus) ornatus ornatus (Shiraki, 1911). Dianemobius nigrofasciatus (Matsumura, 1904) syn. is the junior synonym of Dianemobius fascipes (Walker, 1869). Polionemobius mikado (Shiraki, 1911) syn. is the junior synonym of Polionemobius taprobanensis (Walker, 1869). Vietacheta picea Gorochov, 1992, Oecanthus euryelytra Ichikawa, 2001, Oecanthus similator Ichikawa, 2001, Xabea levissima Gorochov, 1992, Pteronemobius (Pteronemobius) yezoensis (Shiraki, 1911), Metioche (Metioche) japonica (Ichikawa, 2001), Natula matsuurai Sugimoto, 2001 are the first records from China.

DNA damage preceding dopamine neuron degeneration in A53T human α-synuclein transgenic mice

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Degui; Yu, Tianyu; Liu, Yongqiang

Defective DNA repair has been linked with age-associated neurodegenerative disorders. Parkinson's disease (PD) is a progressive neurodegenerative disorder caused by genetic and environmental factors. Whether damages to nuclear DNA contribute to neurodegeneration of PD still remain obscure. in this study we aim to explore whether nuclear DNA damage induce dopamine neuron degeneration in A53T human α-Synuclein over expressed mouse model. We investigated the effects of X-ray irradiation on A53T-α-Syn MEFs and A53T-α-Syn transgene mice. Our results indicate that A53T-α-Syn MEFs show a prolonged DNA damage repair process and senescense phenotype. DNA damage preceded onset of motor phenotype in A53T-α-Syn transgenicmore » mice and decrease the number of nigrostriatal dopaminergic neurons. Neurons of A53T-α-Syn transgenic mice are more fragile to DNA damages. - Highlights: • This study explore contribution of DNA damage to neurodegeneration in Parkinson's disease mice. • A53T-α-Syn MEF cells show a prolonged DNA damage repair process and senescense phenotype. • DNA damage preceded onset of motor phenotype in A53T-α-Syn transgenic mice. • DNA damage decrease the number of nigrostriatal dopaminergic neurons. • Neurons of A53T-α-Syn transgenic mice are more fragile to DNA damages.« less

Phospholipase D1 downregulation by α-synuclein: Implications for neurodegeneration in Parkinson's disease.

PubMed

Conde, Melisa A; Alza, Natalia P; Iglesias González, Pablo A; Scodelaro Bilbao, Paola G; Sánchez Campos, Sofía; Uranga, Romina M; Salvador, Gabriela A

2018-06-01

We have previously shown that phospholipase D (PLD) pathways have a role in neuronal degeneration; in particular, we found that PLD activation is associated with synaptic injury induced by oxidative stress. In the present study, we investigated the effect of α-synuclein (α-syn) overexpression on PLD signaling. Wild Type (WT) α-syn was found to trigger the inhibition of PLD1 expression as well as a decrease in ERK1/2 phosphorylation and expression levels. Moreover, ERK1/2 subcellular localization was shown to be modulated by WT α-syn in a PLD1-dependent manner. Indeed, PLD1 inhibition was found to alter the neurofilament network and F-actin distribution regardless of the presence of WT α-syn. In line with this, neuroblastoma cells expressing WT α-syn exhibited a degenerative-like phenotype characterized by a marked reduction in neurofilament light subunit (NFL) expression and the rearrangement of the F-actin organization, compared with either the untransfected or the empty vector-transfected cells. The gain of function of PLD1 through the overexpression of its active form had the effect of restoring NFL expression in WT α-syn neurons. Taken together, our findings reveal an unforeseen role for α-syn in PLD regulation: PLD1 downregulation may constitute an early mechanism in the initial stages of WT α-syn-triggered neurodegeneration. Copyright © 2018 Elsevier B.V. All rights reserved.

Synaptic activity induces input-specific rearrangements in a targeted synaptic protein interaction network.

PubMed

Lautz, Jonathan D; Brown, Emily A; VanSchoiack, Alison A Williams; Smith, Stephen E P

2018-05-27

Cells utilize dynamic, network level rearrangements in highly interconnected protein interaction networks to transmit and integrate information from distinct signaling inputs. Despite the importance of protein interaction network dynamics, the organizational logic underlying information flow through these networks is not well understood. Previously, we developed the quantitative multiplex co-immunoprecipitation platform, which allows for the simultaneous and quantitative measurement of the amount of co-association between large numbers of proteins in shared complexes. Here, we adapt quantitative multiplex co-immunoprecipitation to define the activity dependent dynamics of an 18-member protein interaction network in order to better understand the underlying principles governing glutamatergic signal transduction. We first establish that immunoprecipitation detected by flow cytometry can detect activity dependent changes in two known protein-protein interactions (Homer1-mGluR5 and PSD-95-SynGAP). We next demonstrate that neuronal stimulation elicits a coordinated change in our targeted protein interaction network, characterized by the initial dissociation of Homer1 and SynGAP-containing complexes followed by increased associations among glutamate receptors and PSD-95. Finally, we show that stimulation of distinct glutamate receptor types results in different modular sets of protein interaction network rearrangements, and that cells activate both modules in order to integrate complex inputs. This analysis demonstrates that cells respond to distinct types of glutamatergic input by modulating different combinations of protein co-associations among a targeted network of proteins. Our data support a model of synaptic plasticity in which synaptic stimulation elicits dissociation of preexisting multiprotein complexes, opening binding slots in scaffold proteins and allowing for the recruitment of additional glutamatergic receptors. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

A modified grapefruit juice eliminates two compound classes as major mediators of the Grapefruit Juice-Fexofenadine Interaction: an In Vitro-In Vivo 'Connect'

USDA-ARS?s Scientific Manuscript database

The grapefruit juice (GFJ)-fexofenadine interaction involves inhibition of intestinal organic anion transporting polypeptide (OATP)-mediated active uptake of fexofenadine by GFJ, manifesting as a decrease in systemic drug exposure. Flavonoids, furanocoumarins, and polymethoxyflavones have been ident...

SynGenics Optimization System (SynOptSys)

NASA Technical Reports Server (NTRS)

Ventresca, Carol; McMilan, Michelle L.; Globus, Stephanie

2013-01-01

The SynGenics Optimization System (SynOptSys) software application optimizes a product with respect to multiple, competing criteria using statistical Design of Experiments, Response-Surface Methodology, and the Desirability Optimization Methodology. The user is not required to be skilled in the underlying math; thus, SynOptSys can help designers and product developers overcome the barriers that prevent them from using powerful techniques to develop better pro ducts in a less costly manner. SynOpt-Sys is applicable to the design of any product or process with multiple criteria to meet, and at least two factors that influence achievement of those criteria. The user begins with a selected solution principle or system concept and a set of criteria that needs to be satisfied. The criteria may be expressed in terms of documented desirements or defined responses that the future system needs to achieve. Documented desirements can be imported into SynOptSys or created and documented directly within SynOptSys. Subsequent steps include identifying factors, specifying model order for each response, designing the experiment, running the experiment and gathering the data, analyzing the results, and determining the specifications for the optimized system. The user may also enter textual information as the project progresses. Data is easily edited within SynOptSys, and the software design enables full traceability within any step in the process, and facilitates reporting as needed. SynOptSys is unique in the way responses are defined and the nuances of the goodness associated with changes in response values for each of the responses of interest. The Desirability Optimization Methodology provides the basis of this novel feature. Moreover, this is a complete, guided design and optimization process tool with embedded math that can remain invisible to the user. It is not a standalone statistical program; it is a design and optimization system.

Human serum antibodies against EBV latent membrane protein 1 cross-react with α-synuclein

PubMed Central

Gray, Madison T.; Ganesh, Munisha S.; Middeldorp, Jaap M.

2016-01-01

Objectives: To identify the epitope on α-synuclein (α-syn) to which antibodies against the Epstein-Barr virus (EBV) latent membrane protein 1 (LMP1) bind and to determine whether antibodies targeting this mimicry domain are present in human sera. Methods: Reactivity of the α-syn-cross-reacting anti-LMP1 monoclonal antibody CS1-4 to a synthetic peptide containing the putative mimicry domain was compared to those in which this domain was mutated and to murine and rat α-syn (which differ from human α-syn at this site) in Western blots. Using ELISA, sera from EBV+ (n = 4) and EBV− (n = 12) donors as well as those with infectious mononucleosis (IM; n = 120), and Hodgkin disease (HD; n = 33) were interrogated for antibody reactivity to synthetic peptides corresponding to regions of α-syn and LMP1 containing the mimicry domain. Results: CS1-4 showed strong reactivity to wild-type human α-syn, but not to the mutant peptides or rodent α-syn. Control EBV− and EBV+ sera showed no reactivity to α-syn or LMP1 peptides. However, a significant proportion of IM and HD sera contained immunoglobulin M (IgM) (59% and 70%, in IM and HD, respectively), immunoglobulin G (IgG) (40% and 48%), and immunoglobulin A (IgA) (28% and 36%) antibodies to both peptides, as well as a significant correlation in the titers of IgM (ρ = 0.606 and 0.664, for IM and HD, respectively), IgG (0.526 and 0.836), and IgA (0.569 and 0.728) antibodies targeting LMP1 and α-syn peptides. Conclusions: Anti-EBV-LMP1 antibodies cross-reacting with a defined epitope in α-syn are present in human patients. These findings may have implications for the pathogenesis of synucleinopathies. PMID:27218119

Purine 3':5'-cyclic nucleotides with the nucleobase in a syn orientation: cAMP, cGMP and cIMP.

PubMed

Řlepokura, Katarzyna Anna

2016-06-01

Purine 3':5'-cyclic nucleotides are very well known for their role as the secondary messengers in hormone action and cellular signal transduction. Nonetheless, their solid-state conformational details still require investigation. Five crystals containing purine 3':5'-cyclic nucleotides have been obtained and structurally characterized, namely adenosine 3':5'-cyclic phosphate dihydrate, C10H12N5O6P·2H2O or cAMP·2H2O, (I), adenosine 3':5'-cyclic phosphate 0.3-hydrate, C10H12N5O6P·0.3H2O or cAMP·0.3H2O, (II), guanosine 3':5'-cyclic phosphate pentahydrate, C10H12N5O7P·5H2O or cGMP·5H2O, (III), sodium guanosine 3':5'-cyclic phosphate tetrahydrate, Na(+)·C10H11N5O7P(-)·4H2O or Na(cGMP)·4H2O, (IV), and sodium inosine 3':5'-cyclic phosphate tetrahydrate, Na(+)·C10H10N4O7P(-)·4H2O or Na(cIMP)·4H2O, (V). Most of the cyclic nucleotide zwitterions/anions [two from four cAMP present in total in (I) and (II), cGMP in (III), cGMP(-) in (IV) and cIMP(-) in (V)] are syn conformers about the N-glycosidic bond, and this nucleobase arrangement is accompanied by Crib-H...Npur hydrogen bonds (rib = ribose and pur = purine). The base orientation is tuned by the ribose pucker. An analysis of data obtained from the Cambridge Structural Database made in the context of syn-anti conformational preferences has revealed that among the syn conformers of various purine nucleotides, cyclic nucleotides and dinucleotides predominate significantly. The interactions stabilizing the syn conformation have been indicated. The inter-nucleotide contacts in (I)-(V) have been systematized in terms of the chemical groups involved. All five structures display three-dimensional hydrogen-bonded networks.

Kinetic Study of Denatonium Sorption to Smectite Clay Minerals.

PubMed

Crosson, Garry S; Sandmann, Emily

2013-06-01

The denatonium cation, as a benzoate salt, is the most bitter cation known to modern society and is frequently added to consumer products to reduce accidental and intentional consumption by humans and animals. Denatonium can enter the environment by accidental discharges, potentially rendering water supplies undrinkable. Interactions of denatonium with soil components ( i.e. , smectite minerals) ultimately control the environmental fate of denatonium, but the current literature is devoid of studies that evaluate denatonium sorption to smectite minerals. This study investigated the mechanism and kinetics of denatonium sorption to smectite clay minerals as a function of smectite type, temperature, pH and ionic strength. Uptake by synthetic mica montmorillonite (Syn-1), Wyoming montmorillonite (SWy-2), and Texas montmorillonite (STx-1b) at 305K was rapid, with equilibrium being reached within 2 min for all clays. Complete removal of denatonium was observed for STx-1b at pH 6.9, while partial removal was observed for Syn-1 and SWy-2. Kinetic behavior of SWy-2 and Syn-1 is consistent with a pseudo-second-order model at 305K. An activation energy of +25.9 kJ/mol was obtained for sorption to Syn-1 and was independent of temperature between 286K and 338K. Activation-free energy (Δ G *), activation enthalpy (Δ H *), and activation entropy (Δ S *) for Syn-1 were found to be +62.91 kJ/mol, +23.36 kJ/mol, and -0.130 kJ/(K·mol), respectively. Sorption capacities at pH 3.6, 6.9, and 8.2 were constant at 1.3×10 -2 g denatonium/g clay; however, the kinetic rate constant increased by 56%, going from acidic to basic solution conditions. Distribution coefficients were negatively correlated with ionic strength, suggesting cation exchange. Collectively, results suggested that smectite minerals can serve as efficient sinks for denatonium cations. This is much-needed information for agencies developing regulations regarding denatonium usage and for water treatment professionals who may ultimately have to treat denatonium-impacted water supplies.

Kinetic Study of Denatonium Sorption to Smectite Clay Minerals

PubMed Central

Crosson, Garry S.; Sandmann, Emily

2013-01-01

Abstract The denatonium cation, as a benzoate salt, is the most bitter cation known to modern society and is frequently added to consumer products to reduce accidental and intentional consumption by humans and animals. Denatonium can enter the environment by accidental discharges, potentially rendering water supplies undrinkable. Interactions of denatonium with soil components (i.e., smectite minerals) ultimately control the environmental fate of denatonium, but the current literature is devoid of studies that evaluate denatonium sorption to smectite minerals. This study investigated the mechanism and kinetics of denatonium sorption to smectite clay minerals as a function of smectite type, temperature, pH and ionic strength. Uptake by synthetic mica montmorillonite (Syn-1), Wyoming montmorillonite (SWy-2), and Texas montmorillonite (STx-1b) at 305K was rapid, with equilibrium being reached within 2 min for all clays. Complete removal of denatonium was observed for STx-1b at pH 6.9, while partial removal was observed for Syn-1 and SWy-2. Kinetic behavior of SWy-2 and Syn-1 is consistent with a pseudo–second-order model at 305K. An activation energy of +25.9 kJ/mol was obtained for sorption to Syn-1 and was independent of temperature between 286K and 338K. Activation-free energy (ΔG*), activation enthalpy (ΔH*), and activation entropy (ΔS*) for Syn-1 were found to be +62.91 kJ/mol, +23.36 kJ/mol, and −0.130 kJ/(K·mol), respectively. Sorption capacities at pH 3.6, 6.9, and 8.2 were constant at 1.3×10−2 g denatonium/g clay; however, the kinetic rate constant increased by 56%, going from acidic to basic solution conditions. Distribution coefficients were negatively correlated with ionic strength, suggesting cation exchange. Collectively, results suggested that smectite minerals can serve as efficient sinks for denatonium cations. This is much-needed information for agencies developing regulations regarding denatonium usage and for water treatment professionals who may ultimately have to treat denatonium-impacted water supplies. PMID:23781128

Cutoff size need not strongly influence molecular dynamics results for solvated polypeptides.

PubMed

Beck, David A C; Armen, Roger S; Daggett, Valerie

2005-01-18

The correct treatment of van der Waals and electrostatic nonbonded interactions in molecular force fields is essential for performing realistic molecular dynamics (MD) simulations of solvated polypeptides. The most computationally tractable treatment of nonbonded interactions in MD utilizes a spherical distance cutoff (typically, 8-12 A) to reduce the number of pairwise interactions. In this work, we assess three spherical atom-based cutoff approaches for use with all-atom explicit solvent MD: abrupt truncation, a CHARMM-style electrostatic shift truncation, and our own force-shifted truncation. The chosen system for this study is an end-capped 17-residue alanine-based alpha-helical peptide, selected because of its use in previous computational and experimental studies. We compare the time-averaged helical content calculated from these MD trajectories with experiment. We also examine the effect of varying the cutoff treatment and distance on energy conservation. We find that the abrupt truncation approach is pathological in its inability to conserve energy. The CHARMM-style shift truncation performs quite well but suffers from energetic instability. On the other hand, the force-shifted spherical cutoff method conserves energy, correctly predicts the experimental helical content, and shows convergence in simulation statistics as the cutoff is increased. This work demonstrates that by using proper and rigorous techniques, it is possible to correctly model polypeptide dynamics in solution with a spherical cutoff. The inherent computational advantage of spherical cutoffs over Ewald summation (and related) techniques is essential in accessing longer MD time scales.

α-Synuclein fibril-induced paradoxical structural and functional defects in hippocampal neurons.

PubMed

Froula, Jessica M; Henderson, Benjamin W; Gonzalez, Jose Carlos; Vaden, Jada H; Mclean, John W; Wu, Yumei; Banumurthy, Gokulakrishna; Overstreet-Wadiche, Linda; Herskowitz, Jeremy H; Volpicelli-Daley, Laura A

2018-05-01

Neuronal inclusions composed of α-synuclein (α-syn) characterize Parkinson's Disease (PD) and Dementia with Lewy bodies (DLB). Cognitive dysfunction defines DLB, and up to 80% of PD patients develop dementia. α-Syn inclusions are abundant in the hippocampus, yet functional consequences are unclear. To determine if pathologic α-syn causes neuronal defects, we induced endogenous α-syn to form inclusions resembling those found in diseased brains by treating hippocampal neurons with α-syn fibrils. At seven days after adding fibrils, α-syn inclusions are abundant in axons, but there is no cell death at this time point, allowing us to assess for potential alterations in neuronal function that are not caused by neuron death. We found that exposure of neurons to fibrils caused a significant reduction in mushroom spine densities, adding to the growing body of literature showing that altered spine morphology is a major pathologic phenotype in synucleinopathies. The reduction in spine densities occurred only in wild type neurons and not in neurons from α-syn knockout mice, suggesting that the changes in spine morphology result from fibril-induced corruption of endogenously expressed α-syn. Paradoxically, reduced postsynaptic spine density was accompanied by increased frequency of miniature excitatory postsynaptic currents (EPSCs) and presynaptic docked vesicles, suggesting enhanced presynaptic function. Action-potential dependent activity was unchanged, suggesting compensatory mechanisms responding to synaptic defects. Although activity at the level of the synapse was unchanged, neurons exposed to α-syn fibrils, showed reduced frequency and amplitudes of spontaneous Ca 2+ transients. These findings open areas of research to determine the mechanisms that alter neuronal function in brain regions critical for cognition at time points before neuron death.

Synaptic Regulator α-Synuclein in Dopaminergic Fibers Is Essentially Required for the Maintenance of Subependymal Neural Stem Cells.

PubMed

Perez-Villalba, Ana; Sirerol-Piquer, M Salomé; Belenguer, Germán; Soriano-Cantón, Raúl; Muñoz-Manchado, Ana Belén; Villadiego, Javier; Alarcón-Arís, Diana; Soria, Federico N; Dehay, Benjamin; Bezard, Erwan; Vila, Miquel; Bortolozzi, Analía; Toledo-Aral, Juan José; Pérez-Sánchez, Francisco; Fariñas, Isabel

2018-01-24

Synaptic protein α-synuclein (α-SYN) modulates neurotransmission in a complex and poorly understood manner and aggregates in the cytoplasm of degenerating neurons in Parkinson's disease. Here, we report that α-SYN present in dopaminergic nigral afferents is essential for the normal cycling and maintenance of neural stem cells (NSCs) in the brain subependymal zone of adult male and female mice. We also show that premature senescence of adult NSCs into non-neurogenic astrocytes in mice lacking α-SYN resembles the effects of dopaminergic fiber degeneration resulting from chronic exposure to 1-methyl-4-phenyl-1,2,3,6-tetra-hydropyridine or intranigral inoculation of aggregated toxic α-SYN. Interestingly, NSC loss in α-SYN-deficient mice can be prevented by viral delivery of human α-SYN into their sustantia nigra or by treatment with l-DOPA, suggesting that α-SYN regulates dopamine availability to NSCs. Our data indicate that α-SYN, present in dopaminergic nerve terminals supplying the subependymal zone, acts as a niche component to sustain the neurogenic potential of adult NSCs and identify α-SYN and DA as potential targets to ameliorate neurogenic defects in the aging and diseased brain. SIGNIFICANCE STATEMENT We report an essential role for the protein α-synuclein present in dopaminergic nigral afferents in the regulation of adult neural stem cell maintenance, identifying the first synaptic regulator with an implication in stem cell niche biology. Although the exact role of α-synuclein in neural transmission is not completely clear, our results indicate that it is required for stemness and the preservation of neurogenic potential in concert with dopamine. Copyright © 2018 the authors 0270-6474/18/380815-12$15.00/0.

The novel Parkinson's disease linked mutation G51D attenuates in vitro aggregation and membrane binding of α-synuclein, and enhances its secretion and nuclear localization in cells

PubMed Central

Fares, Mohamed-Bilal; Ait-Bouziad, Nadine; Dikiy, Igor; Mbefo, Martial K.; Jovičić, Ana; Kiely, Aoife; Holton, Janice L.; Lee, Seung-Jae; Gitler, Aaron D.; Eliezer, David; Lashuel, Hilal A.

2014-01-01

A novel mutation in the α-Synuclein (α-Syn) gene “G51D” was recently identified in two familial cases exhibiting features of Parkinson's disease (PD) and multiple system atrophy (MSA). In this study, we explored the impact of this novel mutation on the aggregation, cellular and biophysical properties of α-Syn, in an attempt to unravel how this mutant contributes to PD/MSA. Our results show that the G51D mutation significantly attenuates α-Syn aggregation in vitro. Moreover, it disrupts local helix formation in the presence of SDS, decreases binding to lipid vesicles C-terminal to the site of mutation and severely inhibits helical folding in the presence of acidic vesicles. When expressed in yeast, α-SynG51D behaves similarly to α-SynA30P, as both exhibit impaired membrane association, form few inclusions and are non-toxic. In contrast, enhanced secreted and nuclear levels of the G51D mutant were observed in mammalian cells, as well as in primary neurons, where α-SynG51D was enriched in the nuclear compartment, was hyper-phosphorylated at S129 and exacerbated α-Syn-induced mitochondrial fragmentation. Finally, post-mortem human brain tissues of α-SynG51D cases were examined, and revealed only partial colocalization with nuclear membrane markers, probably due to post-mortem tissue delay and fixation. These findings suggest that the PD-linked mutations may cause neurodegeneration via different mechanisms, some of which may be independent of α-Syn aggregation. PMID:24728187

Validation of α-Synuclein as a CSF Biomarker for Sporadic Creutzfeldt-Jakob Disease.

PubMed

Llorens, Franc; Kruse, Niels; Karch, André; Schmitz, Matthias; Zafar, Saima; Gotzmann, Nadine; Sun, Ting; Köchy, Silja; Knipper, Tobias; Cramm, Maria; Golanska, Ewa; Sikorska, Beata; Liberski, Pawel P; Sánchez-Valle, Raquel; Fischer, Andre; Mollenhauer, Brit; Zerr, Inga

2018-03-01

The analysis of cerebrospinal fluid (CSF) biomarkers gains importance in the differential diagnosis of prion diseases. However, no single diagnostic tool or combination of them can unequivocally confirm prion disease diagnosis. Electrochemiluminescence (ECL)-based immunoassays have demonstrated to achieve high diagnostic accuracy in a variety of sample types due to their high sensitivity and dynamic range. Quantification of CSF α-synuclein (a-syn) by an in-house ECL-based ELISA assay has been recently reported as an excellent approach for the diagnosis of sporadic Creutzfeldt-Jakob disease (sCJD), the most prevalent form of human prion disease. In the present study, we validated a commercially available ECL-based a-syn ELISA platform as a diagnostic test for correct classification of sCJD cases. CSF a-syn was analysed in 203 sCJD cases with definite diagnosis and in 445 non-CJD cases. We investigated reproducibility and stability of CSF a-syn and made recommendations for its analysis in the sCJD diagnostic workup. A sensitivity of 98% and a specificity of 97% were achieved when using an optimal cut-off of 820 pg/mL a-syn. Moreover, we were able to show a negative correlation between a-syn levels and disease duration suggesting that CSF a-syn may be a good prognostic marker for sCJD patients. The present study validates the use of a-syn as a CSF biomarker of sCJD and establishes the clinical and pre-analytical parameters for its use in differential diagnosis in clinical routine. Additionally, the current test presents some advantages compared to other diagnostic approaches: it is fast, economic, requires minimal amount of CSF and a-syn levels are stable along disease progression.

Chemical Compensation of Mitochondrial Phospholipid Depletion in Yeast and Animal Models of Parkinson’s Disease

PubMed Central

Wang, Shaoxiao; Zhang, Siyuan; Xu, Chuan; Barron, Addie; Galiano, Floyd; Patel, Dhaval; Lee, Yong Joo; Caldwell, Guy A.; Caldwell, Kim A.

2016-01-01

We have been investigating the role that phosphatidylethanolamine (PE) and phosphatidylcholine (PC) content plays in modulating the solubility of the Parkinson’s disease protein alpha-synuclein (α-syn) using Saccharomyces cerevisiae and Caenorhabditis elegans. One enzyme that synthesizes PE is the conserved enzyme phosphatidylserine decarboxylase (Psd1/yeast; PSD-1/worms), which is lodged in the inner mitochondrial membrane. We previously found that decreasing the level of PE due to knockdown of Psd1/psd-1 affects the homeostasis of α-syn in vivo. In S. cerevisiae, the co-occurrence of low PE and α-syn in psd1Δ cells triggers mitochondrial defects, stress in the endoplasmic reticulum, misprocessing of glycosylphosphatidylinositol-anchored proteins, and a 3-fold increase in the level of α-syn. The goal of this study was to identify drugs that rescue this phenotype. We screened the Prestwick library of 1121 Food and Drug Administration-approved drugs using psd1Δ + α-syn cells and identified cyclosporin A, meclofenoxate hydrochloride, and sulfaphenazole as putative protective compounds. The protective activity of these drugs was corroborated using C. elegans in which α-syn is expressed specifically in the dopaminergic neurons, with psd-1 depleted by RNAi. Worm populations were examined for dopaminergic neuron survival following psd-1 knockdown. Exposure to cyclosporine, meclofenoxate, and sulfaphenazole significantly enhanced survival at day 7 in α-syn-expressing worm populations whereby 50–55% of the populations displayed normal neurons, compared to only 10–15% of untreated animals. We also found that all three drugs rescued worms expressing α-syn in dopaminergic neurons that were deficient in the phospholipid cardiolipin following cardiolipin synthase (crls-1) depletion by RNAi. We discuss how these drugs might block α-syn pathology in dopaminergic neurons. PMID:27736935

Specific pesticide-dependent increases in α-synuclein levels in human neuroblastoma (SH-SY5Y) and melanoma (SK-MEL-2) cell lines.

PubMed

Chorfa, Areski; Bétemps, Dominique; Morignat, Eric; Lazizzera, Corinne; Hogeveen, Kevin; Andrieu, Thibault; Baron, Thierry

2013-06-01

Epidemiological studies indicate a role of genetic and environmental factors in Parkinson's disease involving alterations of the neuronal α-synuclein (α-syn) protein. In particular, a relationship between Parkinson's disease and occupational exposure to pesticides has been repeatedly suggested. Our objective was to precisely assess changes in α-syn levels in human neuroblastoma (SH-SY5Y) and melanoma (SK-MEL-2) cell lines following acute exposure to pesticides (rotenone, paraquat, maneb, and glyphosate) using Western blot and flow cytometry. These human cell lines express α-syn endogenously, and overexpression of α-syn (wild type or mutated A53T) can be obtained following recombinant adenoviral transduction. We found that endogenous α-syn levels in the SH-SY5Y neuroblastoma cell line were markedly increased by paraquat, and to a lesser extent by rotenone and maneb, but not by glyphosate. Rotenone also clearly increased endogenous α-syn levels in the SK-MEL-2 melanoma cell line. In the SH-SY5Y cell line, similar differences were observed in the α-syn adenovirus-transduced cells, with a higher increase of the A53T mutated protein. Paraquat markedly increased α-syn in the SK-MEL-2 adenovirus-transduced cell line, similarly for the wild-type or A53T proteins. The observed differences in the propensities of pesticides to increase α-syn levels are in agreement with numerous reports that indicate a potential role of exposure to certain pesticides in the development of Parkinson's disease. Our data support the hypothesis that pesticides can trigger some molecular events involved in this disease and also in malignant melanoma that consistently shows a significant but still unexplained association with Parkinson's disease.

The universality of β-hairpin misfolding indicated by molecular dynamics simulations.

PubMed

Shao, Qiang; Wang, Jinan; Shi, Jiye; Zhu, Weiliang

2013-10-28

Previous molecular dynamics simulations showed that besides the experimentally measured folded structures, several β-structured polypeptides could also have misfolded "out-of-register" structures. Through the enhanced sampling molecular dynamics simulations on a series of polypeptides using either implicit or explicit solvent model, the present study systematically investigated the universality of β-hairpin misfolding and its determinants. It was observed that the misfolding could take place for almost all polypeptides under study, especially in the presence of weak side chain hydrophobicity. Moreover, the observed misfolded structures for various polypeptides share the following common features: (1) the turn length in misfolded structure is one-residue shorter than that in folded structure; (2) the hydrophobic side chains on the two strands are pointed to the opposite directions instead of packing in the same direction to form hydrophobic core cluster in the folded structure; and (3) the misfolded structure is one-residue-shifted asymmetric β-hairpin structure. The detailed analysis suggested that the misfolding of β-hairpin is the result of the competition between the formation of the alterable turn configurations and the inter-strand hydrophobic interactions. These predictions are desired to be tested by experiments.

The universality of β-hairpin misfolding indicated by molecular dynamics simulations

NASA Astrophysics Data System (ADS)

Shao, Qiang; Wang, Jinan; Shi, Jiye; Zhu, Weiliang

2013-10-01

Previous molecular dynamics simulations showed that besides the experimentally measured folded structures, several β-structured polypeptides could also have misfolded "out-of-register" structures. Through the enhanced sampling molecular dynamics simulations on a series of polypeptides using either implicit or explicit solvent model, the present study systematically investigated the universality of β-hairpin misfolding and its determinants. It was observed that the misfolding could take place for almost all polypeptides under study, especially in the presence of weak side chain hydrophobicity. Moreover, the observed misfolded structures for various polypeptides share the following common features: (1) the turn length in misfolded structure is one-residue shorter than that in folded structure; (2) the hydrophobic side chains on the two strands are pointed to the opposite directions instead of packing in the same direction to form hydrophobic core cluster in the folded structure; and (3) the misfolded structure is one-residue-shifted asymmetric β-hairpin structure. The detailed analysis suggested that the misfolding of β-hairpin is the result of the competition between the formation of the alterable turn configurations and the inter-strand hydrophobic interactions. These predictions are desired to be tested by experiments.

A systematic catalogue of butterflies of the former Soviet Union (Armenia, Azerbaijan, Belarus, Estonia, Georgia, Kyrgyzstan, Kazakhstan, Latvia, Lituania, Moldova, Russia, Tajikistan, Turkmenistan, Ukraine, Uzbekistan) with special account to their type specimens (Lepidoptera: Hesperioidea, Papilionoidea).

PubMed

Korb, Stanislav K; Bolshakov, Lavr V

2016-09-01

A catalogue of butterflies of Russia and adjacent countries is given, with special account to the name-bearing types depository. This catalogue contains data about 86 species (3 of them are questionable) of Hesperiidae (22 genera); 47 species of Papilionidae (14 genera); 89 species of Pieridae (5 of them are questionable)  (15 genera); 1 species (1 genus) of Libytheinae(dae); 2 species of Danainae(dae) (2 genera); 160 species of Nymphalinae(dae) (1 of them is questionable) (23 genera); 259 species of Satyrinae(dae) (14 of them are questionable, mainly from genera Oeneis and Pseudochazara) (34 genera); 3 species of Riodinidae (2 genera); 318 species of Lycaenidae (11 of them are questionable, mainly from genera Neolycaena and Plebeius) (57 genera). In total: 965 species of butterflies, 174 genera, by countries: Armenia-244, Azerbaijan-225, Belarus-107, Estonia-113, Georgia-211, Kyrgyzstan-316, Kazakhstan-344, Latvia-115, Lituania-126, Moldova-87, Russia-522, Tajikistan-295, Turkmenistan-159, Ukraine-192, Uzbekistan-241. Detailed distribution and subspecific structure (if present) for every species is provided. Lectotypes of the following species-group taxa are designated: Hesperia poggei Lederer, 1858, Parnassius felderi Bremer, 1861, P. eversmanni Eversmann, 1851, P. boedromius Püngeler, 1901, Limenitis moltrechti Kardakov, 1928, L. sydyi Kindermann, 1853, L. amphyssa Ménétriès, 1859, L. doerriesi Staudinger, 1892, L. helmanni duplicata Staudinger, 1892, L. homeyeri Tancré, 1881, Argynnis penelope Staudinger, 1891, A. thore borealis Staudinger, 1861, Vanessa io geisha Stichel, [1908], Melitaea maturna staudingeri Wnukowsky, 1929 (=uralensis Staudinger, 1871), M. didymina Staudinger, 1895, Papilio fascelis Esper, 1783, Thecla quercivora Staudinger, 1887, Lycaena orion var. ornata Staudinger, 1892. The following nomenclatural acts are established: Neolycaena submontana baitenovi (Zhdanko, 2011), comb. et stat.n. The following new synonymy is provided: Hesperia comma repugnans (Staudinger, 1892) = lena Korshunov et Gorbunov, 1995, syn.n.; Argynnis niobe orientalis Alphéraky, 1881 =ornata Staudinger, 1901, syn.n. =tanjusha Zhdanko, 2011, syn.n.; Boloria frigga gibsoni (Barnes & Benjamin, 1926) = kosarevi Korb, 2011, syn.n., B. erubescens houri Wyatt, 1961 =ancilla Churkin, 2004, syn.n.; Melitaea fergana maracandica Staudinger, 1882 = irinae Churkin, Kolesnichenko et Tremasov, 2012, syn.n.; M. asteroida clara Staudinger, 1887 =ludmilla Churkin, Kolesnichenko et Tuzov, 2000, syn.n.; Paralasa jordana jordana (Staudinger, 1882) =khramovi Churkin et Pletnev, 2012, syn.n.; P. jordana subocellata (Staudinger, 1901) =kipnisi Churkin et Pletnev, 2012, syn.n.; P. kusnezovi kusnezovi (Avinov, 1910) =bosbutaensis Churkin et Pletnev, 2012, syn.n.; Erebia meta Staudinger, 1886 =gertha Staudinger, 1886, syn.n.; Oeneis ammon ammon Elwes, 1899 =smirnovi Yakovlev, 2011, syn.n.; O. norna tundra A.Bang-Haas, 1912 =ivonini Yakovlev, 2011, syn.n.; Chazara briseis ianthe (Pallas, 1771) =lyrnessus Fruhstorfer, 1908, syn.n., Plebejides stekolnikovi (Stradomsky et Tikhonov, 2015), comb.n.

Taxonomic revision of Afrotropical Laccophilus Leach, 1815 (Coleoptera, Dytiscidae)

PubMed Central

Biström, Olof; Nilsson, Anders N.; Bergsten, Johannes

2015-01-01

Abstract The African species of the genus Laccophilus Leach, 1815, are revised, on the basis of study of adult specimens. In all, 105 species are now recognized. A phenetic character-analysis was undertaken, which resulted in a split of the genus into 17 species groups. Diagnoses and a description of each species are given together with keys for identification of species groups and species. We also provide habitus photos, illustration of male genitalia and distribution maps for all species. New species are described as follows: Laccophilus grossus sp. n. (Angola, Namibia), Laccophilus rocchii sp. n. (Tanzania, Namibia, Botswana, Mozambique), Laccophilus ferrugo sp. n. (Mozambique), Laccophilus furthi sp. n. (Madagascar), Laccophilus isamberti sp. n. (Madagascar), Laccophilus inobservatus sp. n. (Gambia, Senegal, Mali, Niger, Sudan, Chad, Ethiopia, Burkina Faso, Ivory Coast, Ghana, Nigeria, Cameroon, Zaire and Asia: Yemen), Laccophilus cryptos sp. n. (Zaire, Mozambique, Zimbabwe, Namibia, Botswana, South Africa), Laccophilus enigmaticus sp. n. (Nigeria, Sudan), Laccophilus bellus sp. n. (Benin, Nigeria), Laccophilus guentheri sp. n. (Guinea, Ghana), Laccophilus guineensis sp. n. (Guinea), Laccophilus decorosus sp. n. (Uganda), Laccophilus empheres sp. n. (Kenya), Laccophilus inconstans sp. n. (Guinea, Ivory Coast, Ghana, Nigeria, Cameroon), Laccophilus brancuccii sp. n. (Central African Republic), Laccophilus incomptus sp. n. (Cameroon), Laccophilus australis sp. n. (Tanzania, South Africa), Laccophilus minimus sp. n. (Namibia), Laccophilus eboris sp. n. (Ivory Coast), Laccophilus insularum sp. n. (Madagascar), Laccophilus occidentalis sp. n. (Gambia, Senegal, Mali, Guinea, Sierra Leone, Ivory Coast, Ghana, Benin, Nigeria, Central African Republic, Zaire) and Laccophilus transversovittatus sp. n. (Madagascar). Laccophilus restrictus Sharp, 1882, is restored as good species; not junior synonym of Laccophilus pictipennis Sharp, 1882. New synonyms are established as follows: Laccophilus continentalis Gschwendtner, 1935 = Laccophilus perplexus Omer-Cooper, 1970, syn. n., Laccophilus taeniolatus Régimbart, 1889 = Laccophilus congener Omer-Cooper, 1957, syn. n., Laccophilus adspersus Boheman, 1848 = Laccophilus vitshumbii Guignot, 1959, syn. n. = Laccophilus adspersus nigeriensis Omer-Cooper, 1970, syn. n. = Laccophilus adspersus sudanensis Omer-Cooper, 1970, syn. n., Laccophilus modestus Régimbart, 1895 = Laccophilus espanyoli Hernando, 1990, syn. n., Laccophilus flaveolus Régimbart, 1906 = Laccophilus pampinatus Guignot, 1941, syn. n., Laccophilus trilineola Régimbart, 1889 = Laccophilus simulator Omer-Cooper, 1958, syn. n., Laccophilus mediocris Guignot, 1952 = Laccophilus meii Rocchi, 2000, syn. n., Laccophilus epinephes Guignot, 1955 = Laccophilus castaneus Guignot, 1956, syn. n., Laccophilus saegeri Guignot, 1958 = Laccophilus comoensis Pederzani & Reintjes, 2002, syn. n., Laccophilus restrictus Sharp, 1882 = Laccophilus evanescens Régimbart, 1895, syn. n., Laccophilus incrassatus Gschwendtner, 1933 = Laccophilus virgatus Guignot, 1953, syn. n., Laccophilus cyclopis Sharp, 1882 = Laccophilus shephardi Omer-Cooper, 1965, syn. n., Laccophilus burgeoni Gschwendtner, 1930 = Laccophilus wittei Guignot, 1952, syn. n., Laccophilus secundus Régimbart, 1895 = Laccophilus torquatus Guignot, 1956, syn. n., Laccophilus desintegratus Régimbart, 1895 = Laccophilus sanguinosus Régimbart, 1895, syn. n. and Laccophilus flavopictus Régimbart, 1889 = Laccophilus bergeri Guignot, 1953, syn. n. = Laccophilus segmentatus Omer-Cooper, 1957, syn. n. Lectotypes are designated for the following taxa: Laccophilus productus Régimbart, 1906, Laccophilus ruficollis Zimmermann, 1919, Laccophilus sordidus Sharp, 1882, Laccophilus alluaudi Régimbart, 1899, Laccophilus pictipennis Sharp, 1882, Laccophilus wehnckei Sharp, 1882, Laccophilus continentalis Gschwendtner, 1935, Laccophilus simplicistriatus Gschwendtner, 1932, Laccophilus complicatus Sharp, 1882, Laccophilus rivulosus Klug, 1833, Laccophilus ampliatus Régimbart, 1895, Laccophilus pilitarsis Régimbart, 1906, Laccophilus adspersus Boheman, 1848, Laccophilus livens Régimbart, 1895, Laccophilus modestus Régimbart, 1895, Laccophilus nodieri Régimbart, 1895, Laccophilus flaveolus Régimbart, 1906, Laccophilus pallescens Régimbart, 1903, Laccophilus restrictus Sharp, 1882, Laccophilus vermiculosus Gerstaecker, 1867, Laccophilus mocquerysi Régimbart, 1895, Laccophilus bizonatus Régimbart, 1895, Laccophilus tschoffeni Régimbart, 1895, Laccophilus persimilis Régimbart, 1895, Laccophilus poecilus Klug, 1834, Laccophilus lateralis Sharp, 1882, Laccophilus lateralis var. polygrammus Régimbart, 1903, Laccophilus cyclopis Sharp, 1882, Laccophilus shephardi Omer-Cooper, 1965, Laccophilus conjunctus Guignot, 1950, Laccophilus grammicus Sharp, 1882, Laccophilus flavoscriptus Régimbart, 1895, Laccophilus flavosignatus Régimbart, 1895, Laccophilus brevicollis Sharp, 1882, Laccophilus secundus Régimbart, Laccophilus desintegratus Régimbart, 1895, Laccophilus gutticollis Régimbart, 1895, Laccophilus luctuosus Sharp, 1882 and Laccophilus inornatus Zimmermann, 1926. Laccophilus remex Guignot, 1952, comprises a species complex with uncertain taxonomic delimitation; the complex includes Laccophilus concisus Guignot, 1953, Laccophilus turneri Omer-Cooper, 1957 and Laccophilus praeteritus Omer-Cooper, 1957, as tentative synonyms of Laccophilus remex Guignot, 1952. PMID:26798285

Notice of release of Syn1 Tall Fescue

USDA-ARS?s Scientific Manuscript database

The Agricultural Research Service, U.S. Department of Agriculture announces the release of Syn1 tall fescue [Festuca arundinacea (syn., Lolium arundinaceum Darbyshire; Schedonorus phoenix (Scop.) Holub)] (PI xxxx, PI xxxx) germplasm developed by Dr. Bryan K. Kindiger at the USDA-ARS Grazinglands Res...

Nanomechanical properties of distinct fibrillar polymorphs of the protein α-synuclein.

PubMed

Makky, Ali; Bousset, Luc; Polesel-Maris, Jérôme; Melki, Ronald

2016-11-30

Alpha-synuclein (α-Syn) is a small presynaptic protein of 140 amino acids. Its pathologic intracellular aggregation within the central nervous system yields protein fibrillar inclusions named Lewy bodies that are the hallmarks of Parkinson's disease (PD). In solution, pure α-Syn adopts an intrinsically disordered structure and assembles into fibrils that exhibit considerable morphological heterogeneity depending on their assembly conditions. We recently established tightly controlled experimental conditions allowing the assembly of α-Syn into highly homogeneous and pure polymorphs. The latter exhibited differences in their shape, their structure but also in their functional properties. We have conducted an AFM study at high resolution and performed a statistical analysis of fibrillar α-Syn shape and thermal fluctuations to calculate the persistence length to further assess the nanomechanical properties of α-Syn polymorphs. Herein, we demonstrated quantitatively that distinct polymorphs made of the same protein (wild-type α-Syn) show significant differences in their morphology (height, width and periodicity) and physical properties (persistence length, bending rigidity and axial Young's modulus).

Assigning the stereochemistry of syn and anti β-trimethylsiloxy-α-trimethylsilyl alkanoic acid silyl esters using GIAO 1H NMR chemical shift calculations

NASA Astrophysics Data System (ADS)

Hadj Mohamed, Slim; Trabelsi, Mahmoud; Champagne, Benoît

2017-08-01

The stereostructure of β-trimethylsiloxy-α-trimethylsilyl alkanoic acid silyl esters synthesized by Bellassoued et al. [J. Org. Chem. 2001, 66, 5054-5057] using Mukaiyama aldol reaction has been reassigned using density functional theory NMR 1H chemical shifts calculations. It is now concluded that the major diastereoisomer is syn and the minor is anti. Within this assignment, for all silyl esters, δHa(anti) > δHa(syn), δHb(anti) < δHb(syn), and 3JHa-Hb (anti) > 3JHa-Hb (syn). Since the experimental assignment was based on the stereostructure (E/Z) of the cinnamic acid obtained by elimination of trimethylsilyl 3-phenyl-3-(trimethylsiloxy)-2-(trimethylsilyl)propanoate in the presence of TiCl4 and on the assumption that this elimination is anti stereospecific in acidic medium, one arrives at the conclusion that the elimination of syn and anti β-trimethylsiloxy-α-trimethylsilyl alkanoic acid silyl esters is not anti stereospecific.

Alpha-synuclein functions in the nucleus to protect against hydroxyurea-induced replication stress in yeast

PubMed Central

Liu, Xianpeng; Lee, Yong Joo; Liou, Liang-Chun; Ren, Qun; Zhang, Zhaojie; Wang, Shaoxiao; Witt, Stephan N.

2011-01-01

Hydroxyurea (HU) inhibits ribonucleotide reductase (RNR), which catalyzes the rate-limiting synthesis of deoxyribonucleotides for DNA replication. HU is used to treat HIV, sickle-cell anemia and some cancers. We found that, compared with vector control cells, low levels of alpha-synuclein (α-syn) protect S. cerevisiae cells from the growth inhibition and reactive oxygen species (ROS) accumulation induced by HU. Analysis of this effect using different α-syn mutants revealed that the α-syn protein functions in the nucleus and not the cytoplasm to modulate S-phase checkpoint responses: α-syn up-regulates histone acetylation and RNR levels, maintains helicase minichromosome maintenance protein complexes (Mcm2–7) on chromatin and inhibits HU-induced ROS accumulation. Strikingly, when residues 2–10 or 96–140 are deleted, this protective function of α-syn in the nucleus is abolished. Understanding the mechanism by which α-syn protects against HU could expand our knowledge of the normal function of this neuronal protein. PMID:21642386

Nanomechanical properties of distinct fibrillar polymorphs of the protein α-synuclein

NASA Astrophysics Data System (ADS)

Makky, Ali; Bousset, Luc; Polesel-Maris, Jérôme; Melki, Ronald

2016-11-01

Alpha-synuclein (α-Syn) is a small presynaptic protein of 140 amino acids. Its pathologic intracellular aggregation within the central nervous system yields protein fibrillar inclusions named Lewy bodies that are the hallmarks of Parkinson’s disease (PD). In solution, pure α-Syn adopts an intrinsically disordered structure and assembles into fibrils that exhibit considerable morphological heterogeneity depending on their assembly conditions. We recently established tightly controlled experimental conditions allowing the assembly of α-Syn into highly homogeneous and pure polymorphs. The latter exhibited differences in their shape, their structure but also in their functional properties. We have conducted an AFM study at high resolution and performed a statistical analysis of fibrillar α-Syn shape and thermal fluctuations to calculate the persistence length to further assess the nanomechanical properties of α-Syn polymorphs. Herein, we demonstrated quantitatively that distinct polymorphs made of the same protein (wild-type α-Syn) show significant differences in their morphology (height, width and periodicity) and physical properties (persistence length, bending rigidity and axial Young’s modulus).

Development of SYN-004, an oral beta-lactamase treatment to protect the gut microbiome from antibiotic-mediated damage and prevent Clostridium difficile infection.

PubMed

Kaleko, Michael; Bristol, J Andrew; Hubert, Steven; Parsley, Todd; Widmer, Giovanni; Tzipori, Saul; Subramanian, Poorani; Hasan, Nur; Koski, Perrti; Kokai-Kun, John; Sliman, Joseph; Jones, Annie; Connelly, Sheila

2016-10-01

The gut microbiome, composed of the microflora that inhabit the gastrointestinal tract and their genomes, make up a complex ecosystem that can be disrupted by antibiotic use. The ensuing dysbiosis is conducive to the emergence of opportunistic pathogens such as Clostridium difficile. A novel approach to protect the microbiome from antibiotic-mediated dysbiosis is the use of beta-lactamase enzymes to degrade residual antibiotics in the gastrointestinal tract before the microflora are harmed. Here we present the preclinical development and early clinical studies of the beta-lactamase enzymes, P3A, currently referred to as SYN-004, and its precursor, P1A. Both P1A and SYN-004 were designed as orally-delivered, non-systemically available therapeutics for use with intravenous beta-lactam antibiotics. SYN-004 was engineered from P1A, a beta-lactamase isolated from Bacillus licheniformis, to broaden its antibiotic degradation profile. SYN-004 efficiently hydrolyses penicillins and cephalosporins, the most widely used IV beta-lactam antibiotics. In animal studies, SYN-004 degraded ceftriaxone in the GI tract of dogs and protected the microbiome of pigs from ceftriaxone-induced changes. Phase I clinical studies demonstrated SYN-004 safety and tolerability. Phase 2 studies are in progress to assess the utility of SYN-004 for the prevention of antibiotic-associated diarrhea and Clostridium difficile disease. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

In Situ Proximity Ligation Assay Reveals Co-Localization of Alpha-Synuclein and SNARE Proteins in Murine Primary Neurons.

PubMed

Almandoz-Gil, Leire; Persson, Emma; Lindström, Veronica; Ingelsson, Martin; Erlandsson, Anna; Bergström, Joakim

2018-01-01

The aggregation of alpha-synuclein (αSyn) is the pathological hallmark of Parkinson's disease, dementia with Lewy bodies and related neurological disorders. However, the physiological function of the protein and how this function relates to its pathological effects remain poorly understood. One of the proposed roles of αSyn is to promote the soluble N -ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complex assembly by binding to VAMP-2. The objective of this study was to visualize the co-localization between αSyn and the SNARE proteins (VAMP-2, SNAP-25, and syntaxin-1) for the first time using in situ proximity ligation assay (PLA). Cortical primary neurons were cultured from either non-transgenic or transgenic mice expressing human αSyn with the A30P mutation under the Thy-1 promoter. With an antibody recognizing both mouse and human αSyn, a PLA signal indicating close proximity between αSyn and the three SNARE proteins was observed both in the soma and throughout the processes. No differences in the extent of PLA signals were seen between non-transgenic and transgenic neurons. With an antibody specific against human αSyn, the PLA signal was mostly located to the soma and was only present in a few cells. Taken together, in situ PLA is a method that can be used to investigate the co-localization of αSyn and the SNARE proteins in primary neuronal cultures.

p27Kip1 regulates alpha-synuclein expression

PubMed Central

Gallastegui, Edurne; Domuro, Carla; Serratosa, Joan; Larrieux, Alejandra; Sin, Laura; Martinez, Jonatan; Besson, Arnaud; Morante-Redolat, José Manuel; Orlando, Serena; Aligue, Rosa; Fariñas, Isabel; Pujol, María Jesús; Bachs, Oriol

2018-01-01

Alpha-synuclein (α-SYN) is the main component of anomalous protein aggregates (Lewy bodies) that play a crucial role in several neurodegenerative diseases (synucleinopathies) like Parkinson’s disease and multiple system atrophy. However, the mechanisms involved in its transcriptional regulation are poorly understood. We investigated here the role of the cyclin-dependent kinase (Cdk) inhibitor and transcriptional regulator p27Kip1 (p27) in the regulation of α-SYN expression. We observed that selective deletion of p27 by CRISPR/Cas9 technology in neural cells resulted in increased levels of α-SYN. Knock-down of the member of the same family p21Cip1 (p21) also led to increased α-SYN levels, indicating that p27 and p21 collaborate in the repression of α-SYN transcription. We demonstrated that this repression is mediated by the transcription factor E2F4 and the member of the retinoblastoma protein family p130 and that it is dependent of Cdk activity. Chromatin immunoprecipitation analysis revealed specific binding sites for p27, p21 and E2F4 in the proximal α-SYN gene promoter. Finally, luciferase assays revealed a direct action of p27, p21 and E2F4 in α-SYN gene expression. Our findings reveal for the first time a negative regulatory mechanism of α-SYN expression, suggesting a putative role for cell cycle regulators in the etiology of synucleinopathies. PMID:29662651

Spine Topographical Distribution of Skin α-Synuclein Deposits in Idiopathic Parkinson Disease.

PubMed

Donadio, Vincenzo; Incensi, Alex; Rizzo, Giovanni; Scaglione, Cesa; Capellari, Sabina; Fileccia, Enrico; Avoni, Patrizia; Liguori, Rocco

2017-05-01

Phosphorylated α-synuclein (p-syn) in skin nerves mainly in the proximal sites is a promising neurodegenerative biomarker for idiopathic Parkinson disease (IPD). However, the p-syn spine distribution particularly in patients with unilateral motor dysfunctions remains undefined. This study aimed to investigate in IPD p-syn differences between left and right cervical spine sites in patients with prevalent unilateral motor symptoms, and cervical and thoracic spine sites in patients with bilateral motor symptoms. We enrolled 28 IPD patients fulfilling clinical diagnostic criteria associated with abnormal nigro-striatal DatScan and cardiac MIBG: 15 with prevalently unilateral motor symptoms demonstrated by DatScan; 13 with bilateral motor symptoms and DatScan abnormalities. Patients underwent skin biopsy searching for intraneural p-syn deposits: skin samples were taken from C7 paravertebral left and right sites in unilateral patients and from cervical (C7) and thoracic (Th12) paravertebral spine regions in bilateral patients. Unilateral patients displayed 20% of abnormal p-syn deposits in the affected motor site, 60% in both sites and 20% only in the non-affected site. P-syn was found in all patients in C7 but in only 62% of patients in Th12. Our data showed that cervical p-syn deposits displayed a uniform distribution between both sides not following the motor dysfunction in unilateral patients, and skin nerve p-syn deposits demonstrated a spine gradient with the cervical site expressing the highest positivity. © 2017 American Association of Neuropathologists, Inc. All rights reserved.

In vivo disruption of T cell development by expression of a dominant-negative polypeptide designed to abolish the SLP-76/Gads interaction.

PubMed

Jordan, Martha S; Maltzman, Jonathan S; Kliche, Stefanie; Shabason, Jacob; Smith, Jennifer E; Obstfeld, Amrom; Schraven, Burkhart; Koretzky, Gary A

2007-10-01

Multi-molecular complexes nucleated by adaptor proteins play a central role in signal transduction. In T cells, one central axis consists of the assembly of several signaling proteins linked together by the adaptors linker of activated T cells (LAT), Src homology 2 domain-containing leukocyte-specific phosphoprotein of 76 kDa (SLP-76), and Grb2-related adaptor downstream of Shc (Gads). Each of these adaptors has been shown to be important for normal T cell development, and their proper sub-cellular localization is critical for optimal function in cell lines. We previously demonstrated in Jurkat T cells and a rat basophilic leukemic cell line that expression of a 50-amino acid polypeptide identical to the site on SLP-76 that binds to Gads blocks proper localization of SLP-76 and SLP-76-dependent signaling events. Here we extend these studies to investigate the ability of this polypeptide to inhibit TCR-induced integrin activity in Jurkat cells and to inhibit in vivo thymocyte development and primary T cell function. These data provide evidence for the in vivo function of a dominant-negative peptide based upon the biology of SLP-76 action and suggest the possibility of therapeutic potential of targeting the SLP-76/Gads interaction.

MaxSynBio - Avenues towards creating cells from the bottom up.

PubMed

Schwille, Petra; Spatz, Joachim; Landfester, Katharina; Bodenschatz, Eberhard; Herminghaus, Stephan; Sourjik, Victor; Erb, Tobias; Bastiaens, Philippe; Lipowsky, Reinhard; Hyman, Anthony; Dabrock, Peter; Baret, Jean-Christophe; Vidakovic-Koch, Tanja; Bieling, Peter; Dimova, Rumiana; Mutschler, Hannes; Robinson, Tom; Tang, Dora; Wegner, Seraphine; Sundmacher, Kai

2018-05-11

A large Max Planck-based German research consortium ('MaxSynBio') was formed to investigate living systems from a fundamental perspective. The research program of MaxSynBio relies solely on the bottom-up approach to Synthetic Biology. MaxSynBio focuses on the detailed analysis and understanding of essential processes of life, via their modular reconstitution in minimal synthetic systems. The ultimate goal is to construct a basic living unit entirely from non-living components. The fundamental insights gained from the activities in MaxSynBio can eventually be utilized for establishing a new generation of biotechnological processes, which would be based on synthetic cell constructs that replace natural cells currently used in conventional biotechnology. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

The cellular transcription factor CREB corresponds to activating transcription factor 47 (ATF-47) and forms complexes with a group of polypeptides related to ATF-43.

PubMed

Hurst, H C; Masson, N; Jones, N C; Lee, K A

1990-12-01

Promoter elements containing the sequence motif CGTCA are important for a variety of inducible responses at the transcriptional level. Multiple cellular factors specifically bind to these elements and are encoded by a multigene family. Among these factors, polypeptides termed activating transcription factor 43 (ATF-43) and ATF-47 have been purified from HeLa cells and a factor referred to as cyclic AMP response element-binding protein (CREB) has been isolated from PC12 cells and rat brain. We demonstrated that CREB and ATF-47 are identical and that CREB and ATF-43 form protein-protein complexes. We also found that the cis requirements for stable DNA binding by ATF-43 and CREB are different. Using antibodies to ATF-43 we have identified a group of polypeptides (ATF-43) in the size range from 40 to 43 kDa. ATF-43 polypeptides are related by their reactivity with anti-ATF-43, DNA-binding specificity, complex formation with CREB, heat stability, and phosphorylation by protein kinase A. Certain cell types vary in their ATF-43 complement, suggesting that CREB activity is modulated in a cell-type-specific manner through interaction with ATF-43. ATF-43 polypeptides do not appear simply to correspond to the gene products of the ATF multigene family, suggesting that the size of the ATF family at the protein level is even larger than predicted from cDNA-cloning studies.

Global investigation of protein-protein interactions in yeast Saccharomyces cerevisiae using re-occurring short polypeptide sequences.

PubMed

Pitre, S; North, C; Alamgir, M; Jessulat, M; Chan, A; Luo, X; Green, J R; Dumontier, M; Dehne, F; Golshani, A

2008-08-01

Protein-protein interaction (PPI) maps provide insight into cellular biology and have received considerable attention in the post-genomic era. While large-scale experimental approaches have generated large collections of experimentally determined PPIs, technical limitations preclude certain PPIs from detection. Recently, we demonstrated that yeast PPIs can be computationally predicted using re-occurring short polypeptide sequences between known interacting protein pairs. However, the computational requirements and low specificity made this method unsuitable for large-scale investigations. Here, we report an improved approach, which exhibits a specificity of approximately 99.95% and executes 16,000 times faster. Importantly, we report the first all-to-all sequence-based computational screen of PPIs in yeast, Saccharomyces cerevisiae in which we identify 29,589 high confidence interactions of approximately 2 x 10(7) possible pairs. Of these, 14,438 PPIs have not been previously reported and may represent novel interactions. In particular, these results reveal a richer set of membrane protein interactions, not readily amenable to experimental investigations. From the novel PPIs, a novel putative protein complex comprised largely of membrane proteins was revealed. In addition, two novel gene functions were predicted and experimentally confirmed to affect the efficiency of non-homologous end-joining, providing further support for the usefulness of the identified PPIs in biological investigations.

Investigation of light-induced conformation changes in spiropyran-modified succinylated poly(L-lysine).

PubMed

Cooper, T M; Stone, M O; Natarajan, L V; Crane, R L

1995-08-01

To determine the maximum range of coupling between side-chain photochromism and polypeptide conformation change, we modified the carboxylate side chains of succinylated poly(L-lysine) with a spiropyran to form polypeptide I. The extent of modification was determined to be 35.5%. The spacer group length between the polypeptide alpha-carbon and the dye was 12 atoms, providing minimum polypeptide-dye interaction. Conformation changes were monitored by circular dichroism as a function of light adaptation and solvent composition (hexafluoroisopropanol [HFIP] vs trifluoroethanol [TFE]). Under all solvent compositions, the dark-adapted dye was in the merocyanine form. Light adaptation by visible light converted the dye to the spiropyran form. When dissolved in TFE, I adopted a helical conformation insensitive to light adaptation. With increasing percentage HFIP, a solvent-induced helix-to-coil transition was observed around 80% (vol/vol) HFIP. At 100% HFIP, both light- and dark-adapted forms of I were in the coil state. Near the midpoint of the solvent-induced helix-to-coil transition, light adaptation caused conformation changes. Applying helix-to-coil transition theory, we measured a statistically significant difference in coil segment-HFIP binding constant for light- vs dark-adapted solutions (6.38 +/- 0.03 M-1 vs 6.56 +/- 0.03 M-1), but not for the nucleation parameter sigma (1.2 +/- 0.4 10(-3) vs 1.3 +/- 0.3 x 10(-3). The small binding constant difference translated to a light-induced binding energy difference of 17 cal/mol/monomer. Near the midpoint of the helix-to-coil transition, collective interactions between monomer units made possible the translation of a small energy difference (less than RT) into large macromolecular conformation changes.(ABSTRACT TRUNCATED AT 250 WORDS)

Fluoxetine Ameliorates Behavioral and Neuropathological Deficits in a Transgenic Model Mouse of α-synucleinopathy

PubMed Central

Ubhi, Kiren; Inglis, Chandra; Mante, Michael; Patrick, Christina; Adame, Anthony; Spencer, Brian; Rockenstein, Edward; May, Verena; Winkler, Juergen; Masliah, Eliezer

2013-01-01

The term α-synucleinopathies refers to a group of age-related neurological disorders including Parkinson’s disease (PD), Dementia with Lewy Bodies (DLB) and Multiple System Atrophy (MSA) that display an abnormal accumulation of alpha-synuclein (α-syn). In contrast to the neuronal α-syn accumulation observed in PD and DLB, MSA is characterized by a widespread oligodendrocytic α-syn accumulation. Transgenic mice expressing human α-syn under the oligodendrocyte-specific myelin basic protein promoter (MBP1-hαsyn tg mice) model many of the behavioral and neuropathological alterations observed in MSA. Fluoxetine, a selective serotonin reuptake inhibitor, has been shown to be protective in toxin-induced models of PD, however its effects in an in vivo transgenic model of α-synucleinopathy remain unclear. In this context, this study examined the effect of fluoxetine in the MBP1-hαsyn tg mice, a model of MSA. Fluoxetine adminstration ameliorated motor deficits in the MBP1-hαsyn tg mice, with a concomitant decrease in neurodegenerative pathology in the basal ganglia, neocortex and hippocampus. Fluoxetine adminstration also increased levels of the neurotrophic factors, GDNF (glial-derived neurotrophic factor) and BDNF (brain-derived neurotrophic factor) in the MBP1-hαsyn tg mice compared to vehicle-treated tg mice. This fluoxetine-induced increase in GDNF and BDNF protein levels was accompanied by activation of the ERK signaling pathway. The effects of fluoxetine adminstration on myelin and serotonin markers were also examined. Collectively these results indicate that fluoxetine may represent a novel therapeutic intervention for MSA and other neurodegenerative disorders. PMID:22281106

Updates to the Nomenclature of Platygastroidea in the Zoological Institute of the Russian Academy of Sciences

USDA-ARS?s Scientific Manuscript database

Parabaryconus Kozlov & Kononova n. syn. is treated as a junior synonym of Cremastobaeus Ashmead; Cremastobaeus artus (Kozlov & Kononova) n. comb. is transferred from Parabaryconus; Paridris macrurous Kozlov & Le n. syn. and P. taekuli Talamas & Masner n. syn. are treated as junior synonyms of P. bis...

Nanomechanical properties of distinct fibrillar polymorphs of the protein α-synuclein

PubMed Central

Makky, Ali; Bousset, Luc; Polesel-Maris, Jérôme; Melki, Ronald

2016-01-01

Alpha-synuclein (α-Syn) is a small presynaptic protein of 140 amino acids. Its pathologic intracellular aggregation within the central nervous system yields protein fibrillar inclusions named Lewy bodies that are the hallmarks of Parkinson’s disease (PD). In solution, pure α-Syn adopts an intrinsically disordered structure and assembles into fibrils that exhibit considerable morphological heterogeneity depending on their assembly conditions. We recently established tightly controlled experimental conditions allowing the assembly of α-Syn into highly homogeneous and pure polymorphs. The latter exhibited differences in their shape, their structure but also in their functional properties. We have conducted an AFM study at high resolution and performed a statistical analysis of fibrillar α-Syn shape and thermal fluctuations to calculate the persistence length to further assess the nanomechanical properties of α-Syn polymorphs. Herein, we demonstrated quantitatively that distinct polymorphs made of the same protein (wild-type α-Syn) show significant differences in their morphology (height, width and periodicity) and physical properties (persistence length, bending rigidity and axial Young’s modulus). PMID:27901068

Novel Benzothiazole Derivatives as Fluorescent Probes for Detection of β-Amyloid and α-Synuclein Aggregates.

PubMed

Watanabe, Hiroyuki; Ono, Masahiro; Ariyoshi, Taisuke; Katayanagi, Rikako; Saji, Hideo

2017-08-16

Deposits of β-amyloid (Aβ) and α-synuclein (α-syn) are the hallmark of Alzheimer's disease (AD) and Parkinson's disease (PD), respectively. The detection of these protein aggregates with fluorescent probes is particularly of interest for preclinical studies using fluorescence microscopy on human brain tissue. In this study, we newly designed and synthesized three push-pull benzothiazole (PP-BTA) derivatives as fluorescent probes for detection of Aβ and α-syn aggregates. Fluorescence intensity of all PP-BTA derivatives significantly increased upon binding to Aβ(1-42) and α-syn aggregates in solution. In in vitro saturation binding assays, PP-BTA derivatives demonstrated affinity for both Aβ(1-42) (K d = 40-148 nM) and α-syn (K d = 48-353 nM) aggregates. In particular, PP-BTA-4 clearly stained senile plaques composed of Aβ aggregates in the AD brain section. Moreover, it also labeled Lewy bodies composed of α-syn aggregates in the PD brain section. These results suggest that PP-BTA-4 may serve as a promising fluorescent probe for the detection of Aβ and α-syn aggregates.

Dissociation of glucocerebrosidase dimer in solution by its co-factor, saposin C

DOE PAGES

Gruschus, James M.; Jiang, Zhiping; Yap, Thai Leong; ...

2015-01-16

Mutations in the gene for the lysosomal enzyme glucocerebrosidase (GCase) cause Gaucher disease and are the most common risk factor for Parkinson disease (PD). Analytical ultracentrifugation of 8 μM GCase shows equilibrium between monomer and dimer forms. However, in the presence of its co-factor saposin C (Sap C), only monomer GCase is seen. Isothermal calorimetry confirms that Sap C associates with GCase in solution in a 1:1 complex (K d = 2.1 ± 1.1 μM). Saturation cross-transfer NMR determined that the region of Sap C contacting GCase includes residues 63–66 and 74–76, which is distinct from the region known tomore » enhance GCase activity. Because α-synuclein (α-syn), a protein closely associated with PD etiology, competes with Sap C for GCase binding, its interaction with GCase was also measured by ultracentrifugation and saturation cross-transfer. Unlike Sap C, binding of α-syn to GCase does not affect multimerization. However, adding α-syn reduces saturation cross-transfer from Sap C to GCase, confirming displacement. To explore where Sap C might disrupt multimeric GCase, GCase x-ray structures were analyzed using the program PISA, which predicted stable dimer and tetramer forms. In conclusion, for the most frequently predicted multimer interface, the GCase active sites are partially buried, suggesting that Sap C might disrupt the multimer by binding near the active site.« less

Generation of a neurodegenerative disease mouse model using lentiviral vectors carrying an enhanced synapsin I promoter.

PubMed

Matsuzaki, Yasunori; Oue, Miho; Hirai, Hirokazu

2014-02-15

Certain inherited progressive neurodegenerative disorders, such as spinocerebellar ataxia (SCA), affect neurons in large areas of the central nervous system (CNS). The selective expression of disease-causing and therapeutic genes in susceptible regions and cell types is critical for the generation of animal models and development of gene therapies for these diseases. Previous studies have demonstrated the advantages of the short synapsin I (SynI) promoter (0.5 kb) as a neuron-specific promoter for robust transgene expression. However, the short SynI promoter has also shown some promoter activity in glia and a lack of transgene expression in significant areas of the CNS. New methods: To improve the SynI promoter, we used a SynI promoter that is twice as long (1.0 kb) as the short SynI promoter and incorporated a minimal CMV (minCMV) sequence. We observed that the 1.0 kb rat SynI promoter with minCMV [rSynI(1.0)-minCMV] exhibited robust promoter strength, excellent neuronal specificity and wide-ranging transgene expression throughout the CNS. Comparison with existing methods: Compared with the two previously reported short (0.5 kb) promoters, the new promoter was superior with respect to neuronal specificity and more efficiently transduced neurons. Moreover, transgenic mice expressing the mutant protein ATXN1[Q98], which causes SCA type 1 (SCA1), under the control of the rSynI(1.0)-minCMV promoter showed robust transgene expression specifically in neurons throughout the CNS and exhibited progressive ataxia. rSynI(1.0)-minCMV drives robust and neuron-specific transgene expression throughout the CNS and is therefore useful for viral vector-mediated neuron-specific gene delivery and generation of neuron-specific transgenic animals. Copyright © 2013 Elsevier B.V. All rights reserved.

SynFind: Compiling Syntenic Regions across Any Set of Genomes on Demand.

PubMed

Tang, Haibao; Bomhoff, Matthew D; Briones, Evan; Zhang, Liangsheng; Schnable, James C; Lyons, Eric

2015-11-11

The identification of conserved syntenic regions enables discovery of predicted locations for orthologous and homeologous genes, even when no such gene is present. This capability means that synteny-based methods are far more effective than sequence similarity-based methods in identifying true-negatives, a necessity for studying gene loss and gene transposition. However, the identification of syntenic regions requires complex analyses which must be repeated for pairwise comparisons between any two species. Therefore, as the number of published genomes increases, there is a growing demand for scalable, simple-to-use applications to perform comparative genomic analyses that cater to both gene family studies and genome-scale studies. We implemented SynFind, a web-based tool that addresses this need. Given one query genome, SynFind is capable of identifying conserved syntenic regions in any set of target genomes. SynFind is capable of reporting per-gene information, useful for researchers studying specific gene families, as well as genome-wide data sets of syntenic gene and predicted gene locations, critical for researchers focused on large-scale genomic analyses. Inference of syntenic homologs provides the basis for correlation of functional changes around genes of interests between related organisms. Deployed on the CoGe online platform, SynFind is connected to the genomic data from over 15,000 organisms from all domains of life as well as supporting multiple releases of the same organism. SynFind makes use of a powerful job execution framework that promises scalability and reproducibility. SynFind can be accessed at http://genomevolution.org/CoGe/SynFind.pl. A video tutorial of SynFind using Phytophthrora as an example is available at http://www.youtube.com/watch?v=2Agczny9Nyc. © The Author(s) 2015. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

Clinical Validation of Multiplex Real-Time PCR Assays for Detection of Bacterial Meningitis Pathogens

PubMed Central

Theodore, M. Jordan; Mair, Raydel; Trujillo-Lopez, Elizabeth; du Plessis, Mignon; Wolter, Nicole; Baughman, Andrew L.; Hatcher, Cynthia; Vuong, Jeni; Lott, Lisa; von Gottberg, Anne; Sacchi, Claudio; McDonald, J. Matthew; Messonnier, Nancy E.; Mayer, Leonard W.

2012-01-01

Neisseria meningitidis, Haemophilus influenzae, and Streptococcus pneumoniae are important causes of meningitis and other infections, and rapid, sensitive, and specific laboratory assays are critical for effective public health interventions. Singleplex real-time PCR assays have been developed to detect N. meningitidis ctrA, H. influenzae hpd, and S. pneumoniae lytA and serogroup-specific genes in the cap locus for N. meningitidis serogroups A, B, C, W135, X, and Y. However, the assay sensitivity for serogroups B, W135, and Y is low. We aimed to improve assay sensitivity and develop multiplex assays to reduce time and cost. New singleplex real-time PCR assays for serogroup B synD, W135 synG, and Y synF showed 100% specificity for detecting N. meningitidis species, with high sensitivity (serogroup B synD, 99% [75/76]; W135 synG, 97% [38/39]; and Y synF, 100% [66/66]). The lower limits of detection (LLD) were 9, 43, and 10 copies/reaction for serogroup B synD, W135 synG, and Y synF assays, respectively, a significant improvement compared to results for the previous singleplex assays. We developed three multiplex real-time PCR assays for detection of (i) N. meningitidis ctrA, H. influenzae hpd, and S. pneumoniae lytA (NHS assay); (ii) N. meningitidis serogroups A, W135, and X (AWX assay); and (iii) N. meningitidis serogroups B, C, and Y (BCY assay). Each multiplex assay was 100% specific for detecting its target organisms or serogroups, and the LLD was similar to that for the singleplex assay. Pairwise comparison of real-time PCR between multiplex and singleplex assays showed that cycle threshold values of the multiplex assay were similar to those for the singleplex assay. There were no substantial differences in sensitivity and specificity between these multiplex and singleplex real-time PCR assays. PMID:22170919

The small heat shock proteins αB-crystallin (HSPB5) and Hsp27 (HSPB1) inhibit the intracellular aggregation of α-synuclein.

PubMed

Cox, Dezerae; Ecroyd, Heath

2017-07-01

Protein homeostasis, or proteostasis, is the process of maintaining the conformational and functional integrity of the proteome. Proteostasis is preserved in the face of stress by a complex network of cellular machinery, including the small heat shock molecular chaperone proteins (sHsps), which act to inhibit the aggregation and deposition of misfolded protein intermediates. Despite this, the pathogenesis of several neurodegenerative diseases has been inextricably linked with the amyloid fibrillar aggregation and deposition of α-synuclein (α-syn). The sHsps are potent inhibitors of α-syn aggregation in vitro. However, the limited availability of a robust, cell-based model of α-syn aggregation has, thus far, restricted evaluation of sHsp efficacy in the cellular context. As such, this work sought to establish a robust model of intracellular α-syn aggregation using Neuro-2a cells. Aggregation of α-syn was found to be sensitive to inhibition of autophagy and the proteasome, resulting in a significant increase in the proportion of cells containing α-syn inclusions. This model was then used to evaluate the capacity of the sHsps, αB-c and Hsp27, to prevent α-syn aggregation in cells. To do so, we used bicistronic expression plasmids to express the sHsps. Unlike traditional fluorescent fusion constructs, these bicistronic expression plasmids enable only individual transfected cells expressing the sHsps (via expression of the fluorescent reporter) to be analysed, but without the need to tag the sHsp, which can affect its oligomeric structure and chaperone activity. Overexpression of both αB-c and Hsp27 significantly reduced the intracellular aggregation of α-syn. Thus, these findings suggest that overexpressing or boosting the activity of sHsps may be a way of preventing amyloid fibrillar aggregation of α-syn in the context of neurodegenerative disease.

Clinical validation of multiplex real-time PCR assays for detection of bacterial meningitis pathogens.

PubMed

Wang, Xin; Theodore, M Jordan; Mair, Raydel; Trujillo-Lopez, Elizabeth; du Plessis, Mignon; Wolter, Nicole; Baughman, Andrew L; Hatcher, Cynthia; Vuong, Jeni; Lott, Lisa; von Gottberg, Anne; Sacchi, Claudio; McDonald, J Matthew; Messonnier, Nancy E; Mayer, Leonard W

2012-03-01

Neisseria meningitidis, Haemophilus influenzae, and Streptococcus pneumoniae are important causes of meningitis and other infections, and rapid, sensitive, and specific laboratory assays are critical for effective public health interventions. Singleplex real-time PCR assays have been developed to detect N. meningitidis ctrA, H. influenzae hpd, and S. pneumoniae lytA and serogroup-specific genes in the cap locus for N. meningitidis serogroups A, B, C, W135, X, and Y. However, the assay sensitivity for serogroups B, W135, and Y is low. We aimed to improve assay sensitivity and develop multiplex assays to reduce time and cost. New singleplex real-time PCR assays for serogroup B synD, W135 synG, and Y synF showed 100% specificity for detecting N. meningitidis species, with high sensitivity (serogroup B synD, 99% [75/76]; W135 synG, 97% [38/39]; and Y synF, 100% [66/66]). The lower limits of detection (LLD) were 9, 43, and 10 copies/reaction for serogroup B synD, W135 synG, and Y synF assays, respectively, a significant improvement compared to results for the previous singleplex assays. We developed three multiplex real-time PCR assays for detection of (i) N. meningitidis ctrA, H. influenzae hpd, and S. pneumoniae lytA (NHS assay); (ii) N. meningitidis serogroups A, W135, and X (AWX assay); and (iii) N. meningitidis serogroups B, C, and Y (BCY assay). Each multiplex assay was 100% specific for detecting its target organisms or serogroups, and the LLD was similar to that for the singleplex assay. Pairwise comparison of real-time PCR between multiplex and singleplex assays showed that cycle threshold values of the multiplex assay were similar to those for the singleplex assay. There were no substantial differences in sensitivity and specificity between these multiplex and singleplex real-time PCR assays.

Wnt-related SynGAP1 is a neuroprotective factor of glutamatergic synapses against Aβ oligomers

PubMed Central

Codocedo, Juan F.; Montecinos-Oliva, Carla; Inestrosa, Nibaldo C.

2015-01-01

Wnt-5a is a synaptogenic factor that modulates glutamatergic synapses and generates neuroprotection against Aβ oligomers. It is known that Wnt-5a plays a key role in the adult nervous system and synaptic plasticity. Emerging evidence indicates that miRNAs are actively involved in the regulation of synaptic plasticity. Recently, we showed that Wnt-5a is able to control the expression of several miRNAs including miR-101b, which has been extensively studied in carcinogenesis. However, its role in brain is just beginning to be explored. That is why we aim to study the relationship between Wnt-5a and miRNAs in glutamatergic synapses. We performed in silico analysis which predicted that miR-101b may inhibit the expression of synaptic GTPase-Activating Protein (SynGAP1), a Ras GTPase-activating protein critical for the development of cognition and proper synaptic function. Through overexpression of miR-101b, we showed that miR-101b is able to regulate the expression of SynGAP1 in an hippocampal cell line. Moreover and consistent with a decrease of miR-101b, Wnt-5a enhances SynGAP expression in cultured hippocampal neurons. Additionally, Wnt-5a increases the activity of SynGAP in a time-dependent manner, with a similar kinetic to CaMKII phosphorylation. This also, correlates with a modulation in the SynGAP clusters density. On the other hand, Aβ oligomers permanently decrease the number of SynGAP clusters. Interestingly, when neurons are co-incubated with Wnt-5a and Aβ oligomers, we do not observe the detrimental effect of Aβ oligomers, indicating that, Wnt-5a protects neurons from the synaptic failure triggered by Aβ oligomers. Overall, our findings suggest that SynGAP1 is part of the signaling pathways induced by Wnt-5a. Therefore, possibility exists that SynGAP is involved in the synaptic protection against Aβ oligomers. PMID:26124704

An Overview on the Role of α -Synuclein in Experimental Models of Parkinson's Disease from Pathogenesis to Therapeutics.

PubMed

Javed, Hayate; Kamal, Mohammad Amjad; Ojha, Shreesh

2016-01-01

Parkinson's disease (PD) is a devastating and progressive movement disorder characterized by symptoms of muscles rigidity, tremor, postural instability and slow physical movements. Biochemically, PD is characterized by lack of dopamine production and its action due to loss of dopaminergic neurons and neuropathologically by the presence of intracytoplasmic inclusions known as Lewy bodies, which mainly consist of presynaptic neuronal protein, α-synuclein (α-syn). It is believed that alteration in α-syn homeostasis leads to increased accumulation and aggregation of α-syn in Lewy body. Based on the important role of α-syn from pathogenesis to therapeutics, the recent researches are mainly focused on deciphering the critical role of α-syn at advanced level. Being a major protein in Lewy body that has a key role in pathogenesis of PD, several model systems including immortalized cell lines (SH-SY5Y), primary neuronal cultures, yeast (saccharomyces cerevisiae), drosophila (fruit flies), nematodes (Caenorhabditis elegans) and rodents are being employed to understand the PD pathogenesis and treatment. In order to study the etiopathogensis and develop novel therapeutic target for α -syn aggregation, majority of investigators rely on toxin (rotenone, 1-Methyl-4-Phenyl-1,2,3,6-Tetrahydropyridine, 6-hydroxydopamine, paraquat)-induced animal models of PD as a tool for basic research. Whereas, cell and tissue based models are mostly utilized to elucidate the mechanistic and molecular pathways underlying the α -syn induced toxicity and therapeutic approaches in PD. Gene modified mouse models based on α-syn expression are fascinating for modeling familial PD and toxin induced models provide a suitable approach for sporadic PD. The purpose of this review is to provide a summary and a critical review of the involvement of α-syn in various in vitro and in vivo models of PD based on use of neurotoxins as well as genetic modifications.

SU-G-JeP2-08: Image-Guided Radiation Therapy Using Synthetic CTs in Brain Cancer

DOE Office of Scientific and Technical Information (OSTI.GOV)

Price, R.G.; Glide-Hurst, C.; Henry Ford Health System, Detroit, MI

Purpose: Synthetic-CTs(synCTs) are essential for MR-only treatment planning. However, the performance of synCT for IGRT must be carefully assessed. This work evaluated the accuracy of synCT and synCT-generated DRRs and determined their performance for IGRT in brain cancer radiation therapy. Methods: MR-SIM and CT-SIM images were acquired of a novel anthropomorphic phantom and a cohort of 12 patients. SynCTs were generated by combining an ultra-short echo time (UTE) sequence with other MRI datasets using voxel-based weighted summation. For the phantom, DRRs from synCT and CT were compared via bounding box and landmark analysis. Planar (MV/KV) and volumetric (CBCT) IGRT performancemore » were evaluated across several platforms. In patients, retrospective analysis was conducted to register CBCTs (n=34) to synCTs and CTs using automated rigid registration in the treatment planning system using whole brain and local registration techniques. A semi-automatic registration program was developed and validated to rigidly register planar MV/KV images (n=37) to synCT and CT DRRs. Registration reproducibility was assessed and margin differences were characterized using the van Herk formalism. Results: Bounding box and landmark analysis of phantom synCT DRRs were within 1mm of CT DRRs. Absolute 2D/2D registration shift differences ranged from 0.0–0.7mm for phantom DRRs on all treatment platforms and 0.0–0.4mm for volumetric registrations. For patient planar registrations, mean shift differences were 0.4±0.5mm (range: −0.6–1.6mm), 0.0±0.5mm, (range: −0.9–1.2mm), and 0.1±0.3mm (range: −0.7–0.6mm) for the superior-inferior(S-I), left-right(L–R), and anterior-posterior(A-P) axes, respectively. Mean shift differences in volumetric registrations were 0.6±0.4mm (range: −0.2–1.6mm), 0.2±0.4mm (range: −0.3–1.2mm), and 0.2±0.3mm (range: −0.2–1.2mm) for S-I, L–R, and A–P axes, respectively. CT-SIM and synCT derived margins were within 0.3mm. Conclusion: DRRs generated via synCT agreed well with CT-SIM. Planar and volumetric registrations to synCT-derived targets were comparable to CT. This validation is the next step toward clinical implementation of MR-only planning for the brain. The submitting institution has research agreements with Philips Healthcare. Research sponsored by a Henry Ford Health System Internal Mentored Grant.« less

Some studies on the composition and surface properties of oil bodies from the seed cotyledons of safflower (Carthamus tinctorius) and linseed (Linum ustatissimum).

PubMed Central

Slack, C R; Bertaud, W S; Shaw, B D; Holland, R; Browse, J; Wright, H

1980-01-01

1. The average oil-body diameter in intact cells of developing linseed (Linum usitatissimum) and safflower (Carthamus tinctorius) cotyledons was similar (about 1.4 micrometer), and there was little change in size after oil bodies were isolated and repeatedly washed. 2. The glycerolipid composition of washed oil bodies from both developing and mature cotyledons of the two species was similar; oil bodies from ten different batches of cotyledons contained 4.3 +/- 0.16 mumol of 3-sn-phosphatidylcholine and 25.2 +/- 1.7 mumol of diacylglycerol per 1000 mumol of triacylglycerol. During four successive washings of a once-washed oil-body preparation, the proportion of diacylglycerol to triacylglycerol remained constant and that of 3-sn-phosphatidylcholine to triacylglycerol decreased by only 20%. 3. The protein content of thrice-washed oil bodies from the two species was similar, about 2.4% of the weight of glycerolipids, and appeared to be independent of the stage of cotyledon maturity. Sodium dodecyl sulphate/polyacrylamide-gel electrophoresis indicated that the protein of purified oil bodies from the two species consisted mainly of only four polypeptides and that two of the polypeptides from each species had apparent mol.wts. of 17500 and 15500. Similar patterns of polypeptides were obtained after the hydrolysis of the 15500-mol.wt. polypeptides from linseed and safflower oil bodies by Staphylococcus aureus V8 proteinase, whereas the proteolysis of the 17500-mol.wt. polypeptides from the two species produced different patterns of polypeptides. 4. The 3-sn-phosphatidylcholine in oil-body preparations was hydrolysed about 85% by bee-venom phospholipase A2 without any apparent coalescence of the oil bodies. Incubation with lipase from Rhizopus arrhizus caused rapid coalescence of the oil bodies, and this lipase appeared to initially hydrolyse diacylglycerols in preference to triacylglycerol. 5. Oil bodies from both species were almost completely dispersed in suspensions of pH between 7.1 and 8.3, but formed large aggregates at pH values between 6.7 and 3.9; pH-induced aggregation caused no coalescence. Aggregates formed under acidic conditions were dispersed by re-adjusting the pH of suspensions to 8.3. 6. A freeze-etch electron-microscopic examination of isolated oil bodies indicated that these organelles were bounded by some form of membrane with a particle-free outer surface. Images Fig. 1. Fig. 2. PLATE 1 PLATE 2 PMID:7008782

Residue length and solvation model dependency of elastinlike polypeptides

NASA Astrophysics Data System (ADS)

Bilsel, Mustafa; Arkin, Handan

2010-05-01

We have performed exhaustive multicanonical Monte Carlo simulations of elastinlike polypeptides with a chain including amino acids (valine-proline-glycine-valine-glycine)n or in short (VPGVG)n , where n changes from 1 to 4, in order to investigate the thermodynamic and structural properties. To predict the characteristic secondary structure motifs of the molecules, Ramachandran plots were prepared and analyzed as well. In these studies, we utilized a realistic model where the interactions between all types of atoms were taken into account. Effects of solvation were also simulated by using an implicit-solvent model with two commonly used solvation parameter sets and compared with the vacuum case.

A low molecular weight protein tyrosine phosphatase from Synechocystis sp. strain PCC 6803: enzymatic characterization and identification of its potential substrates

PubMed Central

Mukhopadhyay, Archana; Kennelly, Peter J.

2011-01-01

The predicted protein product of open reading frame slr0328 from Synechocystis sp. PCC 6803, SynPTP, possesses significant amino acid sequence similarity with known low molecular weight protein tyrosine phosphatases (PTPs). To determine the functional properties of this hypothetical protein, open reading frame slr0328 was expressed in Escherichia coli. The purified recombinant protein, SynPTP, displayed its catalytic phosphatase activity towards several tyrosine, but not serine, phosphorylated exogenous protein substrates. The protein phosphatase activity of SynPTP was inhibited by sodium orthovanadate, a known inhibitor of tyrosine phosphatases, but not by okadaic acid, an inhibitor for many serine/threonine phosphatases. Kinetic analysis indicated that the Km and Vmax values for SynPTP towards p-nitrophenyl phosphate are similar to those of other known bacterial low molecular weight PTPs. Mutagenic alteration of the predicted catalytic cysteine of PTP, Cys7, to serine abolished enzyme activity. Using a combination of immunodetection, mass spectrometric analysis and mutagenically altered Cys7SerAsp125Ala-SynPTP, we identified PsaD (photosystem I subunit II), CpcD (phycocyanin rod linker protein) and phycocyanin-α and -β subunits as possible endogenous substrates of SynPTP in this cyanobacterium. These results indicate that SynPTP might be involved in the regulation of photosynthesis in Synechocystis sp. PCC 6803. PMID:21288886

New species and distributional records of Aleocharinae (Coleoptera, Staphylinidae) from Ontario, Canada, with a checklist of recorded species

PubMed Central

Brunke, Adam J.; Klimaszewski, Jan; Dorval, Julie-Anne; Bourdon, Caroline; Paiero, Steven M.; Marshall, Stephen A.

2012-01-01

Abstract The Aleocharinae (Coleoptera: Staphylinidae) of Ontario were reviewed in the context of recently studied material, primarily from insect surveys conducted by the University of Guelph Insect Collection (Ontario, Canada). Aleochara daviesi Klimaszewski & Brunke sp. n., Agaricomorpha websteri Klimaszewski & Brunke sp. n., Atheta (Microdota) alesi Klimaszewski & Brunke sp. n., Dinaraea backusensis Klimaszewski & Brunke sp. n., and Strigota obscurata Klimaszewski & Brunke sp. n. are described as new to science. We also report 47 new Ontario records and 24 new Canadian records. Callicerus rigidicornis (Erichson) and Alevonota gracilenta (Erichson) are newly reported from North America as adventive species. A checklist, with Canadian distributions by province, of the 224 species of Aleocharinae known from Ontario is given. The following species are placed in subjective synonymy with Dexiogyia angustiventris (Casey): (Dexiogyia asperata (Casey) syn. n., Dexiogyia abscissa (Casey) syn. n., Dexiogyia tenuicauda (Casey) syn. n., Dexiogyia intenta (Casey) syn. n., Dexiogyia alticola (Casey) syn. n.). The following species are placed in subjective synonymy with Acrotona subpygmaea (Bernhauer): (Acrotona avia (Casey) syn. n., Acrotona puritana (Casey) syn. n.). Lectotypes are designated for Thiasophila angustiventris Casey, Thiasophila asperata Casey, Ischnoglossa intenta Casey, Oxypoda rubescans Casey, Chilopora americana Casey, Chilopora fuliginosa Casey, Coprothassa smithi Casey, Atheta subpygmaea Bernhauer, Colpodota puritana Casey, Strigota seducens Casey, Trichiusa compacta Casey, Trichiusa hirsuta Casey and Trichiusa robustula Casey. PMID:22577320

Parkinson disease and progressive supranuclear palsy: protein expression in skin.

PubMed

Rodríguez-Leyva, Ildefonso; Chi-Ahumada, Erika G; Carrizales, Juan; Rodríguez-Violante, Mayela; Velázquez-Osuna, Salvador; Medina-Mier, Verónica; Martel-Gallegos, María G; Zarazúa, Sergio; Enríquez-Macías, Lourdes; Castro, Adriana; Calderón-Garcidueñas, Ana Laura; Jiménez-Capdeville, María E

2016-03-01

This study characterizes the expression of tau (p-tau) and α-synuclein (α-syn) by immunohistochemistry in the skin of three different populations: healthy control (HC), Parkinson disease (PD), and progressive supranuclear paralysis (PSP) subjects, with the purpose of finding a biomarker that could differentiate between subjects with PD and PSP. We evaluated the presence of p-tau and α-syn in a pilot study in the skin of three distinct groups of patients: 17 healthy subjects, 17 patients with PD, and 10 patients with PSP. Four millimeters punch biopsies were obtained from the occipital area and analyzed by immunohistochemistry using antibodies against α-syn and phosphorylated species of tau. PHF (paired helical filaments) antibody identifies p-tau in both normal and pathological conditions and AT8 recognizes p-tau characteristic of pathological conditions. Differences between the three groups were assessed by quantification of immunopositive areas in the epidermis. The immunopositivity pattern of p-tau and α-syn was significantly different among the three groups. Healthy subjects showed minimal staining using AT8 and α-syn. The PD group showed significantly higher α-syn and AT8 immunopositivity, while the PSP group only expressed higher AT8 immunopositivity than HCs. These data suggest that the skin reflects brain pathology. Therefore, immunohistochemical analysis of p-tau and α-syn in the skin can be useful for further characterization of PD and PSP.

α-synuclein transfer through tunneling nanotubes occurs in SH-SY5Y cells and primary brain pericytes from Parkinson’s disease patients

PubMed Central

Dieriks, Birger Victor; Park, Thomas I-H.; Fourie, Chantelle; Faull, Richard L. M.; Dragunow, Mike; Curtis, Maurice A.

2017-01-01

Parkinson’s disease (PD) is characterized by the presence of inclusions known as Lewy bodies, which mainly consist of α-synuclein (α-syn) aggregates. There is growing evidence that α-syn self-propagates in non-neuronal cells, thereby contributing to the progression and spread of PD pathology in the brain. Tunneling nanotubes (TNTs) are long, thin, F-actin-based membranous channels that connect cells and have been proposed to act as conduits for α-syn transfer between cells. SH-SY5Y cells and primary human brain pericytes, derived from postmortem PD brains, frequently form TNTs that allow α-syn transfer and long-distance electrical coupling between cells. Pericytes in situ contain α-syn precipitates like those seen in neurons. Exchange through TNTs was rapid, but dependent on the size of the protein. Proteins were able to spread throughout a network of cells connected by TNTs. Transfer through TNTs was not restricted to α-syn; fluorescent control proteins and labeled membrane were also exchanged through TNTs. Most importantly the formation of TNTs and transfer continued during mitosis. Together, our results provide a detailed description of TNTs in SH-SY5Y cells and human brain PD pericytes, demonstrating their role in α-syn transfer and further emphasize the importance that non-neuronal cells, such as pericytes play in disease progression. PMID:28230073

A Synthetic Lethal Screen Identifies a Role for Lin-44/Wnt in C. elegans Embryogenesis.

PubMed

Hartin, Samantha N; Hudson, Martin L; Yingling, Curtis; Ackley, Brian D

2015-01-01

The C. elegans proteins PTP-3/LAR-RPTP and SDN-1/Syndecan are conserved cell adhesion molecules. Loss-of-function (LOF) mutations in either ptp-3 or sdn-1 result in low penetrance embryonic developmental defects. Work from other systems has shown that syndecans can function as ligands for LAR receptors in vivo. We used double mutant analysis to test whether ptp-3 and sdn-1 function in a linear genetic pathway during C. elegans embryogenesis. We found animals with LOF in both sdn-1 and ptp-3 exhibited a highly penetrant synthetic lethality (SynLet), with only a small percentage of animals surviving to adulthood. Analysis of the survivors demonstrated that these animals had a synergistic increase in the penetrance of embryonic developmental defects. Together, these data strongly suggested PTP-3 and SDN-1 function in parallel during embryogenesis. We subsequently used RNAi to knockdown ~3,600 genes predicted to encode secreted and/or transmembrane molecules to identify genes that interacted with ptp-3 or sdn-1. We found that the Wnt ligand, lin-44, was SynLet with sdn-1, but not ptp-3. We used 4-dimensional time-lapse analysis to characterize the interaction between lin-44 and sdn-1. We found evidence that loss of lin-44 caused defects in the polarization and migration of endodermal precursors during gastrulation, a previously undescribed role for lin-44 that is strongly enhanced by the loss of sdn-1. PTP-3 and SDN-1 function in compensatory pathways during C. elegans embryonic and larval development, as simultaneous loss of both genes has dire consequences for organismal survival. The Wnt ligand lin-44 contributes to the early stages of gastrulation in parallel to sdn-1, but in a genetic pathway with ptp-3. Overall, the SynLet phenotype provides a robust platform to identify ptp-3 and sdn-1 interacting genes, as well as other genes that function in development, yet might be missed in traditional forward genetic screens.

A Synthetic Lethal Screen Identifies a Role for Lin-44/Wnt in C. elegans Embryogenesis

PubMed Central

Hartin, Samantha N.; Hudson, Martin L.; Yingling, Curtis; Ackley, Brian D.

2015-01-01

Background The C. elegans proteins PTP-3/LAR-RPTP and SDN-1/Syndecan are conserved cell adhesion molecules. Loss-of-function (LOF) mutations in either ptp-3 or sdn-1 result in low penetrance embryonic developmental defects. Work from other systems has shown that syndecans can function as ligands for LAR receptors in vivo. We used double mutant analysis to test whether ptp-3 and sdn-1 function in a linear genetic pathway during C. elegans embryogenesis. Results We found animals with LOF in both sdn-1 and ptp-3 exhibited a highly penetrant synthetic lethality (SynLet), with only a small percentage of animals surviving to adulthood. Analysis of the survivors demonstrated that these animals had a synergistic increase in the penetrance of embryonic developmental defects. Together, these data strongly suggested PTP-3 and SDN-1 function in parallel during embryogenesis. We subsequently used RNAi to knockdown ~3,600 genes predicted to encode secreted and/or transmembrane molecules to identify genes that interacted with ptp-3 or sdn-1. We found that the Wnt ligand, lin-44, was SynLet with sdn-1, but not ptp-3. We used 4-dimensional time-lapse analysis to characterize the interaction between lin-44 and sdn-1. We found evidence that loss of lin-44 caused defects in the polarization and migration of endodermal precursors during gastrulation, a previously undescribed role for lin-44 that is strongly enhanced by the loss of sdn-1. Conclusions PTP-3 and SDN-1 function in compensatory pathways during C. elegans embryonic and larval development, as simultaneous loss of both genes has dire consequences for organismal survival. The Wnt ligand lin-44 contributes to the early stages of gastrulation in parallel to sdn-1, but in a genetic pathway with ptp-3. Overall, the SynLet phenotype provides a robust platform to identify ptp-3 and sdn-1 interacting genes, as well as other genes that function in development, yet might be missed in traditional forward genetic screens. PMID:25938228

Molecular and Behavioral Changes Associated with Adult Hippocampus-Specific SynGAP1 Knockout

ERIC Educational Resources Information Center

Muhia, Mary; Willadt, Silvia; Yee, Benjamin K.; Feldon, Joram; Paterna, Jean-Charles; Schwendener, Severin; Vogt, Kaspar; Kennedy, Mary B.; Knuesel, Irene

2012-01-01

The synaptic Ras/Rap-GTPase-activating protein (SynGAP1) plays a unique role in regulating specific downstream intracellular events in response to N-methyl-D-aspartate receptor (NMDAR) activation. Constitutive heterozygous loss of SynGAP1 disrupts NMDAR-mediated physiological and behavioral processes, but the disruptions might be of developmental…

Folding of a single domain protein entering the endoplasmic reticulum precedes disulfide formation.

PubMed

Robinson, Philip J; Pringle, Marie Anne; Woolhead, Cheryl A; Bulleid, Neil J

2017-04-28

The relationship between protein synthesis, folding, and disulfide formation within the endoplasmic reticulum (ER) is poorly understood. Previous studies have suggested that pre-existing disulfide links are absolutely required to allow protein folding and, conversely, that protein folding occurs prior to disulfide formation. To address the question of what happens first within the ER, that is, protein folding or disulfide formation, we studied folding events at the early stages of polypeptide chain translocation into the mammalian ER using stalled translation intermediates. Our results demonstrate that polypeptide folding can occur without complete domain translocation. Protein disulfide isomerase (PDI) interacts with these early intermediates, but disulfide formation does not occur unless the entire sequence of the protein domain is translocated. This is the first evidence that folding of the polypeptide chain precedes disulfide formation within a cellular context and highlights key differences between protein folding in the ER and refolding of purified proteins. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Effect of Sequence Blockiness on the Morphologies of Surface-grafted Elastin-like Polypeptides

NASA Astrophysics Data System (ADS)

Albert, Julie; Sintavanon, Kornkanok; Mays, Robin; MacEwan, Sarah; Chilkoti, Ashutosh; Genzer, Jan

2014-03-01

The inter- and intra- molecular interactions among monomeric units of copolymers and polypeptides depend strongly on monomer sequence distribution and dictate the phase behavior of these species both in solution and on surfaces. To study the relationship between sequence and phase behavior, we have designed a series of elastin-like polypeptides (ELPs) with controlled monomer sequences that mimic copolymers with various co-monomer sequence distributions and attached them covalently to silicon substrates from buffer solutions at temperatures below and above the bulk ELPs' lower critical solution temperatures (LCSTs). The dependence of ELP grafting density on solution temperature was examined by ellipsometry and the resultant surface morphologies were examined in air and under water with atomic force microscopy. Depositions performed above the LCST resulted in higher grafting densities and greater surface roughness of ELPs relative to depositions carried out below the LCST. In addition, we are using gradient substrates to examine the effect of ELP grafting density on temperature responsiveness.

Assignment of two mitochondrially synthesized polypeptides to human mitochondrial DNA and their use in the study of intracellular mitochondrial interaction.

PubMed Central

Oliver, N A; Wallace, D C

1982-01-01

Two mitochondrially synthesized marker polypeptides, MV-1 and MV-2, were found in human HeLa and HT1080 cells. These were assigned to the mitochondrial DNA in HeLa-HT1080 cybrids and hybrids by demonstrating their linkage to cytoplasmic genetic markers. These markers include mitochondrial DNA restriction site polymorphisms and resistance to chloramphenicol, an inhibitor of mitochondrial protein synthesis. In the absence of chloramphenicol, the expression of MV-1 and MV-2 in cybrids and hybrids was found to be directly proportional to the ratio of the parental mitochondrial DNAs. In the presence of chloramphenicol, the marker polypeptide linked to the chloramphenicol-sensitive mitochondrial DNA continued to be expressed. This demonstrated that resistant and sensitive mitochondrial DNAs can cooperate within a cell for gene expression and that the CAP-resistant allele was dominant or codominant to sensitive. Such cooperation suggests that mitochondrial DNAs can be exchanged between mitochondria. Images PMID:6955589

BAG-6 is essential for selective elimination of defective proteasomal substrates

PubMed Central

Minami, Ryosuke; Hayakawa, Atsuko; Kagawa, Hiroki; Yanagi, Yuko; Yokosawa, Hideyoshi

2010-01-01

BAG-6/Scythe/BAT3 is a ubiquitin-like protein that was originally reported to be the product of a novel gene located within the human major histocompatibility complex, although the mechanisms of its function remain largely obscure. Here, we demonstrate the involvement of BAG-6 in the degradation of a CL1 model defective protein substrate in mammalian cells. We show that BAG-6 is essential for not only model substrate degradation but also the ubiquitin-mediated metabolism of newly synthesized defective polypeptides. Furthermore, our in vivo and in vitro analysis shows that BAG-6 interacts physically with puromycin-labeled nascent chain polypeptides and regulates their proteasome-mediated degradation. Finally, we show that knockdown of BAG-6 results in the suppressed presentation of MHC class I on the cell surface, a procedure known to be affected by the efficiency of metabolism of defective ribosomal products. Therefore, we propose that BAG-6 is necessary for ubiquitin-mediated degradation of newly synthesized defective polypeptides. PMID:20713601

The use of phage display in neurobiology.

PubMed

Bradbury, Andrew R M

2010-04-01

Phage display has been extensively used to study protein-protein interactions, receptor- and antibody-binding sites, and immune responses, to modify protein properties, and to select antibodies against a wide range of different antigens. In the format most often used, a polypeptide is displayed on the surface of a filamentous phage by genetic fusion to one of the coat proteins, creating a chimeric coat protein, and coupling phenotype (the protein) to genotype (the gene within). As the gene encoding the chimeric coat protein is packaged within the phage, selection of the phage on the basis of the binding properties of the polypeptide displayed on the surface simultaneously results in the isolation of the gene encoding the polypeptide. This unit describes the background to the technique, and illustrates how it has been applied to a number of different problems, each of which has its neurobiological counterparts. Although this overview concentrates on the use of filamentous phage, which is the most popular platform, other systems are also described. (c) 2010 by John Wiley & Sons, Inc.

Amyloidogenic Regions and Interaction Surfaces Overlap in Globular Proteins Related to Conformational Diseases

PubMed Central

Castillo, Virginia; Ventura, Salvador

2009-01-01

Protein aggregation underlies a wide range of human disorders. The polypeptides involved in these pathologies might be intrinsically unstructured or display a defined 3D-structure. Little is known about how globular proteins aggregate into toxic assemblies under physiological conditions, where they display an initially folded conformation. Protein aggregation is, however, always initiated by the establishment of anomalous protein-protein interactions. Therefore, in the present work, we have explored the extent to which protein interaction surfaces and aggregation-prone regions overlap in globular proteins associated with conformational diseases. Computational analysis of the native complexes formed by these proteins shows that aggregation-prone regions do frequently overlap with protein interfaces. The spatial coincidence of interaction sites and aggregating regions suggests that the formation of functional complexes and the aggregation of their individual subunits might compete in the cell. Accordingly, single mutations affecting complex interface or stability usually result in the formation of toxic aggregates. It is suggested that the stabilization of existing interfaces in multimeric proteins or the formation of new complexes in monomeric polypeptides might become effective strategies to prevent disease-linked aggregation of globular proteins. PMID:19696882

Interplay of signal recognition particle and trigger factor at L23 near the nascent chain exit site on the Escherichia coli ribosome

PubMed Central

Ullers, Ronald S.; Houben, Edith N.G.; Raine, Amanda; ten Hagen-Jongman, Corinne M.; Ehrenberg, Måns; Brunner, Joseph; Oudega, Bauke; Harms, Nellie; Luirink, Joen

2003-01-01

As newly synthesized polypeptides emerge from the ribosome, they interact with chaperones and targeting factors that assist in folding and targeting to the proper location in the cell. In Escherichia coli, the chaperone trigger factor (TF) binds to nascent polypeptides early in biosynthesis facilitated by its affinity for the ribosomal proteins L23 and L29 that are situated around the nascent chain exit site on the ribosome. The targeting factor signal recognition particle (SRP) interacts specifically with the signal anchor (SA) sequence in nascent inner membrane proteins (IMPs). Here, we have used photocross-linking to map interactions of the SA sequence in a short, in vitro–synthesized, nascent IMP. Both TF and SRP were found to interact with the SA with partially overlapping binding specificity. In addition, extensive contacts with L23 and L29 were detected. Both purified TF and SRP could be cross-linked to L23 on nontranslating ribosomes with a competitive advantage for SRP. The results suggest a role for L23 in the targeting of IMPs as an attachment site for TF and SRP that is close to the emerging nascent chain. PMID:12756233

Primary light-harvesting system: phycobilisomes and associated membranes. Progress report, January 1, 1981-December 31, 1981

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gantt, E.

1981-01-01

Phycobilisomes, serving as primary light harvesting complexes in cyanobacteria and red algae, were investigated. Structurally the phycobilisomes of both groups have the same fundamental phycobiliprotein arrangement. Allophycocyanin is in the center near the thylakoid. Stacked rods composed of phycocyanin, or phycocyanin-phycoerythrin radiate peripherally from the allophycocyanin core. Phycobilisomes of Nostoc sp. and Fremyella diplosiphon, after separation into separate allophycocyanin and phycoerythrin-phycocyanin fractions have been associated in vitro. Hybrid phycobilisomes, derived from mixtures of phycobiliprotein from these species were also obtained. The interaction is specific since reassociation was not obtained with phycobiliprotein complexes of some other algae. Phycobilisomes, whether native, ormore » associated in vitro, were similar in their sedimentation, absorption, fluorescence excitation, fluorescence emission, and by electron microscopy. Furthermore, many of the colorless polypeptides were also highly similar between Nostoc and Fremyella. The similarity formed may reflect an evolutionary relationship between the two species. The polypeptide composition of Porphyridium cruentum phycobilisomes is the most complex of any thus far examined. The phycobiliprotein containing polypeptides comprised 84% of the total stainable protein, while the remaining were colorless. Most of the colorless polypeptides occurred in a pelletable fraction, which was enriched in allophycocyanin and phycocyanin, it is probable that some are involved in the linking of these phycobiliproteins.« less

Hypertension (High Blood Pressure)

MedlinePlus

... Throughout the 1960s and 1970s, the results of observational studies further strengthened the causal relationship between high ... polypeptide). Results of this study may lead to methods to regulate ANP and thereby prevent ... research that examines the interaction of race/ethnicity with ...

Synthesis, Characterization, and Crystallization of Syndiotactic Alternating Ethylene-Propylene Crystalline Copolymer and its Blends.

NASA Astrophysics Data System (ADS)

Chien, Haoyang

A syndiotactic alternating ethylene-propylene (SYN-ALT-EP) crystalline copolymer was synthesized by complete hydrogenation, using a diimide reduction, of syndiotactic cis-1,4-poly(pentadiene-1,3) (CIS-PPD). The microstructure was studied by both high resolution nuclear magnetic resonance (NMR) spectroscopy and also fourier transform infra-red (FTIR) spectroscopy. The number average length of syndiotactic sequences is about 69 which indicates a high degree of syndiotacticity (97%) in the microstructure of this copolymer. The single FTIR absorbance at 733 cm^{ -1} without any splitting suggests an alternating arrangement of ethylene and propylene units. The solution state characterization of SYN-ALT -EP was studied by gel permeation chromatography using on -line measurements of multi-angle laser light scattering (MALLS), single capillary viscosities (VISC), and concentrations by differential refractive index (DRI) detectors. The Mark-Houwink-Sakurada parameters of "K" and "a" in THF at 30^circC are determined to be 8.99 times 10^ {-5} and 0.8, respectively. The universal GPC calibration curve can be applied to this copolymer in THF at 30^circC. Two different molecular relaxation processes ( alpha and beta relaxations) were found via dynamic mechanical (DM) analysis below room temperature: an alpha relaxation (around -60^ circC) and a beta relaxation (around -125^circ C). The apparent activation energy of the alpha relaxation is 285 kJ/mol, and the activation energy of the beta relaxation is 43 kJ/mol based on the Arrhenius equation. Molecular motion in SYN-ALT-EP copolymer was probed by solid state ^{13}C NMR experiments. At temperatures above T_{rm g} there are two major molecular motions in this copolymer: a backbone motion (the rotational motion about single bonds) and a methyl side group rotation. The backbone motion is frozen below T_{rm g}, but the methyl rotation still occurs. As the temperature is further decreased to about -175 ^circC, well below the beta -transition observed in DM analysis, the methyl side group rotation slows down, suggesting that the methyl rotation may be associated with the observed beta relaxation process. The equilibrium melting temperature is 55 +/- 1^circC; the equilibrium heat of fusion is 8.8 +/- 0.3 kJ/mol. The overall crystallization kinetics show an Avrami exponent (n) that qualitatively increases with crystallization temperature during primary crystallization. The transition from Regime II to Regime III is observed near T_{rm c} = 26 ^circC based on linear crystal growth rate experiments. The fold surface free energy ( sigma_{rm e}) is determined to be 33 erg/cm^2. A monoclinic crystal unit cell was determined (a = 11.19A b = 11.82A c = 9.00A gamma = 67.03^circ) from the fiber pattern via wide angle x-ray diffraction experiments (WAXD). A banded spherulitic morphology was observed by polarized light microscopy (PLM) and transmission electron microscopy (TEM). Such texture is characteristic of the co-twisting of growing lamellae. The morphology changes from regularly banded spherulites to non-regularly banded spherulites and may be correlated with the Regime III to Regime II transition. A plate-like single crystal morphology was also observed by polarized light microscopy after a melt crystallization at small supercooling conditions. Blends of SYN-ALT-EP/IPP, SYN-ALT-EP/HDPE, and SYN-ALT-EP/LDPE were made and examined. Neither T _{rm g} shifting nor co-crystallization using different blending compositions were observed. Therefore, only limited, if any, miscibility exists in these blends.

The type specimens of Tenthredo Linnaeus, 1758 (Hymenoptera: Tenthredinidae) deposited in the Hungarian Natural History Museum.

PubMed

Taeger, Andreas

2013-01-01

The type specimens of Tenthredo sensu lato deposited in the Hungarian Natural History Museum (Budapest) are reviewed. The following synonyms are recognized and discussed: Rhogogastera aenescens Mocsáry, 1909, syn. nov. of Tenthredo (Temuledo) finschi finschi W.F. Kirby, 1882; Macrophya prasinipes Konow, 1891, syn. nov. of Macrophya caucasica (Mocsáry, 1880); Tenthredo concinnoides Malaise, 1945, syn. nov. of Tenthredo concinna Mocsáry, 1883; Tenthredo fulviventris Mocsáry, 1909, syn. nov. of Tenthredo (Tenthredella) crenata (Enslin, 1912); Tenthredo vespa inaffectata Muche, 1965, syn. nov. of Tenthredo (Tenthredo) vespa Retzius, 1783; Rhogogaster kaszabi Zombori, 1973, syn. nov. of Tenthredo (Eurogaster) aaliensis (Strand, 1898); Tenthredo kaszabi Muche, 1965, syn. nov. of Tenthredo (Eurogaster) mesomela Linné, 1758; Rhogogastera opacella Mocsáry, 1909, syn. nov. of Tenthredo (Eurogaster) stulta Jakowlew, 1891; Tenthredo tschinggischanensis Muche, 1965 syn. nov. of Tenthredo (Tenthredella) balteata Klug, 1817; Tenthredo unfasciata Mocsáry, 1909, syn. nov. of Tenthredo (Tenthredella) colon Klug, 1817; Tenthredella cucullata Enslin, 1912, syn. nov. of Tenthredo (Tenthredella) colon Klug, 1817. Macrophya caucasica (Mocsáry, 1880) is a new combination (comb. nov.) for Allantus caucasicus Mocsáry, 1880. Tenthredo semicolon Mol, is a replacement name (nom. nov.) for Tenthredo punctulata Konow, 1887, nom. praeocc., nec Klug, 1817. Tenthredo chanae Taeger & Shinohara spec. nov., a species close to Tenthredo concinna, is described from Taiwan. Lectotypes are designated and illustrated for the following 30 nominal taxa: Rhogogastera aenescens Mocsáry, 1909; Allantus albiventris Mocsáry, 1880; Allantus almasyanus Mocsáry, 1909; Macrophya binaculata Mocsáry, 1909; Allantus brachycerus Mocsáry, 1909; Allantus caucasicus Mocsáry, 1880; Allantus centrorufus Mocsáry, 1909; Tenthredo chyzeri Mocsáry, 1891; Tenthredo concinna Mocsáry, 1883; Tenthredo concinnoides Malaise, 1945; Allantus eburneus Mocsáry, 1909; Allantus eburneus Mocsáry, 1909; Allantus fulvicornis Mocsáry, 1909; Tenthredo fulvicornis Mocsáry, 1909; Allantus fulvipennis Mocsáry, 1909; Tenthredo fulviventris Mocsáry, 1909; Tenthredo gribodoi Konow, 1898; Allantus lateralis Mocsáry, 1909; Allantus limbiferus Mocsáry, 1891; Allantus moestus Mocsáry, 1883; Rhogogastera nigrita Mocsáry, 1909; Allantus obesus Mocsáry, 1880; Macrophya prasinipes Konow, 1891; Tenthredo punctulata Konow, 1887; Allantus rufipes Mocsáry, 1909; Allantus sabariensis Mocsáry, 1880; Allantus sanguinolentus Mocsáry, 1909; Allantus temulus var. scutellatus Mocsáry, 1909; Allantus testaceus Mocsáry, 1909; Allantus variabilis Mocsáry, 1909. The first records of Tenthredo concinna for Vietnam and Nepal are given.

Cellobiohydrolase I enzymes

DOEpatents

Adney, William S; Himmel, Michael E; Decker, Stephen R; Knoshaug, Eric P; Nimlos, Mark R; Crowley, Michael F; Jeoh, Tina

2014-01-28

Provided herein is an isolated Cel7A polypeptide comprising mutations in the catalytic domain of the polypeptide relative to the catalytic domain of a wild type Cel7A polypeptide, wherein the mutations reduce N-linked glycosylation of the isolated polypeptide relative to the wild type polypeptide. Also provided herein is an isolated Cel7A polypeptide comprising increased O-linked glycosylation of the linker domain relative to a linker domain of a wild type Cel7A polypeptide. The increased O-linked glycosylation is a result of the addition of and/or substitution of one or more serine and/or threonine residues to the linker domain relative to the linker domain of the wild type polypeptide. In some embodiments, the isolated Cel7A polypeptide comprising mutations in the catalytic domain of the polypeptide relative to the catalytic domain of a wild type Cel7A polypeptide further comprises increased O-linked glycosylation of the linker domain relative to a linker domain of a wild type Cel7A polypeptide. The mutations in the catalytic domain reduce N-linked glycosylation of the isolated polypeptide relative to the wild type polypeptide. The addition of and/or substitution of one or more serine and/or threonine residues to the linker domain relative to the linker domain of the wild type polypeptide increases O-linked glycosylation of the isolated polypeptide. Further provided are compositions comprising such polypeptides and nucleic acids encoding such polypeptides. Still further provided are methods for making such polypeptides.

FTY720/Fingolimod Reduces Synucleinopathy and Improves Gut Motility in A53T Mice

PubMed Central

Vidal-Martínez, Guadalupe; Vargas-Medrano, Javier; Gil-Tommee, Carolina; Medina, David; Garza, Nathan T.; Yang, Barbara; Segura-Ulate, Ismael; Dominguez, Samantha J.; Perez, Ruth G.

2016-01-01

Patients with Parkinson's disease (PD) often have aggregated α-synuclein (aSyn) in enteric nervous system (ENS) neurons, which may be associated with the development of constipation. This occurs well before the onset of classic PD motor symptoms. We previously found that aging A53T transgenic (Tg) mice closely model PD-like ENS aSyn pathology, making them appropriate for testing potential PD therapies. Here we show that Tg mice overexpressing mutant human aSyn develop ENS pathology by 4 months. We then evaluated the responses of Tg mice and their WT littermates to the Food and Drug Administration-approved drug FTY720 (fingolimod, Gilenya) or vehicle control solution from 5 months of age. Long term oral FTY720 in Tg mice reduced ENS aSyn aggregation and constipation, enhanced gut motility, and increased levels of brain-derived neurotrophic factor (BDNF) but produced no significant change in WT littermates. A role for BDNF was directly assessed in a cohort of young A53T mice given vehicle, FTY720, the Trk-B receptor inhibitor ANA-12, or FTY720 + ANA-12 from 1 to 4 months of age. ANA-12-treated Tg mice developed more gut aSyn aggregation as well as constipation, whereas FTY720-treated Tg mice had reduced aSyn aggregation and less constipation, occurring in part by increasing both pro-BDNF and mature BDNF levels. The data from young and old Tg mice revealed FTY720-associated neuroprotection and reduced aSyn pathology, suggesting that FTY720 may also benefit PD patients and others with synucleinopathy. Another finding was a loss of tyrosine hydroxylase immunoreactivity in gut neurons with aggregated aSyn, comparable with our prior findings in the CNS. PMID:27528608

Dietary intake and the development of the metabolic syndrome: the Atherosclerosis Risk in Communities study.

PubMed

Lutsey, Pamela L; Steffen, Lyn M; Stevens, June

2008-02-12

The role of diet in the origin of metabolic syndrome (MetSyn) is not well understood; thus, we sought to evaluate the relationship between incident MetSyn and dietary intake using prospective data from 9514 participants (age, 45 to 64 years) enrolled in the Atherosclerosis Risk in Communities (ARIC) study. Dietary intake was assessed at baseline via a 66-item food frequency questionnaire. We used principal-components analysis to derive "Western" and "prudent" dietary patterns from 32 food groups and evaluated 10 food groups used in previous studies of the ARIC cohort. MetSyn was defined by American Heart Association guidelines. Proportional-hazards regression was used. Over 9 years of follow-up, 3782 incident cases of MetSyn were identified. After adjustment for demographic factors, smoking, physical activity, and energy intake, consumption of a Western dietary pattern (P(trend)=0.03) was adversely associated with incident MetSyn. After further adjustment for intake of meat, dairy, fruits and vegetables, refined grains, and whole grains, analysis of individual food groups revealed that meat (P(trend)<0.001), fried foods (P(trend)=0.02), and diet soda (P(trend)=< 0.001) also were adversely associated with incident MetSyn, whereas dairy consumption (P(trend)=0.006) was beneficial. No associations were observed between incident MetSyn and a prudent dietary pattern or intakes of whole grains, refined grains, fruits and vegetables, nuts, coffee, or sweetened beverages. These prospective findings suggest that consumption of a Western dietary pattern, meat, and fried foods promotes the incidence of MetSyn, whereas dairy consumption provides some protection. The diet soda association was not hypothesized and deserves further study.

Fluoxetine ameliorates behavioral and neuropathological deficits in a transgenic model mouse of α-synucleinopathy.

PubMed

Ubhi, Kiren; Inglis, Chandra; Mante, Michael; Patrick, Christina; Adame, Anthony; Spencer, Brian; Rockenstein, Edward; May, Verena; Winkler, Juergen; Masliah, Eliezer

2012-04-01

The term α-synucleinopathies refers to a group of age-related neurological disorders including Parkinson's disease (PD), Dementia with Lewy Bodies (DLB) and Multiple System Atrophy (MSA) that display an abnormal accumulation of alpha-synuclein (α-syn). In contrast to the neuronal α-syn accumulation observed in PD and DLB, MSA is characterized by a widespread oligodendrocytic α-syn accumulation. Transgenic mice expressing human α-syn under the oligodendrocyte-specific myelin basic protein promoter (MBP1-hαsyn tg mice) model many of the behavioral and neuropathological alterations observed in MSA. Fluoxetine, a selective serotonin reuptake inhibitor, has been shown to be protective in toxin-induced models of PD, however its effects in an in vivo transgenic model of α-synucleinopathy remain unclear. In this context, this study examined the effect of fluoxetine in the MBP1-hαsyn tg mice, a model of MSA. Fluoxetine administration ameliorated motor deficits in the MBP1-hαsyn tg mice, with a concomitant decrease in neurodegenerative pathology in the basal ganglia, neocortex and hippocampus. Fluoxetine administration also increased levels of the neurotrophic factors, GDNF (glial-derived neurotrophic factor) and BDNF (brain-derived neurotrophic factor) in the MBP1-hαsyn tg mice compared to vehicle-treated tg mice. This fluoxetine-induced increase in GDNF and BDNF protein levels was accompanied by activation of the ERK signaling pathway. The effects of fluoxetine administration on myelin and serotonin markers were also examined. Collectively these results indicate that fluoxetine may represent a novel therapeutic intervention for MSA and other neurodegenerative disorders. Copyright © 2011 Elsevier Inc. All rights reserved.

Revision of the types of male Sciaridae (Diptera) described from Australia by F.A.A. Skuse.

PubMed

Broadley, Adam; Kauschke, Ellen; Mohrig, Werner

2016-11-17

A total of 27 male sciarid types described by Skuse (1888 and 1890), held in the Australian National Insect Collection, Canberra, and the Australian Museum, Sydney, were remounted and examined microscopically. Of these, 25 species were described as Sciara Meigen, one as Zygoneura Meigen and one as Trichosia Winnertz. Revision of these species revealed the following: 13 species belong to the genus Bradysia Winnertz (B. amabilis, B. conjuncta, B. crassicornis, B. exsequialis, B. frequens, B. froggatti, B. luctifica, B. maesta, B. mastersi, B. ornatula, B. pernitida, B. pictipes, B. unica), 1 species to the genus Corynoptera Winnertz (C. minutela), 4 species to the genus Austrosciara Schmitz & Mjöberg (Aus. infrequens, Aus. montivaga, Aus. spectabilis, Aus. winnertzi), 2 species to the genus Pseudolycoriella Menzel & Mohrig (Psl. cavatica, Psl. ignobilis), 1 species to the genus Pseudozygomma Mohrig (Pseudoz. maculipennis), 1 species to the genus Sciara Meigen (Sc. tryoni), and 1 species to the genus Scythropochroa Enderlein (Scyth. macleayi). In total 26 species were new combinations. Eight species names were declared as new synonyms: Bradysia pictipes (Skuse, 1888) = Sciara notata Skuse, 1888 syn. n. and = Bradysia seticornis Vilkamaa, Hippa & Mohrig, 2012 (from New Caledonia) syn. n.; Bradysia conjuncta (Skuse, 1890) = Sciara serenipennis Skuse, 1890 syn. n.; Pseudolycoriella cavatica (Skuse, 1888) = Sciara familiaris Skuse, 1888 syn. n. and = Sciara festiva Skuse, 1888 syn. n.; Bradysia luctifica (Skuse, 1888) = Bradysia planistylata Vilkamaa, Hippa & Mohrig, 2012 syn. n.; Sciara tryoni Skuse, 1890 = Sciara insulana Vilkamaa, Hippa & Mohrig, 2015 syn. n. (both species are from New Caledonia); Austrosciara winnertzi (Skuse, 1888) = Sciara rufulenta Edwards, 1927 syn. n. (from New Zealand). Lectotype specimens were designated for 17 species in order to fix the names.

Syn- and anti-conformations of 5'-deoxy- and 5'-O-methyl-uridine 2',3'-cyclic monophosphate.

PubMed

Grabarkiewicz, Tomasz; Hoffmann, Marcin

2006-01-01

Two uridine 2',3'-cyclic monophosphate (cUMP) derivatives, 5'-deoxy (DcUMP) and 5'-O-methyl (McUMP), were studied by means of quantum chemical methods. Aqueous solvent effects were estimated based on the isodensity-surface polarized-continuum model (IPCM). Gas phase calculations revealed only slight energy differences between the syn- and anti-conformers of both compounds: the relative energies of the syn-structure are -0.9 and 0.2 kcal mol(-1) for DcUMP and McUMP, respectively. According to the results from the IPCM calculations, however, both syn-conformers become about 14 kcal mol(-1) more stable in aqueous solution than their corresponding anti-structures. Additionally, the effects of a countercation and protonation on DcUMP were studied, revealing that the syn-structure is also favored over the anti-one for these systems.

Basic Research on Processing of Ceramics for Space Structures

DTIC Science & Technology

1984-10-01

form. 110 V. References 1. B. Fegley, E. A. Barringer , and H. K. Bowen, J. Am. Ceram. Soc., 1984, 67(6), C113-16. 2. S. Govil and R . C. Mehrotra, Syn...Avenue, Cambridge, MA 02IApt- R h. 8 9 1 708 9. SFONSO1= UNG# WN AGENCY NAME(S) AND 1041SLI. SPONSORING / moNITORING AFOS AO~~IfiI~ jAGENCY REPORT...NUMBER AFOS-83- 192 BLDG O4107OIVIAIUYSAEETlb.OSRUINC I AB DCT 2033g2644 w’wJELE OR8309 U n i t o r mW c p o i t A O Z r 2 1 9 8 9- . 2 ’ a d S C A F w r

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rycyna, R.E.; Alderfer, J.L.

Uridylyl(3'-5')uridine (UpU) is subjected to aqueous acetone photosensitized radiation with sunlamps. These irradiation conditions form only cyclobutane-type photodimers. Purification of a specific configurational photodimer is accomplished by using C-18 reverse-phase high-performance liquid chromatography. Multinuclear NMR analysis is used to analyze photoproduct formation and to determine conformational features of these photodimers. Four photodimers are identified, with the cis-syn isomer predominant. The cis-syn and trans-syn photodimers of UpU exhibit markedly different furanose and exocyclic bond conformations. A comparison of the properties of the cis-syn dimers of UpU with those of dTpdT reveal many similar conformational features but also some that are different.

ImSyn: photonic image synthesis applied to synthetic aperture radar, microscopy, and ultrasound imaging

NASA Astrophysics Data System (ADS)

Turpin, Terry M.; Lafuse, James L.

1993-02-01

ImSynTM is an image synthesis technology, developed and patented by Essex Corporation. ImSynTM can provide compact, low cost, and low power solutions to some of the most difficult image synthesis problems existing today. The inherent simplicity of ImSynTM enables the manufacture of low cost and reliable photonic systems for imaging applications ranging from airborne reconnaissance to doctor's office ultrasound. The initial application of ImSynTM technology has been to SAR processing; however, it has a wide range of applications such as: image correlation, image compression, acoustic imaging, x-ray tomographic (CAT, PET, SPECT), magnetic resonance imaging (MRI), microscopy, range- doppler mapping (extended TDOA/FDOA). This paper describes ImSynTM in terms of synthetic aperture microscopy and then shows how the technology can be extended to ultrasound and synthetic aperture radar. The synthetic aperture microscope (SAM) enables high resolution three dimensional microscopy with greater dynamic range than real aperture microscopes. SAM produces complex image data, enabling the use of coherent image processing techniques. Most importantly SAM produces the image data in a form that is easily manipulated by a digital image processing workstation.

Infrared identification of the Criegee intermediates syn- and anti-CH3CHOO, and their distinct conformation-dependent reactivity

PubMed Central

Lin, Hui-Yu; Huang, Yu-Hsuan; Wang, Xiaohong; Bowman, Joel M.; Nishimura, Yoshifumi; Witek, Henryk A.; Lee, Yuan-Pern

2015-01-01

The Criegee intermediates are carbonyl oxides that play critical roles in ozonolysis of alkenes in the atmosphere. So far, the mid-infrared spectrum of only the simplest Criegee intermediate CH2OO has been reported. Methyl substitution of CH2OO produces two conformers of CH3CHOO and consequently complicates the infrared spectrum. Here we report the transient infrared spectrum of syn- and anti-CH3CHOO, produced from CH3CHI + O2 in a flow reactor, using a step-scan Fourier-transform spectrometer. Guided and supported by high-level full-dimensional quantum calculations, rotational contours of the four observed bands are simulated successfully and provide definitive identification of both conformers. Furthermore, anti-CH3CHOO shows a reactivity greater than syn-CH3CHOO towards NO/NO2; at the later period of reaction, the spectrum can be simulated with only syn-CH3CHOO. Without NO/NO2, anti-CH3CHOO also decays much faster than syn-CH3CHOO. The direct infrared detection of syn- and anti-CH3CHOO should prove useful for field measurements and laboratory investigations of the Criegee mechanism. PMID:25959902

The Effect of Fragmented Pathogenic α-Synuclein Seeds on Prion-like Propagation*

PubMed Central

Tarutani, Airi; Suzuki, Genjiro; Shimozawa, Aki; Nonaka, Takashi; Akiyama, Haruhiko; Hisanaga, Shin-ichi; Hasegawa, Masato

2016-01-01

Aggregates of abnormal proteins are widely observed in neuronal and glial cells of patients with various neurodegenerative diseases, and it has been proposed that prion-like behavior of these proteins can account for not only the onset but also the progression of these diseases. However, it is not yet clear which abnormal protein structures function most efficiently as seeds for prion-like propagation. In this study, we aimed to identify the most pathogenic species of α-synuclein (α-syn), the main component of the Lewy bodies and Lewy neurites that are observed in α-synucleinopathies. We prepared various forms of α-syn protein and examined their seeding properties in vitro in cells and in mouse experimental models. We also characterized these α-syn species by means of electron microscopy and thioflavin fluorescence assays and found that fragmented β sheet-rich fibrous structures of α-syn with a length of 50 nm or less are the most efficient promoters of accumulation of phosphorylated α-syn, which is the hallmark of α-synucleinopathies. These results indicate that fragmented amyloid-like aggregates of short α-syn fibrils are the key pathogenic seeds that trigger prion-like conversion. PMID:27382062

Revision of the Gonioctena nivosa species-group (Coleoptera, Chrysomelidae, Chrysomelinae) in the Holarctic region, with descriptions of two new species

PubMed Central

Cho, Hee-Wook; Kippenberg, Horst; Borowiec, Lech

2016-01-01

Abstract The Gonioctena nivosa species-group of the genus Gonioctena Chevrolat, 1836 is defined and reviewed. It contains six species including two new to science: Gonioctena gracilicornis (Kraatz, 1879), Gonioctena nivosa (Suffrian, 1851), Gonioctena norvegica (Strand, 1936), Gonioctena springlovae (Bechyně, 1948), Gonioctena amurensis Cho & Borowiec, sp. n. and Gonioctena jani Cho & Borowiec, sp. n. Six new synonyms are proposed: Gonioctena nivosa (= Gonioctena arctica alberta Brown, 1952, syn. n., Phytodecta linnaeana bergrothi Jacobson, 1901, syn. n., Phytodecta linnaeanus var. mutatus Achard, 1924, syn. n., Phytodecta linnaeanus var. simplex Achard, 1924, syn. n. and Phytodecta nivosa var. cedehensis Ronchetti, 1922, syn. n.) and Gonioctena norvegica (= Gonioctena janovskii Medvedev, 1976, syn. n.). Phytodecta flavicornis var. limbatipennis Achard, 1924 and Phytodecta nivosa var. bicolor Heyden, 1883 are removed from synonymy with Gonioctena nivosa (Suffrian, 1851) and are synonymized with Gonioctena flavicornis (Suffrian, 1851). Distribution maps, a key to species, color variation, geographic variation of male genitalia and host plants are provided. Ovoviviparity is newly recorded in Gonioctena gracilicornis and Gonioctena nivosa. Lectotypes are designated for Gonioctena affinis, Gonioctena arctica, Gonioctena linnaeana bergrothi and Gonioctena nivosa. PMID:27408579

The lysosomal enzyme alpha-Galactosidase A is deficient in Parkinson's disease brain in association with the pathologic accumulation of alpha-synuclein.

PubMed

Nelson, Michael P; Boutin, Michel; Tse, Tonia E; Lu, Hailin; Haley, Emily D; Ouyang, Xiaosen; Zhang, Jianhua; Auray-Blais, Christiane; Shacka, John J

2018-02-01

The aberrant accumulation of alpha-synuclein (α-syn) is believed to contribute to the onset and pathogenesis of Parkinson's disease (PD). The autophagy-lysosome pathway (ALP) is responsible for the high capacity clearance of α-syn. ALP dysfunction is documented in PD and pre-clinical evidence suggests that inhibiting the ALP promotes the pathological accumulation of α-syn. We previously identified the pathological accumulation of α-syn in the brains of mice deficient for the soluble lysosomal enzyme alpha-Galactosidase A (α-Gal A), a member of the glycosphingolipid metabolism pathway. In the present study, we quantified α-Gal A activity and levels of its glycosphingolipid metabolites in postmortem temporal cortex specimens from control individuals and in PD individuals staged with respect to α-syn containing Lewy body pathology. In late-state PD temporal cortex we observed significant decreases in α-Gal A activity and the 46kDa "active" species of α-Gal A as determined respectively by fluorometric activity assay and western blot analysis. These decreases in α-Gal A activity/levels correlated significantly with increased α-syn phosphorylated at serine 129 (p129S-α-syn) that was maximal in late-stage PD temporal cortex. Mass spectrometric analysis of 29 different isoforms of globotriaosylceramide (Gb 3 ), a substrate of α-Gal A indicated no significant differences with respect to different stages of PD temporal cortex. However, significant correlations were observed between increased levels of several Gb 3 isoforms and with decreased α-Gal A activity and/or increased p129S-α-syn. Deacylated Gb 3 (globotriaosylsphingosine or lyso-Gb 3 ) was also analyzed in PD brain tissue but was below the limit of detection of 20pmol/g. Analysis of other lysosomal enzymes revealed a significant decrease in activity for the lysosomal aspartic acid protease cathepsin D but not for glucocerebrosidase (GCase) or cathepsin B in late-stage PD temporal cortex. However, a significant correlation was observed between decreasing GCase activity and increasing p129S-α-syn. Together our findings indicate α-Gal A deficiency in late-stage PD brain that correlates significantly with the pathological accumulation of α-syn, and further suggest the potential for α-Gal A and its glycosphingolipid substrates as putative biomarkers for PD. Copyright © 2017 Elsevier Inc. All rights reserved.

Dosimetric evaluation of magnetic resonance-generated synthetic CT for radiation treatment of rectal cancer.

PubMed

Wang, Hesheng; Du, Kevin; Qu, Juliet; Chandarana, Hersh; Das, Indra J

2018-01-01

The purpose of this study was to assess the dosimetric equivalence of magnetic resonance (MR)-generated synthetic CT (synCT) and simulation CT for treatment planning in radiotherapy of rectal cancer. This study was conducted on eleven patients who underwent whole-body PET/MR and PET/CT examination in a prospective IRB-approved study. For each patient synCT was generated from Dixon MR using a model-based method. Standard treatment planning directives were used to create a four-field box (4F), an oblique four-field (O4F) and a volumetric modulated arc therapy (VMAT) plan on synCT for treatment of rectal cancer. The plans were recalculated on CT with the same monitor units (MUs) as that of synCT. Dose-volume metrics of planning target volume (PTV) and organs at risk (OARs) as well as gamma analysis of dose distributions were evaluated to quantify the difference between synCT and CT plans. All plans were calculated using the analytical anisotropic algorithm (AAA). The VMAT plans on synCT and CT were also calculated using the Acuros XB algorithm for comparison with the AAA calculation. Medians of absolute differences in PTV metrics between synCT and CT plans were 0.2%, 0.2% and 0.3% for 4F, O4F and VMAT respectively. No significant differences were observed in OAR dose metrics including bladder V40Gy, mean dose in bladder, bowel V45Gy and femoral head V30Gy in any techniques. Gamma analysis with 2%/2mm dose difference/distance to agreement criteria showed median passing rates of 99.8% (range: 98.5 to 100%), 99.9% (97.2 to 100%), and 99.9% (99.4 to 100%) for 4F, O4F and VMAT, respectively. Using Acuros XB dose calculation, 2%/2mm gamma analysis generated a passing rate of 99.2% (97.7 to 99.9%) for VMAT plans. SynCT enabled dose calculation equivalent to conventional CT for treatment planning of 3D conformal treatment as well as VMAT of rectal cancer. The dosimetric agreement between synCT and CT calculated doses demonstrated the potential of MR-only treatment planning for rectal cancer using MR generated synCT.

Sensitivity analyses of exposure estimates from a quantitative job-exposure matrix (SYN-JEM) for use in community-based studies.

PubMed

Peters, Susan; Kromhout, Hans; Portengen, Lützen; Olsson, Ann; Kendzia, Benjamin; Vincent, Raymond; Savary, Barbara; Lavoué, Jérôme; Cavallo, Domenico; Cattaneo, Andrea; Mirabelli, Dario; Plato, Nils; Fevotte, Joelle; Pesch, Beate; Brüning, Thomas; Straif, Kurt; Vermeulen, Roel

2013-01-01

We describe the elaboration and sensitivity analyses of a quantitative job-exposure matrix (SYN-JEM) for respirable crystalline silica (RCS). The aim was to gain insight into the robustness of the SYN-JEM RCS estimates based on critical decisions taken in the elaboration process. SYN-JEM for RCS exposure consists of three axes (job, region, and year) based on estimates derived from a previously developed statistical model. To elaborate SYN-JEM, several decisions were taken: i.e. the application of (i) a single time trend; (ii) region-specific adjustments in RCS exposure; and (iii) a prior job-specific exposure level (by the semi-quantitative DOM-JEM), with an override of 0 mg/m(3) for jobs a priori defined as non-exposed. Furthermore, we assumed that exposure levels reached a ceiling in 1960 and remained constant prior to this date. We applied SYN-JEM to the occupational histories of subjects from a large international pooled community-based case-control study. Cumulative exposure levels derived with SYN-JEM were compared with those from alternative models, described by Pearson correlation ((Rp)) and differences in unit of exposure (mg/m(3)-year). Alternative models concerned changes in application of job- and region-specific estimates and exposure ceiling, and omitting the a priori exposure ranking. Cumulative exposure levels for the study subjects ranged from 0.01 to 60 mg/m(3)-years, with a median of 1.76 mg/m(3)-years. Exposure levels derived from SYN-JEM and alternative models were overall highly correlated (R(p) > 0.90), although somewhat lower when omitting the region estimate ((Rp) = 0.80) or not taking into account the assigned semi-quantitative exposure level (R(p) = 0.65). Modification of the time trend (i.e. exposure ceiling at 1950 or 1970, or assuming a decline before 1960) caused the largest changes in absolute exposure levels (26-33% difference), but without changing the relative ranking ((Rp) = 0.99). Exposure estimates derived from SYN-JEM appeared to be plausible compared with (historical) levels described in the literature. Decisions taken in the development of SYN-JEM did not critically change the cumulative exposure levels. The influence of region-specific estimates needs to be explored in future risk analyses.

Syn-collisional felsic magmatism and continental crust growth: A case study from the North Qilian Orogenic Belt at the northern margin of the Tibetan Plateau

NASA Astrophysics Data System (ADS)

Chen, Shuo; Niu, Yaoling; Xue, Qiqi

2018-05-01

The abundant syn-collisional granitoids produced and preserved at the northern Tibetan Plateau margin provide a prime case for studying the felsic magmatism as well as continental crust growth in response to continental collision. Here we present the results from a systematic study of the syn-collisional granitoids and their mafic magmatic enclaves (MMEs) in the Laohushan (LHS) and Machangshan (MCS) plutons from the North Qilian Orogenic Belt (NQOB). Two types of MMEs from the LHS pluton exhibit identical crystallization age ( 430 Ma) and bulk-rock isotopic compositions to their host granitoids, indicating their genetic link. The phase equilibrium constraints and pressure estimates for amphiboles from the LHS pluton together with the whole rock data suggest that the two types of MMEs represent two evolution products of the same hydrous andesitic magmas. In combination with the data on NQOB syn-collisional granitoids elsewhere, we suggest that the syn-collisional granitoids in the NQOB are material evidence of melting of ocean crust and sediment. The remarkable compositional similarity between the LHS granitoids and the model bulk continental crust in terms of major elements, trace elements, and some key element ratios indicates that the syn-collisional magmatism in the NQOB contributes to net continental crust growth, and that the way of continental crust growth in the Phanerozoic through syn-collisional felsic magmatism (production and preservation) is a straightforward process without the need of petrologically and physically complex processes.

The interplay of intrinsic disorder and macromolecular crowding on α-synuclein fibril formation

NASA Astrophysics Data System (ADS)

Shirai, Nobu C.; Kikuchi, Macoto

2016-02-01

α-synuclein (α-syn) is an intrinsically disordered protein which is considered to be one of the causes of Parkinson's disease. This protein forms amyloid fibrils when in a highly concentrated solution. The fibril formation of α-syn is induced not only by increases in α-syn concentration but also by macromolecular crowding. In order to investigate the coupled effect of the intrinsic disorder of α-syn and macromolecular crowding, we construct a lattice gas model of α-syn in contact with a crowding agent reservoir based on statistical mechanics. The main assumption is that α-syn can be expressed as coarse-grained particles with internal states coupled with effective volume; and disordered states are modeled by larger particles with larger internal entropy than other states. Thanks to the simplicity of the model, we can exactly calculate the number of conformations of crowding agents, and this enables us to prove that the original grand canonical ensemble with a crowding agent reservoir is mathematically equivalent to a canonical ensemble without crowding agents. In this expression, the effect of macromolecular crowding is absorbed in the internal entropy of disordered states; it is clearly shown that the crowding effect reduces the internal entropy. Based on Monte Carlo simulation, we provide scenarios of crowding-induced fibril formation. We also discuss the recent controversy over the existence of helically folded tetramers of α-syn, and suggest that macromolecular crowding is the key to resolving the controversy.

Common structural features of toxic intermediates from α-synuclein and GroES fibrillogenesis detected using cryogenic coherent X-ray diffraction imaging.

PubMed

Kameda, Hiroshi; Usugi, Sayaka; Kobayashi, Mana; Fukui, Naoya; Lee, Seki; Hongo, Kunihiro; Mizobata, Tomohiro; Sekiguchi, Yuki; Masaki, Yu; Kobayashi, Amane; Oroguchi, Tomotaka; Nakasako, Masayoshi; Takayama, Yuki; Yamamoto, Masaki; Kawata, Yasushi

2017-01-01

The aggregation and deposition of α-synuclein (αSyn) in neuronal cells is correlated to pathogenesis of Parkinson's disease. Although the mechanism of αSyn aggregation and fibril formation has been studied extensively, the structural hallmarks that are directly responsible for toxicity toward cells are still under debate. Here, we have compared the structural characteristics of the toxic intermediate molecular species of αSyn and similar toxic species of another protein, GroES, using coherent X-ray diffraction analysis. Using coherent X-ray free electron laser pulses of SACLA, we analysed αSyn and GroES fibril intermediate species and characterized various aggregate structures. Unlike previous studies where an annular oligomeric form of αSyn was identified, particle reconstruction from scattering traces suggested that the specific forms of the toxic particles were varied, with the sizes of the particles falling within a specific range. We did however discover a common structural feature in both αSyn and GroES samples; the edges of the detected particles were nearly parallel and produced a characteristic diffraction pattern in the diffraction experiments. The presence of parallel-edged particles in toxic intermediates of αSyn and GroES fibrillogenesis pointed towards a plausible common molecular interface that leads to the formation of mature fibrils. © The Authors 2016. Published by Oxford University Press on behalf of the Japanese Biochemical Society. All rights reserved.

Synthetic biology in the German press: how implications of metaphors shape representations of morality and responsibility.

PubMed

Döring, Martin

2018-06-24

Synthetic biology (SynBio) represents a relatively young field of research which has developed into an important scientific endeavour. Characterised by a high degree of interdisciplinary work crossing disciplinary boundaries, such as biology, mathematics and engineering, SynBio has been, since its beginning, devoted to creating new biological functions, metabolic pathways or even minimal organisms. Although its often-articulated aim of developing new forms of life has so far not been archived, SynBio nowadays represents a well-established biotechnological approach and it has also attracted public concern, especially since Craig Venter's work on Mycoplasma Mycoides JCVI-syn1.0. Taking these developments as a starting point, the paper empirically investigates the metaphorical representations of SynBio in two leading German media publications, the daily newspaper Die Frankfurter Allgemeine Zeitung and the weekly magazine Der Spiegel between 2000 and 2010. Using a novel combination of metaphor and co-occurrence analysis, the paper engages in a systematic examination of implicit moral implications inherent in linguistic images permeating this news coverage. It demonstrates a method of how media-metaphorical representations and their moral implications of SynBio could analytically be revealed and analysed. In doing so, it aims at contributing to empirical ethical analyses of the news coverage on SynBio in particular and offers an approach that methodologically adds to literature on responsible language use, which is emerging in science and technology studies and ethical analyses of new technologies.

Studies on complex π-π and T-stacking features of imidazole and phenyl/p-halophenyl units in series of 5-amino-1-(phenyl/p-halophenyl)imidazole-4-carboxamides and their carbonitrile derivatives: Role of halogens in tuning of conformation

NASA Astrophysics Data System (ADS)

Das, Aniruddha

2017-11-01

5-amino-1-(phenyl/p-halophenyl)imidazole-4-carboxamides (N-phenyl AICA) (2a-e) and 5-amino-1-(phenyl/p-halophenyl)imidazole-4-carbonitriles (N-phenyl AICN) (3a-e) had been synthesized. X-ray crystallographic studies of 2a-e and 3a-e had been performed to identify any distinct change in stacking patterns in their crystal lattice. Single crystal X-ray diffraction studies of 2a-e revealed π-π stack formations with both imidazole and phenyl/p-halophenyl units in anti and syn parallel-displaced (PD)-type dispositions. No π-π stacking of imidazole occurred when the halogen substituent is bromo or iodo; π-π stacking in these cases occurred involving phenyl rings only. The presence of an additional T-stacking had been observed in crystal lattices of 3a-e. Vertical π-π stacking distances in anti-parallel PD-type arrangements as well as T-stacking distances had shown stacking distances short enough to impart stabilization whereas syn-parallel stacking arrangements had got much larger π-π stacking distances to belie any syn-parallel stacking stabilization. DFT studies had been pursued for quantifying the π-π stacking and T-stacking stabilization. The plotted curves for anti-parallel and T-stacked moieties had similarities to the 'Morse potential energy curve for diatomic molecule'. The minima of the curves corresponded to the most stable stacking distances and related energy values indicated stacking stabilization. Similar DFT studies on syn-parallel systems of 2b corresponded to no π-π stacking stabilization at all. Halogen-halogen interactions had also been observed to stabilize the compounds 2d, 2e and 3d. Nano-structural behaviour of the series of compounds 2a-e and 3a-e were thoroughly investigated.

Using Affinity Chromatography to Investigate Novel Protein–Protein Interactions in an Undergraduate Cell and Molecular Biology Lab Course

PubMed Central

2009-01-01

Inquiry-driven lab exercises require students to think carefully about a question, carry out an investigation of that question, and critically analyze the results of their investigation. Here, we describe the implementation and assessment of an inquiry-based laboratory exercise in which students obtain and analyze novel data that contribute to our understanding of macromolecular trafficking between the nucleus and cytoplasm in eukaryotic cells. Although many of the proteins involved in nucleocytoplasmic transport are known, the physical interactions between some of these polypeptides remain uncharacterized. In this cell and molecular biology lab exercise, students investigate novel protein–protein interactions between factors involved in nuclear RNA export. Using recombinant protein expression, protein extraction, affinity chromatography, SDS-polyacrylamide gel electrophoresis, and Western blotting, undergraduates in a sophomore-level lab course identified a previously unreported association between the soluble mRNA transport factor Mex67 and the C-terminal region of the yeast nuclear pore complex protein Nup1. This exercise immersed students in the process of investigative science, from proposing and performing experiments through analyzing data and reporting outcomes. On completion of this investigative lab sequence, students reported enhanced understanding of the scientific process, increased proficiency with cellular and molecular methods and content, greater understanding of data analysis and the importance of appropriate controls, an enhanced ability to communicate science effectively, and an increased enthusiasm for scientific research and for the lab component of the course. The modular nature of this exercise and its focus on asking novel questions about protein–protein interactions make it easily transferable to undergraduate lab courses performed in a wide variety of contexts. PMID:19723816

Stereodivergent catalytic doubly diastereoselective nitroaldol reactions using heterobimetallic complexes.

PubMed

Sohtome, Yoshihiro; Kato, Yuko; Handa, Shinya; Aoyama, Naohiro; Nagawa, Keita; Matsunaga, Shigeki; Shibasaki, Masakatsu

2008-06-05

Stereodivergent construction of three contiguous stereocenters in catalytic doubly diastereoselective nitroaldol reactions of alpha-chiral aldehydes with nitroacetaldehyde dimethyl acetal using two types of heterobimetallic catalysts is described. A La-Li-BINOL (LLB) catalyst afforded anti,syn-nitroaldol products in >20:1-14:1 selectivity, and a Pd/La/Schiff base catalyst afforded complimentary syn,syn-nitroaldol products in 10:1-5:1 selectivity.

On the Quantitative Analysis of Liquid Flow in Physiological Tubes.

DTIC Science & Technology

1982-12-01

cri- copharyngeal sphincter which is aided by skeletal muscle (Vantrap- pen and hellemans, 1980) relaxes to accept the bolus and the gastro - esophageal ...lower ( gastro -) esophageal junction during peristalsis resulting from the interaction of gastric, esophageal and thoracic pressures. PIP is a pressure...higher than the downstream pressure and a flow velocity profile with no reflux (syn.: retropulsion). The 5 Pumping in Biological Tubes a. Peristaltic

A triple-bridged azido-Cu(II) chain compound fine-tuned by mixed carboxylate/ethanol linkers displays slow-relaxation and ferromagnetic order: synthesis, crystal structure, magnetic properties and DFT calculations.

PubMed

Liu, Xiangyu; Chen, Sanping; Grancha, Thais; Pardo, Emilio; Ke, Hongshan; Yin, Bing; Wei, Qing; Xie, Gang; Gao, Shengli

2014-11-07

A new azido-Cu(II) compound, [Cu(4-fba)(N3)(C2H5OH)] (4-fba = 4-fluorobenzoic acid) (1), has been synthesized and characterized. The X-ray crystal structure analysis demonstrates that only one crystallographically independent Cu(II) ion in the asymmetric unit of 1 exhibits a stretched octahedral geometry in which two azido N atoms and two carboxylic O atoms locate in the equatorial square, while two ethanol O atoms occupy the apical positions, forming a 1D Cu(II) chain with an alternating triple-bridge of EO-azido, syn,syn-carboxylate, and μ2-ethanol. The title compound consists of ferromagnetically interacting ferromagnetic chains, which exhibit ferromagnetic order (T(c) = 7.0 K). The strong ferromagnetic coupling between adjacent Cu(II) ions within each chain is due to the countercomplementarity of the super-exchange pathways, whereas the ferromagnetic interchain interactions--responsible for the long-range magnetic ordering--are most likely due to the presence of coordinated ethanol molecules establishing hydrogen bonds with neighboring chains. DFT calculations have been performed on compound 1 to offer a qualitative theoretical explanation of the magnetic behavior.

First experimental observations on melting and chemical modification of volcanic ash during lightning interaction.

PubMed

Mueller, S P; Helo, C; Keller, F; Taddeucci, J; Castro, J M

2018-01-23

Electrification in volcanic ash plumes often leads to syn-eruptive lightning discharges. High temperatures in and around lightning plasma channels have the potential to chemically alter, re-melt, and possibly volatilize ash fragments in the eruption cloud. In this study, we experimentally simulate temperature conditions of volcanic lightning in the laboratory, and systematically investigate the effects of rapid melting on the morphology and chemical composition of ash. Samples of different size and composition are ejected towards an artificially generated electrical arc. Post-experiment ash morphologies include fully melted spheres, partially melted particles, agglomerates, and vesiculated particles. High-speed imaging reveals various processes occurring during the short lightning-ash interactions, such as particle melting and rounding, foaming, and explosive particle fragmentation. Chemical analyses of the flash-melted particles reveal considerable bulk loss of Cl, S, P and Na through thermal vaporization. Element distribution patterns suggest convection as a key process of element transport from the interior of the melt droplet to rim where volatiles are lost. Modeling the degree of sodium loss delivers maximum melt temperatures between 3290 and 3490 K. Our results imply that natural lighting strikes may be an important agent of syn-eruptive morphological and chemical processing of volcanic ash.

Polypeptide Synthesis in Simian Virus 5-Infected Cells

PubMed Central

Peluso, Richard W.; Lamb, Robert A.; Choppin, Purnell W.

1977-01-01

Polypeptide synthesis in three different cell types infected with simian virus 5 has been examined using high-resolution polyacrylamide slab gel electrophoresis, and all of the known viral polypeptides have been identified above the host cell background. The polypeptides were synthesized in infected cells in unequal proportions, which are approximately the same as they are found in virions, suggesting that their relative rates of synthesis are controlled. The nucleocapsid polypeptide (NP) was the first to be detected in infected cells, and by 12 to 14 h the other virion structural polypeptides were identified, except for the polypeptides comprising the smaller glycoprotein (F). However, a glycosylated precursor (F0) with a molecular weight of 66,000 was found in each cell type, and pulse-chase experiments suggested that this precursor was cleaved to yield polypeptides F1 and F2. No other proteolytic processing was found. In addition to the structural polypeptides, the synthesis of five other polypeptides, designated I through V, has been observed in simian virus 5-infected cells. One of these (V), with a molecular weight of 24,000, was found in all cells examined and may be a nonstructural viral polypeptide. In contrast, there are polypeptides present in uninfected cells that correspond in size to polypeptides I through IV, and similar polypeptides have also been detected in increased amounts in cells infected with Sendai virus. These findings, and the fact that the synthesis of all four of these polypeptides is not increased in every cell type, suggest that they represent host polypeptides whose synthesis may be enhanced upon infection. When a high salt concentration was used to decrease host cell protein synthesis in infected cells, polypeptides IV and (to a lesser extent) I were synthesized in relatively greater amounts than other cellular polypeptides, as were the viral polypeptides. The possibility that these polypeptides may play some role in virus replication is discussed. Images PMID:196101

Dose-dependent interaction between gemfibrozil and repaglinide in humans: strong inhibition of CYP2C8 with subtherapeutic gemfibrozil doses.

PubMed

Honkalammi, Johanna; Niemi, Mikko; Neuvonen, Pertti J; Backman, Janne T

2011-10-01

Gemfibrozil 1-O-β-glucuronide inactivates CYP2C8 irreversibly. We investigated the effect of gemfibrozil dose on CYP2C8 activity in humans using repaglinide as a probe drug. In a randomized, five-phase crossover study, 10 healthy volunteers ingested 0.25 mg of repaglinide 1 h after different doses of gemfibrozil or placebo. Concentrations of plasma repaglinide, gemfibrozil, their metabolites, and blood glucose were measured. A single gemfibrozil dose of 30, 100, 300, and 900 mg increased the area under the concentration-time curve of repaglinide 1.8-, 4.5-, 6.7-, and 8.3-fold (P < 0.001), and its peak concentration 1.4-, 1.7-, 2.1-, and 2.4-fold (P < 0.05), compared with placebo, respectively. Gemfibrozil pharmacokinetics was characterized by a slightly more than dose-proportional increase in the area under the curve of gemfibrozil and its glucuronide. The gemfibrozil-repaglinide interaction could be mainly explained by gemfibrozil 1-O-β-glucuronide concentration-dependent, mechanism-based inhibition of CYP2C8, with a minor contribution by competitive inhibition of organic anion-transporting polypeptide 1B1 at the highest gemfibrozil dose. The findings are consistent with ∼50% inhibition of CYP2C8 already with a single 30-mg dose of gemfibrozil and >95% inhibition with 900 mg. In clinical drug-drug interaction studies, a single 900-mg dose of gemfibrozil can be used to achieve nearly complete inactivation of CYP2C8.

Interaction of coeval felsic and mafic magmas from the Kanker granite, Pithora region, Bastar Craton, Central India

NASA Astrophysics Data System (ADS)

Elangovan, R.; Krishna, Kumar; Vishwakarma, Neeraj; Hari, K. R.; Ram Mohan, M.

2017-10-01

Field and petrographic studies are carried out to characterize the interactions of mafic and felsic magmas from Pithora region of the northeastern part of the Bastar Craton. The MMEs, syn-plutonic mafic dykes, cuspate contacts, magmatic flow textures, mingling and hybridization suggest the coeval emplacement of end member magmas. Petrographic evidences such as disequilibrium assemblages, resorption textures, quartz ocelli, rapakivi and poikilitic textures suggest magma mingling and mixing phenomena. Such features of mingling and mixing of the felsic and mafic magma manifest the magma chamber processes. Introduction of mafic magmas into the felsic magmas before initiation of crystallization of the latter, results in hybrid magmas under the influence of thermal and chemical exchange. The mechanical exchange occurs between the coexisting magmas due to viscosity contrast, if the mafic magma enters slightly later into the magma chamber, then the felsic magma starts to crystallize. Blobs of mafic magma form as MMEs in the felsic magma and they scatter throughout the pluton due to convection. At a later stage, if mafic magma enters the system after partial crystallization of felsic phase, mechanical interaction between the magmas leads to the formation of fragmented dyke or syn-plutonic mafic dyke. All these features are well-documented in the study area. Field and petrographic evidences suggest that the textural variations from Pithora region of Bastar Craton are the outcome of magma mingling, mixing and hybridization processes.

Pulmonary inflammatory myofibroblastic tumor harboring EML4-ALK fusion gene.

PubMed

Sokai, Akihiko; Enaka, Makiko; Sokai, Risa; Mori, Shoichi; Mori, Shunsuke; Gunji, Masaharu; Fujino, Masahiko; Ito, Masafumi

2014-01-01

Inflammatory myofibroblastic tumor is a rare tumor deriving from mesenchymal tissue. Approximately 50% of inflammatory myofibroblastic tumors harbor an anaplastic lymphoma kinase fusion gene. Pulmonary inflammatory myofibroblastic tumors harboring tropomyosin3-anaplastic lymphoma kinase or protein tyrosine phosphatase receptor-type F polypeptide-interacting protein-binding protein 1-anaplastic lymphoma kinase have been reported previously. However, it has not been reported that inflammatory myofibroblastic tumors harbor echinoderm microtubule-associated protein-like 4-anaplastic lymphoma kinase fusion gene which is considered to be very specific to lung cancers. A few tumors harboring echinoderm microtubule-associated protein-like 4-anaplastic lymphoma kinase fusion gene other than lung cancers have been reported and the tumors were all carcinomas. A 67-year-old man had been followed up for a benign tumor for approximately 3 years before the tumor demonstrated malignant transformation. Lobectomy and autopsy revealed that an inflammatory myofibroblastic tumor harboring echinoderm microtubule-associated protein-like 4-anaplastic lymphoma kinase fusion gene had transformed into an undifferentiated sarcoma. This case suggests that echinoderm microtubule-associated protein-like 4-anaplastic lymphoma kinase fusion is an oncogenic event in not only carcinomas but also sarcomas originating from stromal cells.

Tristetraprolin inhibits mitochondrial function through suppression of α-Synuclein expression in cancer cells

PubMed Central

Vo, Mai-Tram; Choi, Seong Hee; Lee, Ji-Heon; Hong, Chung Hwan; Kim, Jong Soo; Lee, Unn Hwa; Chung, Hyung-Min; Lee, Byung Ju; Park, Jeong Woo; Cho, Wha Ja

2017-01-01

Mitochondrial dynamics play critical roles in maintaining mitochondrial functions. Here, we report a novel mechanism for regulation of mitochondrial dynamics mediated by tristetraprolin (TTP), an AU-rich element (ARE)-binding protein. Overexpression of TTP resulted in elongated mitochondria, down-regulation of mitochondrial oxidative phosphorylation, reduced membrane potential, cytochrome c release, and increased apoptotic cell death in cancer cells. TTP overexpression inhibited the expression of α-Synuclein (α-Syn). TTP bound to the ARE within the mRNA 3′-untranslated regions (3′-UTRs) of α-Syn and enhanced the decay of α-Syn mRNA. Overexpression of α-Syn without the 3′-UTR restored TTP-induced defects in mitochondrial morphology, mitochondrial oxidative phosphorylation, membrane potential, and apoptotic cell death. Taken together, our data demonstrate that TTP acts as a regulator of mitochondrial dynamics through enhancing degradation of α-Syn mRNA in cancer cells. This finding will increase understanding of the molecular basis of mitochondrial dynamics. PMID:28410208

Dioleoyl-phosphatidic acid selectively binds to α-synuclein and strongly induces its aggregation.

PubMed

Mizuno, Satoru; Sasai, Hirotaka; Kume, Aiko; Takahashi, Daisuke; Satoh, Mamoru; Kado, Sayaka; Sakane, Fumio

2017-03-01

α-Synuclein (α-syn), which causally links to Parkinson's disease, binds to vesicles containing phosphatidic acid (PA). However, the effects of the fatty acyl chains of PA on its ability to bind to α-syn protein remain unclear. Intriguingly, we reveal that among several PA species, 18:1/18:1-PA is the most strongly bound PA to the α-syn protein. Moreover, 18:1/18:1-PA more strongly enhances secondary structural changes from the random coil form to the α-helical form than 16:0/18:1-PA. Furthermore, 18:1/18:1-PA more markedly accelerates generation of multimeric and proteinase K-resistant α-syn protein compared to 16:0/18:1-PA. These results indicate that among phospholipids examined so far, 18:1/18:1-PA demonstrates the strongest binding to α-syn, as well as the most effective enhancement of its secondary structural changes and aggregation formation. © 2017 Federation of European Biochemical Societies.

DNA damage preceding dopamine neuron degeneration in A53T human α-synuclein transgenic mice.

PubMed

Wang, Degui; Yu, Tianyu; Liu, Yongqiang; Yan, Jun; Guo, Yingli; Jing, Yuhong; Yang, Xuguang; Song, Yanfeng; Tian, Yingxia

2016-12-02

Defective DNA repair has been linked with age-associated neurodegenerative disorders. Parkinson's disease (PD) is a progressive neurodegenerative disorder caused by genetic and environmental factors. Whether damages to nuclear DNA contribute to neurodegeneration of PD still remain obscure. in this study we aim to explore whether nuclear DNA damage induce dopamine neuron degeneration in A53T human α-Synuclein over expressed mouse model. We investigated the effects of X-ray irradiation on A53T-α-Syn MEFs and A53T-α-Syn transgene mice. Our results indicate that A53T-α-Syn MEFs show a prolonged DNA damage repair process and senescense phenotype. DNA damage preceded onset of motor phenotype in A53T-α-Syn transgenic mice and decrease the number of nigrostriatal dopaminergic neurons. Neurons of A53T-α-Syn transgenic mice are more fragile to DNA damages. Copyright © 2016 Elsevier Inc. All rights reserved.

Basic science breaks through: New therapeutic advances in Parkinson's disease.

PubMed

Brundin, Patrik; Atkin, Graham; Lamberts, Jennifer T

2015-09-15

Parkinson's disease (PD) is the second most common neurodegenerative disease and is typically associated with progressive motor dysfunction, although PD patients also exhibit a variety of non-motor symptoms. The neuropathological hallmark of PD is intraneuronal inclusions containing primarily α-Synuclein (α-Syn), and several lines of evidence point to α-Syn as a key contributor to disease progression. Thus, basic research in the field of PD is largely focused on understanding the pathogenic properties of α-Syn. Over the past 2 y, these studies helped to identify several novel therapeutic strategies that have the potential to slow PD progression; such strategies include sequestration of extracellular α-Syn through immunotherapy, reduction of α-Syn multimerization or intracellular toxicity, and attenuation of the neuroinflammatory response. This review describes these and other putative therapeutic strategies, together with the basic science research that led to their identification. The current breadth of novel targets for the treatment of PD warrants cautious optimism in the fight against this devastating disease. © 2015 International Parkinson and Movement Disorder Society.

Nonviral gene editing via CRISPR/Cas9 delivery by membrane-disruptive and endosomolytic helical polypeptide.

PubMed

Wang, Hong-Xia; Song, Ziyuan; Lao, Yeh-Hsing; Xu, Xin; Gong, Jing; Cheng, Du; Chakraborty, Syandan; Park, Ji Sun; Li, Mingqiang; Huang, Dantong; Yin, Lichen; Cheng, Jianjun; Leong, Kam W

2018-05-08

Effective and safe delivery of the CRISPR/Cas9 gene-editing elements remains a challenge. Here we report the development of PEGylated nanoparticles (named P-HNPs) based on the cationic α-helical polypeptide poly(γ-4-((2-(piperidin-1-yl)ethyl)aminomethyl)benzyl-l-glutamate) for the delivery of Cas9 expression plasmid and sgRNA to various cell types and gene-editing scenarios. The cell-penetrating α-helical polypeptide enhanced cellular uptake and promoted escape of pCas9 and/or sgRNA from the endosome and transport into the nucleus. The colloidally stable P-HNPs achieved a Cas9 transfection efficiency up to 60% and sgRNA uptake efficiency of 67.4%, representing an improvement over existing polycation-based gene delivery systems. After performing single or multiplex gene editing with an efficiency up to 47.3% in vitro, we demonstrated that P-HNPs delivering Cas9 plasmid/sgRNA targeting the polo-like kinase 1 (Plk1) gene achieved 35% gene deletion in HeLa tumor tissue to reduce the Plk1 protein level by 66.7%, thereby suppressing the tumor growth by >71% and prolonging the animal survival rate to 60% within 60 days. Capable of delivering Cas9 plasmids to various cell types to achieve multiplex gene knock-out, gene knock-in, and gene activation in vitro and in vivo, the P-HNP system offers a versatile gene-editing platform for biological research and therapeutic applications. Copyright © 2018 the Author(s). Published by PNAS.

Translocation of cell-penetrating peptides into Candida fungal pathogens.

PubMed

Gong, Zifan; Karlsson, Amy J

2017-09-01

Cell-penetrating peptides (CPPs) are small peptides capable of crossing cellular membranes while carrying molecular cargo. Although they have been widely studied for their ability to translocate nucleic acids, small molecules, and proteins into mammalian cells, studies of their interaction with fungal cells are limited. In this work, we evaluated the translocation of eleven fluorescently labeled peptides into the important human fungal pathogens Candida albicans and C. glabrata and explored the mechanisms of translocation. Seven of these peptides (cecropin B, penetratin, pVEC, MAP, SynB, (KFF) 3 K, and MPG) exhibited substantial translocation (>80% of cells) into both species in a concentration-dependent manner, and an additional peptide (TP-10) exhibiting strong translocation into only C. glabrata. Vacuoles were involved in translocation and intracellular trafficking of the peptides in the fungal cells and, for some peptides, escape from the vacuoles and localization in the cytosol were correlated to toxicity toward the fungal cells. Endocytosis was involved in the translocation of cecropin B, MAP, SynB, MPG, (KFF) 3 K, and TP-10, and cecropin B, penetratin, pVEC, and MAP caused membrane permeabilization during translocation. These results indicate the involvement of multiple translocation mechanisms for some CPPs. Although high levels of translocation were typically associated with toxicity of the peptides toward the fungal cells, SynB was translocated efficiently into Candida cells at concentrations that led to minimal toxicity. Our work highlights the potential of CPPs in delivering antifungal molecules and other bioactive cargo to Candida pathogens. © 2017 The Protein Society.

Alpha-synuclein is present in dental calculus but not altered in Parkinson's disease patients in comparison to controls.

PubMed

Schmid, Sabrina; Goldberg-Bockhorn, Eva; Schwarz, Silke; Rotter, Nicole; Kassubek, Jan; Del Tredici, Kelly; Pinkhardt, Elmar; Otto, Markus; Ludolph, Albert C; Oeckl, Patrick

2018-06-01

In autopsy cases staged for sporadic Parkinson's disease (PD), the neuropathology is characterized by a preclinical phase that targets the enteric nervous system of the gastrointestinal tract (GIT). Therefore, the ENS might be a source of potential (presymptomatic) PD biomarkers. In this clinically based study, we examined the alpha-synuclein (αSyn) concentration in an easily accessible protein storage medium of the GIT, dental calculus, in 21/50 patients with PD and 28/50 age- and gender-matched controls using ELISA. αSyn was detectable in dental calculus and the median concentration in the control patients was 8.6 pg/mg calculus (interquartile range 2.6-13.1 pg/mg). αSyn concentrations were significantly influenced by blood contamination and samples with a hemoglobin concentration of > 4000 ng/mL were excluded. There was no significant difference of αSyn concentrations in the dental calculus of PD patients (5.76 pg/mg, interquartile range 2.91-9.74 pg/mg) compared to those in controls (p = 0.40). The total αSyn concentration in dental calculus is not a suitable biomarker for sporadic PD. Disease-related variants such as oligomeric or phosphorylated αSyn in calculus might prove to be more specific.

Site-specific structural dynamics of α-Synuclein revealed by time-resolved fluorescence spectroscopy: a review

NASA Astrophysics Data System (ADS)

Sahay, Shruti; Krishnamoorthy, G.; Maji, Samir K.

2016-12-01

Aggregation of α-Synuclein (α-Syn) into amyloid fibrils is known to be associated with the pathogenesis of Parkinson’s disease (PD). Several missense mutations of the α-Syn gene have been associated with rare, early onset familial forms of PD. Despite several studies done so far, the local/residue-level structure and dynamics of α-Syn in its soluble and aggregated fibril form and how these are affected by the familial PD associated mutations are still not clearly understood. Here, we review studies performed by our group as well as other research groups, where time-resolved fluorescence spectroscopy has been used to understand the site-specific structure and dynamics of α-Syn under physiological conditions as well as under conditions that alter the aggregation properties of the protein such as low pH, high temperature, presence of membrane mimics and familial PD associated mutations. These studies have provided important insights into the critical structural properties of α-Syn that may govern its aggregation. The review also highlights time-resolved fluorescence as a promising tool to study the critical conformational transitions associated with early oligomerization of α-Syn, which are otherwise not accessible using other commonly used techniques such as thioflavin T (ThT) binding assay.

Stereoselective bioaccumulation of syn- and anti-Dechlorane plus isomers in different tissues of common carp (Cyprinus carpio).

PubMed

Tang, Bin; Luo, Xiao-Jun; Huang, Chen-Chen; Sun, Run-Xia; Wang, Tao; Zeng, Yan-Hong; Mai, Bi-Xian

2018-03-01

Common carps (Cyprinus carpio) were exposed to syn- and anti-Dechlorane Plus (DP) isomers to investigate absorption, tissue distribution, and stereoselective bioaccumulation of DP isomers. The absorption efficiencies of anti-DP in the gastrointestinal system were higher than those of syn-DP. A linear accumulation was found for both isomers in all fish tissues except for serum; and the liver and gill exhibited the highest and lowest DP assimilation efficiency, respectively. The elimination of DP isomers in all tissues followed first-order kinetics, with the fastest depuration rate occurring in the liver and serum. The biomagnification factors (BMFs) of both isomers were less than one in all tissues, except for serum. Anti-DP was preferably accumulated in the liver, gill, and serum, whereas syn-DP was selectively accumulated in the carcass and gastrointestinal tract. As a whole, fish did not show selective accumulation of the syn- or anti-DP isomer in the uptake stage, whereas a selective accumulation of syn-DP in fish was observed during the depuration period, which could be due to a selective excretion of anti-DP. Metabolism cannot be ruled out as a possible reason considering the high f anti values and the high elimination rate of DPs in the liver. Copyright © 2017 Elsevier B.V. All rights reserved.

Synfograms: a new generation of holographic applications

NASA Astrophysics Data System (ADS)

Meulien Öhlmann, Odile; Öhlmann, Dietmar; Zacharovas, Stanislovas J.

2008-04-01

The new synthetic Four-dimensional printing technique (Syn4D) Synfogram is introducing time (animation) into spatial configuration of the imprinted three-dimensional shapes. While lenticular solutions offer 2 to 9 stereoscopic images Syn4D offers large format, full colors true 3D visualization printing of 300 to 2500 frames imprinted as holographic dots. This past 2 years Syn4D high-resolution displays proved to be extremely efficient for museums presentation, engineering design, automobile prototyping, and advertising virtual presentation as well as, for portrait and fashion applications. The main advantages of syn4D is that it offers a very easy way of using a variety of digital media, like most of 3D Modelling programs, 3D scan system, video sequences, digital photography, tomography as well as the Syn4D camera track system for life recording of spatial scenes changing in time. The use of digital holographic printer in conjunction with Syn4D image acquiring and processing devices separates printing and imaging creation in such a way that makes four-dimensional printing similar to a conventional digital photography processes where imaging and printing are usually separated in space and time. Besides making content easy to prepare, Syn4D has also developed new display and lighting solutions for trade show, museum, POP, merchandising, etc. The introduction of Synfograms is opening new applications for real life and virtual 4D displays. In this paper we will analyse the 3D market, the properties of the Synfograms and specific applications, the problems we encounter, solutions we find, discuss about customers demand and need for new product development.

Neural control of blood flow during exercise in human metabolic syndrome.

PubMed

Limberg, Jacqueline K; Morgan, Barbara J; Sebranek, Joshua J; Proctor, Lester T; Eldridge, Marlowe W; Schrage, William G

2014-09-01

α-Adrenergic-mediated vasoconstriction is greater during simulated exercise in animal models of metabolic syndrome (MetSyn) when compared with control animals. In an attempt to translate such findings to humans, we hypothesized that adults with MetSyn (n = 14, 35 ± 3 years old) would exhibit greater α-adrenergic responsiveness during exercise when compared with age-matched healthy control subjects (n = 16, 31 ± 3 years old). We measured muscle sympathetic nerve activity (MSNA; microneurography) and forearm blood flow (Doppler ultrasound) during dynamic forearm exercise (15% of maximal voluntary contraction). α-Adrenergic agonists (phenylephrine and clonidine) and an antagonist (phentolamine) were infused intra-arterially to assess α-adrenergic receptor responsiveness and restraint, respectively. Resting MSNA was ∼35% higher in adults with MetSyn (P < 0.05), but did not change in either group with dynamic exercise. Clonidine-mediated vasoconstriction was greater in adults with MetSyn (P < 0.01). Group differences in vascular responses to phenylephrine and phentolamine were not detected (P > 0.05). Interestingly, exercise-mediated vasodilatation was greater in MetSyn (P < 0.05). Adults with MetSyn exhibit greater resting MSNA and clonidine-mediated vasoconstriction, yet preserved functional sympatholysis and higher exercise blood flow during low-intensity hand-grip exercise when compared with age-matched healthy control subjects. These results suggest that adults with MetSyn exhibit compensatory vascular control mechanisms capable of preserving blood flow responses to exercise in the face of augmented sympathetic adrenergic activity. © 2014 The Authors. Experimental Physiology © 2014 The Physiological Society.

Ethylene Regulates the Physiology of the Cyanobacterium Synechocystis sp. PCC 6803 via an Ethylene Receptor.

PubMed

Lacey, Randy F; Binder, Brad M

2016-08-01

Ethylene is a plant hormone that plays a crucial role in the growth and development of plants. The ethylene receptors in plants are well studied, and it is generally assumed that they are found only in plants. In a search of sequenced genomes, we found that many bacterial species contain putative ethylene receptors. Plants acquired many proteins from cyanobacteria as a result of the endosymbiotic event that led to chloroplasts. We provide data that the cyanobacterium Synechocystis (Synechocystis sp. PCC 6803) has a functional receptor for ethylene, Synechocystis Ethylene Response1 (SynEtr1). We first show that SynEtr1 directly binds ethylene. Second, we demonstrate that application of ethylene to Synechocystis cells or disruption of the SynEtr1 gene affects several processes, including phototaxis, type IV pilus biosynthesis, photosystem II levels, biofilm formation, and spontaneous cell sedimentation. Our data suggest a model where SynEtr1 inhibits downstream signaling and ethylene inhibits SynEtr1. This is similar to the inverse-agonist model of ethylene receptor signaling proposed for plants and suggests a conservation of structure and function that possibly originated over 1 billion years ago. Prior research showed that SynEtr1 also contains a light-responsive phytochrome-like domain. Thus, SynEtr1 is a bifunctional receptor that mediates responses to both light and ethylene. To our knowledge, this is the first demonstration of a functional ethylene receptor in a nonplant species and suggests that that the perception of ethylene is more widespread than previously thought. © 2016 American Society of Plant Biologists. All Rights Reserved.

Ethylene Regulates the Physiology of the Cyanobacterium Synechocystis sp. PCC 6803 via an Ethylene Receptor1[OPEN

PubMed Central

2016-01-01

Ethylene is a plant hormone that plays a crucial role in the growth and development of plants. The ethylene receptors in plants are well studied, and it is generally assumed that they are found only in plants. In a search of sequenced genomes, we found that many bacterial species contain putative ethylene receptors. Plants acquired many proteins from cyanobacteria as a result of the endosymbiotic event that led to chloroplasts. We provide data that the cyanobacterium Synechocystis (Synechocystis sp. PCC 6803) has a functional receptor for ethylene, Synechocystis Ethylene Response1 (SynEtr1). We first show that SynEtr1 directly binds ethylene. Second, we demonstrate that application of ethylene to Synechocystis cells or disruption of the SynEtr1 gene affects several processes, including phototaxis, type IV pilus biosynthesis, photosystem II levels, biofilm formation, and spontaneous cell sedimentation. Our data suggest a model where SynEtr1 inhibits downstream signaling and ethylene inhibits SynEtr1. This is similar to the inverse-agonist model of ethylene receptor signaling proposed for plants and suggests a conservation of structure and function that possibly originated over 1 billion years ago. Prior research showed that SynEtr1 also contains a light-responsive phytochrome-like domain. Thus, SynEtr1 is a bifunctional receptor that mediates responses to both light and ethylene. To our knowledge, this is the first demonstration of a functional ethylene receptor in a nonplant species and suggests that that the perception of ethylene is more widespread than previously thought. PMID:27246094

Conformational Equilibria in Monomeric α-Synuclein at the Single-Molecule Level

PubMed Central

Tessari, Isabella; Mammi, Stefano; Bergantino, Elisabetta; Musiani, Francesco; Brucale, Marco; Bubacco, Luigi; Samorì, Bruno

2008-01-01

Human α-Synuclein (αSyn) is a natively unfolded protein whose aggregation into amyloid fibrils is involved in the pathology of Parkinson disease. A full comprehension of the structure and dynamics of early intermediates leading to the aggregated states is an unsolved problem of essential importance to researchers attempting to decipher the molecular mechanisms of αSyn aggregation and formation of fibrils. Traditional bulk techniques used so far to solve this problem point to a direct correlation between αSyn's unique conformational properties and its propensity to aggregate, but these techniques can only provide ensemble-averaged information for monomers and oligomers alike. They therefore cannot characterize the full complexity of the conformational equilibria that trigger the aggregation process. We applied atomic force microscopy–based single-molecule mechanical unfolding methodology to study the conformational equilibrium of human wild-type and mutant αSyn. The conformational heterogeneity of monomeric αSyn was characterized at the single-molecule level. Three main classes of conformations, including disordered and “β-like” structures, were directly observed and quantified without any interference from oligomeric soluble forms. The relative abundance of the “β-like” structures significantly increased in different conditions promoting the aggregation of αSyn: the presence of Cu2+, the pathogenic A30P mutation, and high ionic strength. This methodology can explore the full conformational space of a protein at the single-molecule level, detecting even poorly populated conformers and measuring their distribution in a variety of biologically important conditions. To the best of our knowledge, we present for the first time evidence of a conformational equilibrium that controls the population of a specific class of monomeric αSyn conformers, positively correlated with conditions known to promote the formation of aggregates. A new tool is thus made available to test directly the influence of mutations and pharmacological strategies on the conformational equilibrium of monomeric αSyn. PMID:18198943

Afrotropical flea beetle genera: a key to their identification, updated catalogue and biogeographical analysis (Coleoptera, Chrysomelidae, Galerucinae, Alticini)

PubMed Central

Biondi, Maurizio; D’Alessandro, Paola

2012-01-01

Abstract A revision of the Alticini genera from the Afrotropical region is reported. The paper includes the following for the flea beetle fauna occurring in Sub-Saharan Africa and Madagascar: a key to their identification; habitus photos of all the genera; microscope and scanning electron micrographs of many diagnostic morphological characters; and an updated annotated catalogue with biogeographical notes that include new distributional data. The following new synonymies are proposed: Aphthona Chevrolat, 1836 = Ethiopia Scherer, 1972 syn. n.; Sanckia Duvivier, 1891 = Eugonotes Jacoby, 1897 syn. n.; Eurylegna Weise, 1910a = Eurylegniella Scherer, 1972 syn. n.; Kimongona Bechyné, 1959a = Mesocrepis Scherer, 1963 syn. n.; Diphaulacosoma Jacoby, 1892a = Neoderina Bechyné, 1952 syn. n.; Sesquiphaera Bechyné, 1958a = Paropsiderma Bechyné, 1958a syn. n.; Podagrica Chevrolat, 1836 = Podagricina Csiki in Heikertinger and Csiki 1940 syn. n.; Amphimela Chapuis, 1875 = Sphaerophysa Baly, 1876a syn. n. The following new combinations are proposed: Blepharida insignis Brancsik, 1897 = Xanthophysca insignis (Brancsik, 1897) comb. n.; Blepharida multiguttata Duvivier, 1891 = Xanthophysca multiguttata (Duvivier, 1891) comb. n.; Hemipyxis balyana (Csiki in Heikertinger and Csiki 1940) = Pseudadorium balyanum (Csiki in Heikertinger and Csiki, 1940) comb. n.; Hemipyxis brevicornis (Jacoby, 1892a) = Pseudadorium brevicornis (Jacoby, 1892a) comb. n.; Hemipyxis cyanea (Weise, 1910b) = Pseudadorium cyaneum (Weise, 1910b) comb. n.; Hemipyxis gynandromorpha Bechyné, 1958c = Pseudadorium gynandromorphum (Bechyné, 1958c) comb. n.; Hemipyxis latiuscula Bechyné, 1958c = Pseudadorium latiusculum (Bechyné, 1958c) comb. n.; Hemipyxis soror (Weise, 1910b) = Pseudadorium soror (Weise, 1910b) comb. n. The genera Buphonella Jacoby, 1903aand Halticopsis Fairmaire, 1883a are transferred to the tribe Galerucini; the genus Biodontocnema Biondi, 2000 stat. prom. is considered to be valid and reinstated at generic level. Finally, a zoogeographical analysis of the flea beetle fauna in the Afrotropical region is provided. PMID:23378812

FTY720/Fingolimod Reduces Synucleinopathy and Improves Gut Motility in A53T Mice: CONTRIBUTIONS OF PRO-BRAIN-DERIVED NEUROTROPHIC FACTOR (PRO-BDNF) AND MATURE BDNF.

PubMed

Vidal-Martínez, Guadalupe; Vargas-Medrano, Javier; Gil-Tommee, Carolina; Medina, David; Garza, Nathan T; Yang, Barbara; Segura-Ulate, Ismael; Dominguez, Samantha J; Perez, Ruth G

2016-09-23

Patients with Parkinson's disease (PD) often have aggregated α-synuclein (aSyn) in enteric nervous system (ENS) neurons, which may be associated with the development of constipation. This occurs well before the onset of classic PD motor symptoms. We previously found that aging A53T transgenic (Tg) mice closely model PD-like ENS aSyn pathology, making them appropriate for testing potential PD therapies. Here we show that Tg mice overexpressing mutant human aSyn develop ENS pathology by 4 months. We then evaluated the responses of Tg mice and their WT littermates to the Food and Drug Administration-approved drug FTY720 (fingolimod, Gilenya) or vehicle control solution from 5 months of age. Long term oral FTY720 in Tg mice reduced ENS aSyn aggregation and constipation, enhanced gut motility, and increased levels of brain-derived neurotrophic factor (BDNF) but produced no significant change in WT littermates. A role for BDNF was directly assessed in a cohort of young A53T mice given vehicle, FTY720, the Trk-B receptor inhibitor ANA-12, or FTY720 + ANA-12 from 1 to 4 months of age. ANA-12-treated Tg mice developed more gut aSyn aggregation as well as constipation, whereas FTY720-treated Tg mice had reduced aSyn aggregation and less constipation, occurring in part by increasing both pro-BDNF and mature BDNF levels. The data from young and old Tg mice revealed FTY720-associated neuroprotection and reduced aSyn pathology, suggesting that FTY720 may also benefit PD patients and others with synucleinopathy. Another finding was a loss of tyrosine hydroxylase immunoreactivity in gut neurons with aggregated aSyn, comparable with our prior findings in the CNS. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Monoclonal antibodies to the light-harvesting chlorophyll a/b protein complex of photosystem II

PubMed Central

1986-01-01

A collection of 17 monoclonal antibodies elicited against the light- harvesting chlorophyll a/b protein complex which serves photosystem II (LHC-II) of Pisum sativum shows six classes of binding specificity. Antibodies of two of the classes recognize a single polypeptide (the 28- or the 26- kD polypeptides), thereby suggesting that the two proteins are not derived from a common precursor. Other classes of antibodies cross-react with several polypeptides of LHC-II or with polypeptides of both LHC-II and the light-harvesting chlorophyll a/b polypeptides of photosystem I (LHC-I), indicating that there are structural similarities among the polypeptides of LHC-II and LHC-I. The evidence for protein processing by which the 26-, 25.5-, and 24.5-kD polypeptides are derived from a common precursor polypeptide is discussed. Binding studies using antibodies specific for individual LHC- II polypeptides were used to quantify the number of antigenic polypeptides in the thylakoid membrane. 27 copies of the 26-kD polypeptide and two copies of the 28-kD polypeptide were found per 400 chlorophylls. In the chlorina f2 mutant of barley, and in intermittent light-treated barley seedlings, the amount of the 26-kD polypeptide in the thylakoid membranes was greatly reduced, while the amount of 28-kD polypeptide was apparently not affected. We propose that stable insertion and assembly of the 28-kD polypeptide, unlike the 26-kD polypeptide, is not regulated by the presence of chlorophyll b. PMID:3528171

Non-covalent S···O interactions control conformation in a scaffold that disrupts islet amyloid polypeptide fibrillation† †Electronic supplementary information (ESI) available: Experimental procedures; 1H, 13C and NOESY spectra; HRMS, and IR values. CCDC 1453550 (24), ThT assay data, computation, electron microscopy. For ESI and crystallographic data in CIF or other electronic format see DOI: 10.1039/c6sc00756b Click here for additional data file.

PubMed Central

Peacock, Hayden; Luo, Jinghui; Yamashita, Tohru; Luccarelli, James

2016-01-01

Conformationally-constrained molecules that selectively recognise the surfaces of proteins have the potential to direct the path of protein folding. Such molecules are of therapeutic interest because the misfolding of proteins, especially that which results in fibrillation and aggregation, is strongly correlated with numerous diseases. Here we report the novel use of S···O interactions as a conformational control element in a new class of non-peptidic scaffold that mimics key elements of protein surfaces. These molecules disrupt the fibrillation of islet amyloid polypeptide (IAPP), a process that is implicated in the pathology of type II diabetes. PMID:28451100

Solvent history dependence of gramicidin A conformations in hydrated lipid bilayers.

PubMed Central

LoGrasso, P V; Moll, F; Cross, T A

1988-01-01

Reconstituted lipid bilayers of dimyristoylphosphatidylcholine (DMPC) and gramicidin A' have been prepared by cosolubilizing gramicidin and DMPC in one of three organic solvent systems followed by vacuum drying and hydration. The conformational state of gramicidin as characterized by 23Na NMR, circular dichroism, and solid state 15N NMR is dependent upon the cosolubilizing solvent system. In particular, two conformational states are described; a state in which Na+ has minimal interactions with the polypeptide, referred to as a nonchannel state, and a state in which Na+ interacts very strongly with the polypeptide, referred to as the channel state. Both of these conformations are intimately associated with the hydrophobic core of the lipid bilayer. Furthermore, both of these states are stable in the bilayer at neutral pH and at a temperature above the bilayer phase transition temperature. These results with gramicidin suggest that the conformation of membrane proteins may be dictated by the conformation before membrane insertion and may be dependent upon the mechanism by which the insertion is accomplished. PMID:2462923

Fingerprinting of near-homogeneous DNA ligase I and II from human cells. Similarity of their AMP-binding domains.

PubMed

Yang, S W; Becker, F F; Chan, J Y

1990-10-25

DNA ligases play obligatory roles during replication, repair, and recombination. Multiple forms of DNA ligase have been reported in mammalian cells including DNA ligase I, the high molecular mass species which functions during replication, and DNA ligase II, the low molecular mass species which is associated with repair. In addition, alterations in DNA ligase activities have been reported in acute lymphocytic leukemia cells, Bloom's syndrome cells, and cells undergoing differentiation and development. To better distinguish the biochemical and molecular properties of the various DNA ligases from human cells, we have developed a method of purifying multiple species of DNA ligase from HeLa cells by chromatography through DEAE-Bio-Gel, CM-Bio-Gel, hydroxylapatite, Sephacryl S-300, Mono P, and DNA-cellulose. DNA-cellulose chromatography of the partially purified enzymes resolved multiple species of DNA ligase after labeling the enzyme with [alpha-32P]ATP to form the ligase-[32P]AMP adduct. The early eluting enzyme activity (0.25 M NaCl) contained a major 67-kDa-labeled protein, while the late eluting activity (0.48 M NaCl) contained two major labeled proteins of 90 and 78 kDa. Neutralization experiments with antiligase I antibodies indicated that the early and late eluting activity peaks were DNA ligase II and I, respectively. The three major ligase-[32P]AMP polypeptides (90, 78, and 67 kDa) were subsequently purified to near homogeneity by elution from preparative sodium dodecyl sulfate-polyacrylamide gels. All three polypeptides retained DNA ligase activities after gel elution and renaturation. To further reveal the relationship between these enzymes, partial digestion by V8-protease was performed. All three purified polypeptides gave rise to a common 22-kDa-labeled fragment for their AMP-binding domains, indicating that the catalytic sites of ligase I and II are quite similar, if not identical. Similar findings were obtained from the two-dimensional gel electrophoresis of their AMP-binding domains in the trypsin-digested protein fragments. The results also suggested that these isozymes have been derived from the same primordial DNA sequence or from the same precursor protein. The purification scheme and the data obtained will be instrumental for the further elucidation of the biological roles of various DNA ligases from human cells.

Crystal structures of Hsp104 N-terminal domains from Saccharomyces cerevisiae and Candida albicans suggest the mechanism for the function of Hsp104 in dissolving prions

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Peng; Li, Jingzhi; Weaver, Clarissa

Hsp104 is a yeast member of the Hsp100 family which functions as a molecular chaperone to disaggregate misfolded polypeptides. To understand the mechanism by which the Hsp104 N-terminal domain (NTD) interacts with its peptide substrates, crystal structures of the Hsp104 NTDs fromSaccharomyces cerevisiae(ScHsp104NTD) andCandida albicans(CaHsp104NTD) have been determined at high resolution. The structures of ScHsp104NTD and CaHsp104NTD reveal that the yeast Hsp104 NTD may utilize a conserved putative peptide-binding groove to interact with misfolded polypeptides. In the crystal structures ScHsp104NTD forms a homodimer, while CaHsp104NTD exists as a monomer. The consecutive residues Gln105, Gln106 and Lys107, and Lys141 around themore » putative peptide-binding groove mediate the monomer–monomer interactions within the ScHsp104NTD homodimer. Dimer formation by ScHsp104NTD suggests that the Hsp104 NTD may specifically interact with polyQ regions of prion-prone proteins. The data may reveal the mechanism by which Hsp104 NTD functions to suppress and/or dissolve prions.« less

Packaging of the virion host shutoff (Vhs) protein of herpes simplex virus: two forms of the Vhs polypeptide are associated with intranuclear B and C capsids, but only one is associated with enveloped virions.

PubMed

Read, G Sullivan; Patterson, Mary

2007-02-01

The virion host shutoff (Vhs) protein (UL41) is a minor component of herpes simplex virus virions which, following penetration, accelerates turnover of host and viral mRNAs. Infected cells contain 58-kDa and 59.5-kDa forms of Vhs, which differ in the extent of phosphorylation, yet only a 58-kDa polypeptide is incorporated into virions. In pulse-chase experiments, the primary Vhs translation product comigrated in sodium dodecyl sulfate-polyacrylamide gel electrophoresis with the 58-kDa virion polypeptide, and could be chased to 59.5 kDa. While both 59.5-kDa and 58-kDa forms were found in nuclear and cytoplasmic fractions, the 59.5-kDa form was significantly enriched in the nucleus. Both forms were associated with intranuclear B and C capsids, yet only the 58-kDa polypeptide was found in enveloped cytoplasmic virions. A 58-kDa form, but not the 59.5-kDa form, was found in L particles, noninfectious particles that contain an envelope and tegument but no capsid. The data suggest that virions contain two populations of Vhs that are packaged by different pathways. In the first pathway, the primary translation product is processed to 59.5 kDa, is transported to the nucleus, binds intranuclear capsids, and is converted to 58 kDa at some stage prior to final envelopment. The second pathway does not involve the 59.5-kDa form or interactions between Vhs and capsids. Instead, the primary translation product is phosphorylated to the 58-kDa virion form and packaged through interactions with other tegument proteins in the cytoplasm or viral envelope proteins at the site of final envelopment.

Role of aromatic interactions in amyloid formation by islet amyloid polypeptide.

PubMed

Tu, Ling-Hsien; Raleigh, Daniel P

2013-01-15

Aromatic-aromatic and aromatic-hydrophobic interactions have been proposed to play a role in amyloid formation by a range of polypeptides, including islet amyloid polypeptide (IAPP or amylin). IAPP is responsible for amyloid formation in patients with type 2 diabetes. The polypeptide is 37 residues long and contains three aromatic residues, Phe-15, Phe-23, and Tyr-37. The ability of all single aromatic to leucine mutants, all double aromatic to leucine mutants, and the triple leucine mutant to form amyloid were examined. Amyloid formation was almost twice as rapid for the F15L mutant as for the wild type but was almost 3-fold slower for the Y37L mutant and almost 2-fold slower for the F23L mutant. Amyloid fibrils formed from each of the single mutants were effective at seeding amyloid formation by wild-type IAPP, implying that the fibril structures are similar. The F15L/F23L double mutant has a larger effect than the F15L/Y37L double mutant on the rate of amyloid formation, even though a Y37L substitution has more drastic consequences in the wild-type background than does the F23L mutation, suggesting nonadditive effects between the different sites. The triple leucine mutant and the F23L/Y37L double mutant are the slowest to form amyloid. F15 has been proposed to make important contacts early in the aggregation pathway, but the data for the F15L mutant indicate that they are not optimal. A set of variants containing natural and unnatural amino acids at position 15, which were designed to conserve hydrophobicity, but alter α-helix and β-sheet propensity, were analyzed to determine the properties of this position that control the rate of amyloid formation. There is no correlation between β-sheet propensity at this position and the rate of amyloid formation, but there is a correlation with α-helical propensity.

Py4Syn: Python for synchrotrons.

PubMed

Slepicka, H H; Canova, H F; Beniz, D B; Piton, J R

2015-09-01

In this report, Py4Syn, an open-source Python-based library for data acquisition, device manipulation, scan routines and other helper functions, is presented. Driven by easy-to-use and scalability ideals, Py4Syn offers control system agnostic solution and high customization level for scans and data output, covering distinct techniques and facilities. Here, most of the library functionalities are described, examples of use are shown and ideas for future implementations are presented.

Keys to genera of the spider wasps (Hymenoptera: Pompilidae) of Russia and neighbouring countries, with check-list of genera.

PubMed

Loktionov, Valery M; Lelej, Arkady S

2015-10-28

Keys to 55 genera of spider wasps of Russia and neighbouring countries in females and males are given. Of them 34 genera are distributed in Russia. An annotated list of genera with type species and distribution data within Russia and biogeographical regions is given. The genus Xenaporus Ashmead, 1902 and X. eremocanus Wolf, 1990 are newly recorded from Russia. According to ICZN 1995 (Opinion 1820) new synonymy (valid name first) is proposed for the type species of genus Cryptocheilus Panzer, 1806: Sphex annulata Fabricius, 1798 (=Pompilus alternatus Lepeletier de Saint Fargeau, 1845, syn. nov.; =Pompilus comparatus Smith, 1855, syn. nov.; =Priocnemis culpabilis Costa, 1893, syn. nov.; Salius annulatilis Richards, 1935, syn. nov.).

(67/68)Ga-labeling agent that liberates (67/68)Ga-NOTA-methionine by lysosomal proteolysis of parental low molecular weight polypeptides to reduce renal radioactivity levels.

PubMed

Uehara, Tomoya; Rokugawa, Takemi; Kinoshita, Mai; Nemoto, Souki; Fransisco Lazaro, Guerra Gomez; Hanaoka, Hirofumi; Arano, Yasushi

2014-11-19

The renal localization of gallium-67 or gallium-68 ((67/68)Ga)-labeled low molecular weight (LMW) probes such as peptides and antibody fragments constitutes a problem in targeted imaging. Wu et al. previously showed that (67)Ga-labeled S-2-(4-isothiocyanatobenzyl)-1,4,7-triazacyclononane-1,4,7-triacetic acid (SCN-Bz-NOTA)-conjugated methionine ((67)Ga-NOTA-Met) was rapidly excreted from the kidney in urine following lysosomal proteolysis of the parental (67)Ga-NOTA-Bz-SCN-disulfide-stabilized Fv fragment (Bioconjugate Chem., (1997) 8, 365-369). In the present study, a new (67/68)Ga-labeling reagent for LMW probes that liberates (67/68)Ga-NOTA-Met was designed, synthesized, and evaluated using longer-lived (67)Ga in order to reduce renal radioactivity levels. We employed a methionine-isoleucine (MI) dipeptide bond as the cleavable linkage. The amine residue of MI was coupled with SCN-Bz-NOTA for (67)Ga-labeling, while the carboxylic acid residue of MI was derivatized to maleimide for antibody conjugation in order to synthesize NOTA-MI-Mal. A Fab fragment of the anti-Her2 antibody was thiolated with iminothiolane, and NOTA-MI-Mal was conjugated with the antibody fragment by maleimide-thiol chemistry. The Fab fragment was also conjugated with SCN-Bz-NOTA (NOTA-Fab) for comparison. (67)Ga-NOTA-MI-Fab was obtained at radiochemical yields of over 95% and was stable in murine serum for 24 h. In the biodistribution study using normal mice, (67)Ga-NOTA-MI-Fab registered significantly lower renal radioactivity levels from 1 to 6 h postinjection than those of (67)Ga-NOTA-Fab. An analysis of urine samples obtained 6 h after the injection of (67)Ga-NOTA-MI-Fab showed that the majority of radioactivity was excreted as (67)Ga-NOTA-Met. In the biodistribution study using tumor-bearing mice, the tumor to kidney ratios of (67)Ga-NOTA-MI-Fab were 4 times higher (6 h postinjection) than those of (67)Ga-NOTA-Fab. Although further studies including the structure of radiometabolites and/or cleavable linkages are required, the results of the present study indicate that the current chemical design is applicable to the development of (67)Ga-labeled Fabs for low renal radioactivity levels.

Investigating antimalarial drug interactions of emetine dihydrochloride hydrate using CalcuSyn-based interactivity calculations

PubMed Central

Matthews, Holly; Deakin, Jon; Rajab, May; Idris-Usman, Maryam

2017-01-01

The widespread introduction of artemisinin-based combination therapy has contributed to recent reductions in malaria mortality. Combination therapies have a range of advantages, including synergism, toxicity reduction, and delaying the onset of resistance acquisition. Unfortunately, antimalarial combination therapy is limited by the depleting repertoire of effective drugs with distinct target pathways. To fast-track antimalarial drug discovery, we have previously employed drug-repositioning to identify the anti-amoebic drug, emetine dihydrochloride hydrate, as a potential candidate for repositioned use against malaria. Despite its 1000-fold increase in in vitro antimalarial potency (ED50 47 nM) compared with its anti-amoebic potency (ED50 26–32 uM), practical use of the compound has been limited by dose-dependent toxicity (emesis and cardiotoxicity). Identification of a synergistic partner drug would present an opportunity for dose-reduction, thus increasing the therapeutic window. The lack of reliable and standardised methodology to enable the in vitro definition of synergistic potential for antimalarials is a major drawback. Here we use isobologram and combination-index data generated by CalcuSyn software analyses (Biosoft v2.1) to define drug interactivity in an objective, automated manner. The method, based on the median effect principle proposed by Chou and Talalay, was initially validated for antimalarial application using the known synergistic combination (atovaquone-proguanil). The combination was used to further understand the relationship between SYBR Green viability and cytocidal versus cytostatic effects of drugs at higher levels of inhibition. We report here the use of the optimised Chou Talalay method to define synergistic antimalarial drug interactivity between emetine dihydrochloride hydrate and atovaquone. The novel findings present a potential route to harness the nanomolar antimalarial efficacy of this affordable natural product. PMID:28257497

Image Guided Radiation Therapy Using Synthetic Computed Tomography Images in Brain Cancer

DOE Office of Scientific and Technical Information (OSTI.GOV)

Price, Ryan G.; Department of Radiation Oncology, Wayne State University School of Medicine, Detroit, Michigan; Kim, Joshua P.

Purpose: The development of synthetic computed tomography (CT) (synCT) derived from magnetic resonance (MR) images supports MR-only treatment planning. We evaluated the accuracy of synCT and synCT-generated digitally reconstructed radiographs (DRRs) relative to CT and determined their performance for image guided radiation therapy (IGRT). Methods and Materials: Magnetic resonance simulation (MR-SIM) and CT simulation (CT-SIM) images were acquired of an anthropomorphic skull phantom and 12 patient brain cancer cases. SynCTs were generated using fluid attenuation inversion recovery, ultrashort echo time, and Dixon data sets through a voxel-based weighted summation of 5 tissue classifications. The DRRs were generated from the phantommore » synCT, and geometric fidelity was assessed relative to CT-generated DRRs through bounding box and landmark analysis. An offline retrospective analysis was conducted to register cone beam CTs (n=34) to synCTs and CTs using automated rigid registration in the treatment planning system. Planar MV and KV images (n=37) were rigidly registered to synCT and CT DRRs using an in-house script. Planar and volumetric registration reproducibility was assessed and margin differences were characterized by the van Herk formalism. Results: Bounding box and landmark analysis of phantom synCT DRRs were within 1 mm of CT DRRs. Absolute planar registration shift differences ranged from 0.0 to 0.7 mm for phantom DRRs on all treatment platforms and from 0.0 to 0.4 mm for volumetric registrations. For patient planar registrations, the mean shift differences were 0.4 ± 0.5 mm (range, −0.6 to 1.6 mm), 0.0 ± 0.5 mm (range, −0.9 to 1.2 mm), and 0.1 ± 0.3 mm (range, −0.7 to 0.6 mm) for the superior-inferior (S-I), left-right (L-R), and anterior-posterior (A-P) axes, respectively. The mean shift differences in volumetric registrations were 0.6 ± 0.4 mm (range, −0.2 to 1.6 mm), 0.2 ± 0.4 mm (range, −0.3 to 1.2 mm), and 0.2 ± 0.3 mm (range, −0.2 to 1.2 mm) for the S-I, L-R, and A-P axes, respectively. The CT-SIM and synCT derived margins were <0.3 mm different. Conclusion: DRRs generated by synCT were in close agreement with CT-SIM. Planar and volumetric image registrations to synCT-derived targets were comparable with CT for phantom and patients. This validation is the next step toward MR-only planning for the brain.« less

Targeted polypeptide degradation

DOEpatents

Church, George M [Brookline, MA; Janse, Daniel M [Brookline, MA

2008-05-13

This invention pertains to compositions, methods, cells and organisms useful for selectively localizing polypeptides to the proteasome for degradation. Therapeutic methods and pharmaceutical compositions for treating disorders associated with the expression and/or activity of a polypeptide by targeting these polypeptides for degradation, as well as methods for targeting therapeutic polypeptides for degradation and/or activating therapeutic polypeptides by degradation are provided. The invention provides methods for identifying compounds that mediate proteasome localization and/or polypeptide degradation. The invention also provides research tools for the study of protein function.

Hand-rim forces and gross mechanical efficiency in asynchronous and synchronous wheelchair propulsion: a comparison.

PubMed

Lenton, J P; van der Woude, L; Fowler, N; Nicholson, G; Tolfrey, K; Goosey-Tolfrey, V

2014-03-01

To compare the force application characteristics at various push frequencies of asynchronous (ASY) and synchronous (SYN) hand-rim propulsion, 8 able-bodied participants performed a separate sub-maximal exercise test on a wheelchair roller ergometer for each propulsion mode. Each test consisted of a series of 5, 4-min exercise blocks at 1.8 m · s-1 - initially at their freely chosen frequency (FCF), followed by four counter-balanced trials at 60, 80, 120 and 140% FCF. Kinetic data was obtained using a SMARTWheel, measuring forces and moments. The gross efficiency (GE) was determined as the ratio of external work done and the total energy expended. The ASY propulsion produced higher force measures for FRES, FTAN, rate of force development & FEF (P<0.05), while there was no difference in GE values (P=0.518). In pair-matched push frequencies (ASY80:SYN60, ASY100:SYN80, ASY120:SYN100 and ASY140:SYN120), ASY propulsion forces remained significantly higher (FRES, FTAN, rate of force development & FEF P<0.05), and there was no significant effect on GE (P=0.456). Both ASY and SYN propulsion demonstrate similar trends: changes in push frequency are accompanied by changes in absolute force even without changes in the gross pattern/trend of force application, FEF or GE. Matched push frequencies continue to produce significant differences in force measures but not GE. This suggests ASY propulsion is the predominant factor in force application differences. The ASY would appear to offer a kinetic disadvantage to SYN propulsion and no physiological advantage under current testing conditions. © Georg Thieme Verlag KG Stuttgart · New York.

Trehalose does not improve neuronal survival on exposure to alpha-synuclein pre-formed fibrils.

PubMed

Redmann, Matthew; Wani, Willayat Y; Volpicelli-Daley, Laura; Darley-Usmar, Victor; Zhang, Jianhua

2017-04-01

Parkinson's disease is a debilitating neurodegenerative disorder that is pathologically characterized by intracellular inclusions comprised primarily of alpha-synuclein (αSyn) that can also be transmitted from neuron to neuron. Several lines of evidence suggest that these inclusions cause neurodegeneration. Thus exploring strategies to improve neuronal survival in neurons with αSyn aggregates is critical. Previously, exposure to αSyn pre-formed fibrils (PFFs) has been shown to induce aggregation of endogenous αSyn resulting in cell death that is exacerbated by either starvation or inhibition of mTOR by rapamycin, both of which are able to induce autophagy, an intracellular protein degradation pathway. Since mTOR inhibition may also inhibit protein synthesis and starvation itself can be detrimental to neuronal survival, we investigated the effects of autophagy induction on neurons with αSyn inclusions by a starvation and mTOR-independent autophagy induction mechanism. We exposed mouse primary cortical neurons to PFFs to induce inclusion formation in the presence and absence of the disaccharide trehalose, which has been proposed to induce autophagy and stimulate lysosomal biogenesis. As expected, we observed that on exposure to PFFs, there was increased abundance of pS129-αSyn aggregates and cell death. Trehalose alone increased LC3-II levels, consistent with increased autophagosome levels that remained elevated with PFF exposure. Interestingly, trehalose alone increased cell viability over a 14-d time course. Trehalose was also able to restore cell viability to control levels, but PFFs still exhibited toxic effects on the cells. These data provide essential information regarding effects of trehalose on αSyn accumulation and neuronal survival on exposure to PFF. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

Antimüllerian hormone levels and cardiometabolic risk in young women with polycystic ovary syndrome.

PubMed

Feldman, Rebecca A; O'Neill, Kathleen; Butts, Samantha F; Dokras, Anuja

2017-01-01

To determine the association between antimüllerian hormone (AMH) levels and metabolic syndrome (MetSyn) in young women with polycystic ovary syndrome (PCOS). Cross-sectional study. Academic PCOS center. A total of 252 women aged 18-46 years with PCOS. None. Association of AMH with markers of cardiometabolic risk and MetSyn. The median AMH level was 5.1 ng/mL (interquartile range [IQR] 3.0-8.1), and prevalence of MetSyn was 23.8%. AMH levels positively correlated with total T, high-density lipoprotein (HDL) cholesterol, and SHBG and negatively correlated with fasting glucose, homeostasis-model assessment of insulin resistance, body mass index (BMI), and systolic and diastolic blood pressure. A single-unit decrease in AMH was associated with an 11% increase in odds of MetSyn (odds ratio [OR] 1.11, 95% confidence interval [CI] 1.03-1.20); the strength of this association was maintained in the multivariate model (OR 1.09, 95% CI 1.01-1.18) adjusting for age and race. Subjects with AMH values in the lowest tertile were twice as likely as those in the highest tertile to have MetSyn (adjusted OR 2.1, 95% CI 1.01-4.3). Total T was not associated with MetSyn or its individual components. Our findings indicate that in young women with PCOS, low AMH levels predict a greater risk of MetSyn. The role of AMH, an established biomarker of ovarian reserve, in risk stratification of cardiometabolic risk in obese women with PCOS needs to be clarified in longitudinal studies and in the perimenopausal population. Copyright © 2016 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

Anle138b Partly Ameliorates Motor Deficits Despite Failure of Neuroprotection in a Model of Advanced Multiple System Atrophy

PubMed Central

Fellner, Lisa; Kuzdas-Wood, Daniela; Levin, Johannes; Ryazanov, Sergey; Leonov, Andrei; Griesinger, Christian; Giese, Armin; Wenning, Gregor K.; Stefanova, Nadia

2016-01-01

The neurodegenerative disorder multiple system atrophy (MSA) is characterized by autonomic failure, cerebellar ataxia and parkinsonism in any combination associated with predominantly oligodendroglial α-synuclein (α-syn) aggregates (glial cytoplasmic inclusions = GCIs). To date, there is no effective disease modifying therapy. Previous experiments have shown that the aggregation inhibitor anle138b reduces neurodegeneration, as well as behavioral deficits in both transgenic and toxin mouse models of Parkinson's disease (PD). Here we analyzed whether anle138b improves motor skills and reduces neuronal loss, as well as oligodendroglial α-syn aggregation in the PLP-α-syn transgenic mouse challenged with the mitochondrial toxin 3-nitropropionic acid (3-NP) to model full-blown MSA. Following 1 month of treatment with anle138b, MSA mice showed signs of motor improvement affecting stride length, but not pole, grip strength, and beam test performance. Loss of dopaminergic nigral neurons and Purkinje cells was not attenuated and GCI density remained unchanged. These data suggest that the pathology in transgenic PLP-α-syn mice receiving 3-NP might be too advanced to detect significant effects of anle138b treatment on neuronal loss and intracytoplasmic α-syn inclusion bodies. However, the partial motor amelioration may indicate potential efficacy of anle138b treatment that may be mediated by its actions on α-syn oligomers or may reflect improvement of neuronal dysfunction in neural at risk populations. Further studies are required to address the efficacy of anle138b in transgenic α-syn models of early-stage MSA and in the absence of additional toxin application. PMID:27013960

Characterization of cognitive deficits in rats overexpressing human alpha-synuclein in the ventral tegmental area and medial septum using recombinant adeno-associated viral vectors.

PubMed

Hall, Hélène; Jewett, Michael; Landeck, Natalie; Nilsson, Nathalie; Schagerlöf, Ulrika; Leanza, Giampiero; Kirik, Deniz

2013-01-01

Intraneuronal inclusions containing alpha-synuclein (a-syn) constitute one of the pathological hallmarks of Parkinson's disease (PD) and are accompanied by severe neurodegeneration of A9 dopaminergic neurons located in the substantia nigra. Although to a lesser extent, A10 dopaminergic neurons are also affected. Neurodegeneration of other neuronal populations, such as the cholinergic, serotonergic and noradrenergic cell groups, has also been documented in PD patients. Studies in human post-mortem PD brains and in rodent models suggest that deficits in cholinergic and dopaminergic systems may be associated with the cognitive impairment seen in this disease. Here, we investigated the consequences of targeted overexpression of a-syn in the mesocorticolimbic dopaminergic and septohippocampal cholinergic pathways. Rats were injected with recombinant adeno-associated viral vectors encoding for either human wild-type a-syn or green fluorescent protein (GFP) in the ventral tegmental area and the medial septum/vertical limb of the diagonal band of Broca, two regions rich in dopaminergic and cholinergic neurons, respectively. Histopathological analysis showed widespread insoluble a-syn positive inclusions in all major projections areas of the targeted nuclei, including the hippocampus, neocortex, nucleus accumbens and anteromedial striatum. In addition, the rats overexpressing human a-syn displayed an abnormal locomotor response to apomorphine injection and exhibited spatial learning and memory deficits in the Morris water maze task, in the absence of obvious spontaneous locomotor impairment. As losses in dopaminergic and cholinergic immunoreactivity in both the GFP and a-syn expressing animals were mild-to-moderate and did not differ from each other, the behavioral impairments seen in the a-syn overexpressing animals appear to be determined by the long term persisting neuropathology in the surviving neurons rather than by neurodegeneration.

Hsp31 Is a Stress Response Chaperone That Intervenes in the Protein Misfolding Process*

PubMed Central

Tsai, Chai-jui; Aslam, Kiran; Drendel, Holli M.; Asiago, Josephat M.; Goode, Kourtney M.; Paul, Lake N.; Rochet, Jean-Christophe; Hazbun, Tony R.

2015-01-01

The Saccharomyces cerevisiae heat shock protein Hsp31 is a stress-inducible homodimeric protein that is involved in diauxic shift reprogramming and has glyoxalase activity. We show that substoichiometric concentrations of Hsp31 can abrogate aggregation of a broad array of substrates in vitro. Hsp31 also modulates the aggregation of α-synuclein (αSyn), a target of the chaperone activity of human DJ-1, an Hsp31 homolog. We demonstrate that Hsp31 is able to suppress the in vitro fibrillization or aggregation of αSyn, citrate synthase and insulin. Chaperone activity was also observed in vivo because constitutive overexpression of Hsp31 reduced the incidence of αSyn cytoplasmic foci, and yeast cells were rescued from αSyn-generated proteotoxicity upon Hsp31 overexpression. Moreover, we showed that Hsp31 protein levels are increased by H2O2, in the diauxic phase of normal growth conditions, and in cells under αSyn-mediated proteotoxic stress. We show that Hsp31 chaperone activity and not the methylglyoxalase activity or the autophagy pathway drives the protective effects. We also demonstrate reduced aggregation of the Sup35 prion domain, PrD-Sup35, as visualized by fluorescent protein fusions. In addition, Hsp31 acts on its substrates prior to the formation of large aggregates because Hsp31 does not mutually localize with prion aggregates, and it prevents the formation of detectable in vitro αSyn fibrils. These studies establish that the protective role of Hsp31 against cellular stress is achieved by chaperone activity that intervenes early in the protein misfolding process and is effective on a wide spectrum of substrate proteins, including αSyn and prion proteins. PMID:26306045

'Prion-like' propagation of the synucleinopathy of M83 transgenic mice depends on the mouse genotype and type of inoculum.

PubMed

Sargent, Dorian; Verchère, Jérémy; Lazizzera, Corinne; Gaillard, Damien; Lakhdar, Latifa; Streichenberger, Nathalie; Morignat, Eric; Bétemps, Dominique; Baron, Thierry

2017-10-01

The M83 transgenic mouse is a model of human synucleinopathies that develops severe motor impairment correlated with accumulation of the pathological Ser129-phosphorylated α-synuclein (α-syn P ) in the brain and spinal cord. M83 disease can be accelerated by intracerebral inoculation of brain extracts from sick M83 mice. This has also recently been described using peripheral routes, injecting recombinant preformed α-syn fibrils into the muscle or the peritoneum. Here, we inoculated homozygous and/or hemizygous M83 neonates via the intraperitoneal and/or intracerebral routes with two different brain extracts: one from sick M83 mice inoculated with brain extract from other sick M83 mice, and the other derived from a human multiple system atrophy source passaged in M83 mice. Detection of α-syn P using ELISA and western blot confirmed the disease in mice. The distribution of α-syn P in the central nervous system was similar, independently of the inoculum or inoculation route, consistent with previous studies describing M83 disease. ELISA tests revealed higher levels of α-syn P in homozygous than in hemizygous sick M83 mice, at least after IC inoculation. Interestingly, the immunoreactivity of α-syn P detected by ELISA was significantly lower in M83 mice inoculated with the multiple system atrophy inoculum than in M83 mice inoculated with the M83 inoculum, at the first two passages. 'Prion-like' propagation of the synucleinopathy up to the clinical disease was accelerated by both intracerebral and intraperitoneal inoculations of brain extracts from sick mice. This acceleration, however, depends on the levels of α-syn expression by the mouse and the type of inoculum. © 2017 International Society for Neurochemistry.

Decreased expression of serum- and glucocorticoid-inducible kinase 1 (SGK1) promotes alpha-synuclein increase related with down-regulation of dopaminergic cell in the Substantia Nigra of chronic MPTP-induced Parkinsonism mice and in SH-SY5Y cells.

PubMed

Yeo, Sujung; Sung, Backil; Hong, Yeon-Mi; van den Noort, Maurits; Bosch, Peggy; Lee, Sook-Hyun; Song, Jongbeom; Park, Sang-Kyun; Lim, Sabina

2018-06-30

Parkinson's disease (PD) is a chronically progressive neurodegenerative disease, with its main pathological hallmarks being a dramatic loss of dopaminergic neurons predominantly in the Substantia Nigra (SN), and the formations of intracytoplasmic Lewy bodies and dystrophic neurites. Alpha-synuclein (α-syn), widely recognized as the most prominent element of the Lewy body, is one of the representative hallmarks in PD. However, the mechanisms behind the increased α-syn expression and aggregation have not yet been clarified. To examine what causes α-syn expression to increase, we analyzed the pattern of gene expression in the SN of mice intoxicated with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), where down-regulation of dopaminergic cells occurred. We identified serum- and glucocorticoid-dependent kinase 1 (SGK1) as one of the genes that is evidently downregulated in chronic MPTP-intoxication. The results of Western blot analyses showed that, together with the down-regulation of dopaminergic cells, the decrease in SGK1 expression increased α-syn expression in the SN in a chronic MPTP-induced Parkinsonism mouse. For an examination of the expression correlation between SGK1 and α-syn, SH-5YSY cells were knocked down with SGK1 siRNA then, the downregulation of dopaminergic cells and the increase in the expression of α-syn were observed. These results suggest that decreased expression of SGK1 may play a critical role in increasing the expression of α-syn, which is related with dopaminergic cell death in the SN of chronic MPTP-induced Parkinsonism mice and in SH-SY5Y cells. Copyright © 2018. Published by Elsevier B.V.

Loss or Mislocalization of Aquaporin-4 Affects Diffusion Properties and Intermediary Metabolism in Gray Matter of Mice.

PubMed

Pavlin, T; Nagelhus, E A; Brekken, C; Eyjolfsson, E M; Thoren, A; Haraldseth, O; Sonnewald, U; Ottersen, O P; Håberg, A K

2017-01-01

The first aim of this study was to determine how complete or perivascular loss of aquaporin-4 (AQP4) water channels affects membrane permeability for water in the mouse brain grey matter in the steady state. Time-dependent diffusion magnetic resonance imaging was performed on global Aqp4 knock out (KO) and α-syntrophin (α-syn) KO mice, in the latter perivascular AQP4 are mislocalized, but still functioning. Control animals were corresponding wild type (WT) mice. By combining in vivo diffusion measurements with the effective medium theory and previously measured extra-cellular volume fractions, the effects of membrane permeability and extracellular volume fraction were uncoupled for Aqp4 and α-syn KO. The second aim was to assess the effect of α-syn KO on cortical intermediary metabolism combining in vivo [1- 13 C]glucose and [1,2- 13 C]acetate injection with ex vivo 13 C MR spectroscopy. Aqp4 KO increased the effective diffusion coefficient at long diffusion times by 5%, and a 14% decrease in membrane water permeability was estimated for Aqp4 KO compared with WT mice. α-syn KO did not affect the measured diffusion parameters. In the metabolic analyses, significantly lower amounts of [4- 13 C]glutamate and [4- 13 C]glutamine, and percent enrichment in [4- 13 C]glutamate were detected in the α-syn KO mice. [1,2- 13 C]acetate metabolism was unaffected in α-syn KO, but the contribution of astrocyte derived metabolites to GABA synthesis was significantly increased. Taken together, α-syn KO mice appeared to have decreased neuronal glucose metabolism, partly compensated for by utilization of astrocyte derived metabolites.

Properties of Streptococcus mutans Grown in a Synthetic Medium: Binding of Glucosyltransferase and In Vitro Adherence, and Binding of Dextran/Glucan and Glycoprotein and Agglutination

PubMed Central

Wu-Yuan, Christine D.; Tai, Stella; Slade, Hutton D.

1979-01-01

The influence of culture media on various properties of Streptococcus mutans was investigated. Strains of S. mutans (serotypes c, d, f, and g) were grown in a complex medium (Todd-Hewitt broth [THB]) or a synthetic medium (SYN). The SYN cells, in contrast to THB cells, did not bind extracellular glucosyltransferase and did not produce in vitro adherence. Both types of cells possessed constitutive levels of glucosyltransferase. B13 cells grown in SYN plus invertase-treated glucose possessed the same level of constitutive enzyme as THB cells. In contrast to THB cells, the SYN cells of seven serotype strains did not agglutinate upon the addition of high-molecular-weight dextran/glucan. Significant quantities of lower-molecular-weight (2 × 104 or 7 × 104) dextran and B13 glucan were bound by SYN cells. SYN cells agglutinated weakly in anti-glucan serum (titers, 0 to 16), whereas THB cells possessed titers of 32 to 256. Evidence for the existence of a second binding site in agglutination which does not possess a glucan-like polymer has been obtained. B13 cells grown in invertase-treated THB agglutinated to the same degree as normal THB cells. The nature of this site is unknown. SYN cells possess the type-specific polysaccharide antigen. B13 cells did not bind from THB a glycoprotein which reacts with antisera to the A, B, or T blood group antigens or which allows agglutination upon the addition of dextran. The results demonstrate that S. mutans grown in a chemically defined medium possesse markedly different biochemical and biological activities than cells grown in a complex organic medium. PMID:457252

Hydrogenase polypeptide and methods of use

DOEpatents

Adams, Michael W.W.; Hopkins, Robert C.; Jenney, JR, Francis E.; Sun, Junsong

2016-02-02

Provided herein are polypeptides having hydrogenase activity. The polypeptide may be multimeric, and may have hydrogenase activity of at least 0.05 micromoles H.sub.2 produced min.sup.-1 mg protein.sup.-1. Also provided herein are polynucleotides encoding the polypeptides, genetically modified microbes that include polynucleotides encoding one or more subunits of the multimeric polypeptide, and methods for making and using the polypeptides.

An in vivo library-versus-library selection of optimized protein-protein interactions.

PubMed

Pelletier, J N; Arndt, K M; Plückthun, A; Michnick, S W

1999-07-01

We describe a rapid and efficient in vivo library-versus-library screening strategy for identifying optimally interacting pairs of heterodimerizing polypeptides. Two leucine zipper libraries, semi-randomized at the positions adjacent to the hydrophobic core, were genetically fused to either one of two designed fragments of the enzyme murine dihydrofolate reductase (mDHFR), and cotransformed into Escherichia coli. Interaction between the library polypeptides reconstituted enzymatic activity of mDHFR, allowing bacterial growth. Analysis of the resulting colonies revealed important biases in the zipper sequences relative to the original libraries, which are consistent with selection for stable, heterodimerizing pairs. Using more weakly associating mDHFR fragments, we increased the stringency of selection. We enriched the best-performing leucine zipper pairs by multiple passaging of the pooled, selected colonies in liquid culture, as the best pairs allowed for better bacterial propagation. This competitive growth allowed small differences among the pairs to be amplified, and different sequence positions were enriched at different rates. We applied these selection processes to a library-versus-library sample of 2.0 x 10(6) combinations and selected a novel leucine zipper pair that may be appropriate for use in further in vivo heterodimerization strategies.

Disruption of hippocampus-regulated behavioural and cognitive processes by heterozygous constitutive deletion of SynGAP.

PubMed

Muhia, Mary; Yee, Benjamin K; Feldon, Joram; Markopoulos, Foivos; Knuesel, Irene

2010-02-01

The brain-specific Ras/Rap-GTPase activating protein (SynGAP) is a prime candidate linking N-methyl-d-aspartate receptors to the regulation of the ERK/MAP kinase signalling cascade, suggested to be essential for experience-dependent synaptic plasticity. Here, we evaluated the behavioural phenotype of SynGAP heterozygous knockout mice (SG(+/-)), expressing roughly half the normal levels of SynGAP. In the cognitive domain, SG(+/-) mice demonstrated severe working and reference memory deficits in the radial arm maze task, a mild impairment early in the transfer test of the water maze task, and a deficiency in spontaneous alternation in an elevated T-maze. In the non-cognitive domain, SG(+/-) mice were hyperactive in the open field and appeared less anxious in the elevated plus maze test. In contrast, object recognition memory performance was not impaired in SG(+/-) mice. The reduction in SynGAP thus resulted in multiple behavioural traits suggestive of aberrant cognitive and non-cognitive processes normally mediated by the hippocampus. Immunohistochemical evaluation further revealed a significant reduction in calbindin-positive interneurons in the hippocampus and doublecortin-positive neurons in the dentate gyrus of adult SG(+/-) mice. Heterozygous constitutive deletion of SynGAP is therefore associated with notable behavioural as well as morphological phenotypes indicative of hippocampal dysfunction. Any suggestion of a possible causal link between them however remains a matter for further investigation.

Recapitulating the Structural Evolution of Redox Regulation in Adenosine 5'-Phosphosulfate Kinase from Cyanobacteria to Plants.

PubMed

Herrmann, Jonathan; Nathin, David; Lee, Soon Goo; Sun, Tony; Jez, Joseph M

2015-10-09

In plants, adenosine 5'-phosphosulfate (APS) kinase (APSK) is required for reproductive viability and the production of 3'-phosphoadenosine 5'-phosphosulfate (PAPS) as a sulfur donor in specialized metabolism. Previous studies of the APSK from Arabidopsis thaliana (AtAPSK) identified a regulatory disulfide bond formed between the N-terminal domain (NTD) and a cysteine on the core scaffold. This thiol switch is unique to mosses, gymnosperms, and angiosperms. To understand the structural evolution of redox control of APSK, we investigated the redox-insensitive APSK from the cyanobacterium Synechocystis sp. PCC 6803 (SynAPSK). Crystallographic analysis of SynAPSK in complex with either APS and a non-hydrolyzable ATP analog or APS and sulfate revealed the overall structure of the enzyme, which lacks the NTD found in homologs from mosses and plants. A series of engineered SynAPSK variants reconstructed the structural evolution of the plant APSK. Biochemical analyses of SynAPSK, SynAPSK H23C mutant, SynAPSK fused to the AtAPSK NTD, and the fusion protein with the H23C mutation showed that the addition of the NTD and cysteines recapitulated thiol-based regulation. These results reveal the molecular basis for structural changes leading to the evolution of redox control of APSK in the green lineage from cyanobacteria to plants. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Conversion of Natively Unstructured α-Synuclein to Its α-Helical Conformation Significantly Attenuates Production of Reactive Oxygen Species

PubMed Central

Zhou, Binbin; Hao, Yuanqiang; Wang, Chengshan; Li, Ding; Liu, You-Nian; Zhou, Feimeng

2012-01-01

The intracellular α-synuclein (α-syn) protein, whose conformational change and aggregation have been closely linked to the pathology of Parkingson’s disease (PD), is highly populated at the presynaptic termini and remains there in the α-helical conformation. In this study, circular dichroism confirmed that natively unstructured α-syn in aqueous solution was transformed to its α-helical conformation upon addition of trifluoroethanol (TFE). Electrochemical and UV–visible spectroscopic experiments reveal that both Cu(I) and Cu(II) are stabilized, with the former being stabilized by about two orders of magnitude. Compared to unstructured α-syn (Binolfi et al., J. Am. Chem. Soc. 133 (2011) 194–196), α-helical α-syn stabilizes Cu(I) by more than three orders of magnitude. Through the measurements of H2O2 and hydroxyl radicals (OH•) in solutions containing different forms of Cu(II) (free and complexed by unstructured or α-helical α-syn), we demonstrate that the significantly enhanced Cu(I) binding affinity helps inhibit the production of highly toxic reactive oxygen species, especially the hydroxyl radicals. Our study provides strong evidence that, as a possible means to prevent neuronal cell damage, conversion of the natively unstructured α-syn to its α-helical conformation in vivo could significantly attenuate the copper-modulated ROS production. PMID:23123341

(Poly)phenol-digested metabolites modulate alpha-synuclein toxicity by regulating proteostasis.

PubMed

Macedo, Diana; Jardim, Carolina; Figueira, Inês; Almeida, A Filipa; McDougall, Gordon J; Stewart, Derek; Yuste, Jose E; Tomás-Barberán, Francisco A; Tenreiro, Sandra; Outeiro, Tiago F; Santos, Cláudia N

2018-05-03

Parkinson's disease (PD) is an age-related neurodegenerative disease associated with the misfolding and aggregation of alpha-synuclein (aSyn). The molecular underpinnings of PD are still obscure, but nutrition may play an important role in the prevention, onset, and disease progression. Dietary (poly)phenols revert and prevent age-related cognitive decline and neurodegeneration in model systems. However, only limited attempts were made to evaluate the impact of digestion on the bioactivities of (poly)phenols and determine their mechanisms of action. This constitutes a challenge for the development of (poly)phenol-based nutritional therapies. Here, we subjected (poly)phenols from Arbutus unedo to in vitro digestion and tested the products in cell models of PD based on the cytotoxicity of aSyn. The (poly)phenol-digested metabolites from A. unedo leaves (LPDMs) effectively counteracted aSyn and H 2 O 2 toxicity in yeast and human cells, improving viability by reducing aSyn aggregation and inducing its clearance. In addition, LPDMs modulated pathways associated with aSyn toxicity, such as oxidative stress, endoplasmic reticulum (ER) stress, mitochondrial impairment, and SIR2 expression. Overall, LPDMs reduced aSyn toxicity, enhanced the efficiency of ER-associated protein degradation by the proteasome and autophagy, and reduced oxidative stress. In total, our study opens novel avenues for the exploitation of (poly)phenols in nutrition and health.

Active inhibition of herpes simplex virus type 1-induced cell fusion

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bzik, D.J.; Person, S.; Read, G.S.

1982-01-01

Previous studies have demonstrated that syn mutant-infected cells fuse less well with nonsyncytial virus-infected cells than with uninfected cells, a phenomenon defined as function inhibition. The present study characterizes the kinetics as well as the requirements for expression of fusion inhibition. Initially, the capacity of sparse syn mutant-infected cells to fuse with uninfected surrounding cells was determined throughout infection. Of seven syn mutants examined, including representatives with alterations in two different viral genes that affect cell fusion, all showed an increase in fusion capacity up to 12 hr after infection and a decrease at later times. Fusion inhibition was examinedmore » in experiments employing sparse syn20-infected cells which had been incubated to a maximum fusion capacity; it was shown that surrounding cells infected with KOS, the parent of syn20, began to inhibit fusion by the syn20-infected cells at about 4 hr after infection, and that the maximum ability to inhibit fusion was attained at about 6 hr after infection. The metabolic blocking agents actinomycin D (RNA), cycloheximide (protein), 2-deoxyglucose, and tunicamycin (glycoslyation of glycoproteins) all showed the ability to inhibit the expression of fusion inhibition by KOS-infected cells if added shortly after infection. It is concluded that fusion inhibition is an active process that requires the synthesis of RNA, proteins, and glycoproteins. 17 references, 3 figures, 2 tables.« less

A selection that reports on protein–protein interactions within a thermophilic bacterium

PubMed Central

Nguyen, Peter Q.; Silberg, Jonathan J.

2010-01-01

Many proteins can be split into fragments that exhibit enhanced function upon fusion to interacting proteins. While this strategy has been widely used to create protein-fragment complementation assays (PCAs) for discovering protein–protein interactions within mesophilic organisms, similar assays have not yet been developed for studying natural and engineered protein complexes at the temperatures where thermophilic microbes grow. We describe the development of a selection for protein–protein interactions within Thermus thermophilus that is based upon growth complementation by fragments of Thermotoga neapolitana adenylate kinase (AKTn). Complementation studies with an engineered thermophile (PQN1) that is not viable above 75°C because its adk gene has been replaced by a Geobacillus stearothermophilus ortholog revealed that growth could be restored at 78°C by a vector that coexpresses polypeptides corresponding to residues 1–79 and 80–220 of AKTn. In contrast, PQN1 growth was not complemented by AKTn fragments harboring a C156A mutation within the zinc-binding tetracysteine motif unless these fragments were fused to Thermotoga maritima chemotaxis proteins that heterodimerize (CheA and CheY) or homodimerize (CheX). This enhanced complementation is interpreted as arising from chemotaxis protein–protein interactions, since AKTn-C156A fragments having only one polypeptide fused to a chemotaxis protein did not complement PQN1 to the same extent. This selection increases the maximum temperature where a PCA can be used to engineer thermostable protein complexes and to map protein–protein interactions. PMID:20418388

A selection that reports on protein-protein interactions within a thermophilic bacterium.

PubMed

Nguyen, Peter Q; Silberg, Jonathan J

2010-07-01

Many proteins can be split into fragments that exhibit enhanced function upon fusion to interacting proteins. While this strategy has been widely used to create protein-fragment complementation assays (PCAs) for discovering protein-protein interactions within mesophilic organisms, similar assays have not yet been developed for studying natural and engineered protein complexes at the temperatures where thermophilic microbes grow. We describe the development of a selection for protein-protein interactions within Thermus thermophilus that is based upon growth complementation by fragments of Thermotoga neapolitana adenylate kinase (AK(Tn)). Complementation studies with an engineered thermophile (PQN1) that is not viable above 75 degrees C because its adk gene has been replaced by a Geobacillus stearothermophilus ortholog revealed that growth could be restored at 78 degrees C by a vector that coexpresses polypeptides corresponding to residues 1-79 and 80-220 of AK(Tn). In contrast, PQN1 growth was not complemented by AK(Tn) fragments harboring a C156A mutation within the zinc-binding tetracysteine motif unless these fragments were fused to Thermotoga maritima chemotaxis proteins that heterodimerize (CheA and CheY) or homodimerize (CheX). This enhanced complementation is interpreted as arising from chemotaxis protein-protein interactions, since AK(Tn)-C156A fragments having only one polypeptide fused to a chemotaxis protein did not complement PQN1 to the same extent. This selection increases the maximum temperature where a PCA can be used to engineer thermostable protein complexes and to map protein-protein interactions.

Matrix photochemical study and conformational analysis of CH3C(O)NCS and CF3C(O)NCS.

PubMed

Ramos, Luis A; Ulic, Sonia E; Romano, Rosana M; Beckers, Helmut; Willner, Helge; Della Védova, Carlos O

2014-01-30

The vapor of acetyl isocyanide, CH3C(O)NCS, and trifluoroacetyl isocyanide, CF3C(O)NCS, were isolated in solid Ar at 15 K. The existence of rotational isomerism was confirmed when the matrixes were irradiated with broad-band UV-vis light (200 ≤ λ ≤ 800 nm) and also by temperature-dependent Ar-matrix IR spectroscopy. The initial spectra showed the vapor of CH3C(O)NCS and CF3C(O)NCS consist of two conformers syn-syn and syn-anti (with the C═O bond syn with respect to the C-H or C-F bond and syn or anti with respect to the N═C double bond). When CH3C(O)NCS is irradiated, simultaneously with the randomization process, H2CCO and HSCN are produced. In the case of the photolysis of CF3C(O)NCS, the main products are CF3NCS and CO. The assignment of the IR bands to the different photoproducts was made on the basis of the usual criteria, taking account reported antecedents in the literature.

ESCRT-mediated Uptake and Degradation of Brain-targeted α-synuclein Single Chain Antibody Attenuates Neuronal Degeneration In Vivo

PubMed Central

Spencer, Brian; Emadi, Sharareh; Desplats, Paula; Eleuteri, Simona; Michael, Sarah; Kosberg, Kori; Shen, Jay; Rockenstein, Edward; Patrick, Christina; Adame, Anthony; Gonzalez, Tania; Sierks, Michael; Masliah, Eliezer

2014-01-01

Parkinson's disease and dementia with Lewy bodies are neurodegenerative disorders characterized by accumulation of α-synuclein (α-syn). Recently, single-chain fragment variables (scFVs) have been developed against individual conformational species of α-syn. Unlike more traditional monoclonal antibodies, these scFVs will not activate or be endocytosed by Fc receptors. For this study, we investigated an scFV directed against oligomeric α-syn fused to the LDL receptor-binding domain from apolipoprotein B (apoB). The modified scFV showed enhanced brain penetration and was imported into neuronal cells through the endosomal sorting complex required for transport (ESCRT) pathway, leading to lysosomal degradation of α-syn aggregates. Further analysis showed that the scFV was effective at ameliorating neurodegenerative pathology and behavioral deficits observed in the mouse model of dementia with Lewy bodies/Parkinson's disease. Thus, the apoB modification had the effect of both increasing accumulation of the scFV in the brain and directing scFV/α-syn complexes for degradation through the ESCRT pathway, leading to improved therapeutic potential of immunotherapy. PMID:25008355

ESCRT-mediated uptake and degradation of brain-targeted α-synuclein single chain antibody attenuates neuronal degeneration in vivo.

PubMed

Spencer, Brian; Emadi, Sharareh; Desplats, Paula; Eleuteri, Simona; Michael, Sarah; Kosberg, Kori; Shen, Jay; Rockenstein, Edward; Patrick, Christina; Adame, Anthony; Gonzalez, Tania; Sierks, Michael; Masliah, Eliezer

2014-10-01

Parkinson's disease and dementia with Lewy bodies are neurodegenerative disorders characterized by accumulation of α-synuclein (α-syn). Recently, single-chain fragment variables (scFVs) have been developed against individual conformational species of α-syn. Unlike more traditional monoclonal antibodies, these scFVs will not activate or be endocytosed by Fc receptors. For this study, we investigated an scFV directed against oligomeric α-syn fused to the LDL receptor-binding domain from apolipoprotein B (apoB). The modified scFV showed enhanced brain penetration and was imported into neuronal cells through the endosomal sorting complex required for transport (ESCRT) pathway, leading to lysosomal degradation of α-syn aggregates. Further analysis showed that the scFV was effective at ameliorating neurodegenerative pathology and behavioral deficits observed in the mouse model of dementia with Lewy bodies/Parkinson's disease. Thus, the apoB modification had the effect of both increasing accumulation of the scFV in the brain and directing scFV/α-syn complexes for degradation through the ESCRT pathway, leading to improved therapeutic potential of immunotherapy.

Curcumin Modulates α-Synuclein Aggregation and Toxicity

PubMed Central

2012-01-01

In human beings, Parkinson’s disease (PD) is associated with the oligomerization and amyloid formation of α-synuclein (α-Syn). The polyphenolic Asian food ingredient curcumin has proven to be effective against a wide range of human diseases including cancers and neurological disorders. While curcumin has been shown to significantly reduce cell toxicity of α-Syn aggregates, its mechanism of action remains unexplored. Here, using a series of biophysical techniques, we demonstrate that curcumin reduces toxicity by binding to preformed oligomers and fibrils and altering their hydrophobic surface exposure. Further, our fluorescence and two-dimensional nuclear magnetic resonance (2D-NMR) data indicate that curcumin does not bind to monomeric α-Syn but binds specifically to oligomeric intermediates. The degree of curcumin binding correlates with the extent of α-Syn oligomerization, suggesting that the ordered structure of protein is required for effective curcumin binding. The acceleration of aggregation by curcumin may decrease the population of toxic oligomeric intermediates of α-Syn. Collectively; our results suggest that curcumin and related polyphenolic compounds can be pursued as candidate drug targets for treatment of PD and other neurological diseases. PMID:23509976

NMR and rotational angles in solution conformation of polypeptides

NASA Astrophysics Data System (ADS)

Bystrov, V. F.

1985-01-01

Professor San-Ichiro Mizushima and Professor Yonezo Morino's classical contributions provided unique means and firm basis for understanding of conformational states and internal rotation in polypeptide molecules. Now the NMR spectroscopy is the best choice to study molecular conformation, mechanism of action and structure-functional relationships of peptide and proteins in solution under conditions approaching those of their physiological environments. Crucial details of spatial structure and interactions of these molecules in solution are revealed by using proton-proton and carbon-proton vicinal coupling constants, proton nuclear Overhauser effect and spectral perturbation techniques. The results of NMR conformational analysis are presented for valinomycin "bracelet", gramicidin A double helices, honey-bee neurotoxin apamin, scorpion insectotoxins and snake neurotoxins of long and short types.

Breakout character of islet amyloid polypeptide hydrophobic mutations at the onset of type-2 diabetes

NASA Astrophysics Data System (ADS)

Frigori, Rafael B.

2014-11-01

Toxic fibrillar aggregates of islet amyloid polypeptide (IAPP) appear as the physical outcome of a peptidic phase transition signaling the onset of type-2 diabetes mellitus in different mammalian species. In particular, experimentally verified mutations on the amyloidogenic segment 20-29 in humans, cats, and rats are highly correlated with the molecular aggregation propensities. Through a microcanonical analysis of the aggregation of IAPP20 -29 isoforms, we show that a minimalist one-bead hydrophobic-polar continuum model for protein interactions properly quantifies those propensities from free-energy barriers. Our results highlight the central role of sequence-dependent hydrophobic mutations on hot spots for stabilization, and thus for the engineering, of such biological peptides.

Deep functional analysis of synII, a 770 kb synthetic yeast chromosome

PubMed Central

Gao, Feng; Gong, Jianhui; Abramczyk, Dariusz; Walker, Roy; Zhao, Hongcui; Chen, Shihong; Liu, Wei; Luo, Yisha; Müller, Carolin A.; Paul-Dubois-Taine, Adrien; Alver, Bonnie; Stracquadanio, Giovanni; Mitchell, Leslie A.; Luo, Zhouqing; Fan, Yanqun; Zhou, Baojin; Wen, Bo; Tan, Fengji; Wang, Yujia; Zi, Jin; Xie, Zexiong; Li, Bingzhi; Yang, Kun; Richardson, Sarah M.; Jiang, Hui; French, Christopher E.; Nieduszynski, Conrad A.; Koszul, Romain; Marston, Adele L.; Yuan, Yingjin; Wang, Jian; Bader, Joel S.; Dai, Junbiao; Boeke, Jef D.; Xu, Xun; Cai, Yizhi; Yang, Huanming

2017-01-01

Herein we report the successful design, construction and characterization of a 770 kb synthetic yeast chromosome II (synII). Our study incorporates characterization at multiple levels, including phenomics, transcriptomics, proteomics, chromosome segregation and replication analysis to provide a thorough and comprehensive analysis of a synthetic chromosome. Our “Trans-Omics” analyses reveal a modest but potentially significant pervasive up-regulation of translational machinery observed in synII is mainly caused by the deletion of 13 tRNAs. By both complementation assays and SCRaMbLE, we targeted and debuged the origin of a growth defect at 37°C in glycerol medium, which is related to misregulation of the HOG response. Despite the subtle differences, the synII strain shows highly consistent biological processes comparable to the native strain. PMID:28280153

Analysis of polypeptide composition and antigenic components of Rickettsia tsutsugamushi by polyacrylamide gel electrophoresis and immunoblotting.

PubMed Central

Tamura, A; Ohashi, N; Urakami, H; Takahashi, K; Oyanagi, M

1985-01-01

Polyacrylamide gel electrophoresis of lysates of purified Rickettsia tsutsugamushi revealed as many as 30 polypeptide bands, including major bands corresponding to molecular sizes of 70, 60, 54 to 56, and 46 to 47 kilodaltons. Compared with the polypeptide composition of the rickettsiae of Gilliam, Karp, and Kato strains and a newly isolated Shimokoshi strain, the major polypeptide in the Kato strain (54-56K) and in the Karp strain (46-47K) migrated a little faster and slower, respectively, than the corresponding polypeptides in the other strains. The largest major polypeptide (54-56K) was digestible by the treatment of intact rickettsiae with trypsin and variable in content in separate preparations, suggesting that the polypeptide exists on the rickettsial surface and is easily degraded during the handling of these microorganisms. Several surface polypeptides of rickettsiae, including the 54-56K and 46-47K polypeptides, were detected by radioiodination of intact rickettsiae followed by polyacrylamide gel electrophoresis of the lysate; however, the 70K and 60K polypeptides were not labeled. Immunoblotting experiments with hyperimmune sera prepared in guinea pigs against each strain demonstrated that the 70K, 54-56K, and 46-47K polypeptides showed antigenic activities. The 54-56K polypeptide appeared to be strain specific, whereas the 70K and 46-47K polypeptides cross-reacted with the heterologous antisera. Images PMID:3922893

Polypeptide synthesis induced in Nicotiana clevelandii protoplasts by infection with raspberry ringspot nepovirus.

PubMed

Acosta, O; Mayo, M A

1993-01-01

Infection of Nicotiana clevelandii protoplasts by raspberry ringspot nepovirus resulted in the accumulation of about 24 polypeptides that differed in M(r) and pI from polypeptides accumulating in mock-inoculated protoplasts. Similar polypeptides accumulated in protoplasts infected with the S and E strains of RRV but different infection-specific polypeptides were detected in protoplasts infected with tobacco ringspot nepovirus. The M(r) of RRV-specific polypeptides ranged from 210,000 to 18,000 and most are presumed to be derived from others by proteolytic cleavage. No evidence was found for marked changes in polypeptide abundance with time after inoculation or for any virus-specific polypeptide becoming disproportionately abundant in the medium during culture.

Monte Carlo simulations of the counter ion effect on the conformational equilibrium of the N, N'-diphenyl-guanidinium ion in aqueous solution

NASA Astrophysics Data System (ADS)

Nagy, Peter I.; Durant, Graham J.

1996-01-01

Results of calculations for the equilibrium of the syn-syn, anti-syn, and anti-anti conformers of the N, N'-diphenyl-guanidinium ion in aqueous solution are sensitive to whether a counter ion is considered. Relative internal free energies were calculated upon MP2/6-31G*//HF/4-31G energies (second order Møller-Plesset energies obtained when using the 6-31G* basis set at geometries optimized at the Hartree-Fock level and using the 4-31G basis set) and relative solvation free energy terms were obtained by Monte Carlo simulations. Without considering a counter ion only a small fraction of the solute has been predicted to adopt the anti-anti conformation in the solution. When considering acetate and chloride counter ions with salt concentration of 0.11 mol/l at 310 K, mimicking physiological conditions, the counter ion close to the cation stabilizes the anti-anti form significantly. Though there are not local free energy minima for the present systems with close counter ions because of the relatively weak ion-ion interaction due to the largely delocalized total charge and atomic charge alternation in the cation, the constraint for the C(guanidinium)...C(carboxylate) separation of 4.6 Å allows an insight into the arginine...aspartate or glutamate interactions commonly found in peptides. The N-H(guanidinium)...O(carboxylate) hydrogen bonds are not stable due to thermal motion in aqueous solution. The neighboring water molecules, however, move into the space in-between the charged groups and comprise a hydrogen bonded network. Interactions with a chloride counter ion may be significant for the drug delivery process to the receptor site. Close contact between the N, N'-diphenyl guanidinium and a chloride ion is also not favored, though it may occur temporarily and then would favor the anti-anti conformer. Deviation from the relative solvation free energy obtained for the conformational change of the single cation is still some tenths of a kcal/mol with ions separated as much as 12.4 Å. While solvation energetics is affected even at such a separation, solution structure around the ions can be basically characterized without considering the effect of a remote counterpart.

Orpinomyces cellulase CelE protein and coding sequences

DOEpatents

Li, Xin-Liang; Ljungdahl, Lars G.; Chen, Huizhong

2000-08-29

A CDNA designated celE cloned from Orpinomyces PC-2 encodes a polypeptide (CelE) of 477 amino acids. CelE is highly homologous to CelB of Orpinomyces (72.3% identity) and Neocallimastix (67.9% identity), and like them, it has a non-catalytic repeated peptide domain (NCRPD) at the C-terminal end. The catalytic domain of CelE is homologous to glycosyl hydrolases of Family 5, found in several anaerobic bacteria. The gene of celE is devoid of introns. The recombinant proteins CelE and CelB of Orpinomyces PC-2 randomly hydrolyze carboxymethylcellulose and cello-oligosaccharides in the pattern of endoglucanases.

Inhibition of secretin stimulated pancreatic secretion by pancreatic polypeptide.

PubMed

Adrian, T E; Besterman, H S; Mallinson, C N; Greenberg, G R; Bloom, S R

1979-01-01

The effect of PP on secretin-stimulated pancreatic secretion was assessed in five healthy subjects. During an intravenous infusion of BPP at a dose which produced plasma levels similar to those seen after meals in healthy young adults the volume and bicarbonate content of duodenal juice was reduced by 25% (p less than 0.05) and 24% (p less than 0.05) respectively, while protein and bilirubin concentrations were more markedly reduced by 68% (p less than 0.0005) and 67% (p less than 0.0005) respectively. PP, thus, may be an important inhibitory factor in the control of bilirubin and pancreatic enzyme secretion in man.

Inhibition of secretin stimulated pancreatic secretion by pancreatic polypeptide.

PubMed Central

Adrian, T E; Besterman, H S; Mallinson, C N; Greenberg, G R; Bloom, S R

1979-01-01

The effect of PP on secretin-stimulated pancreatic secretion was assessed in five healthy subjects. During an intravenous infusion of BPP at a dose which produced plasma levels similar to those seen after meals in healthy young adults the volume and bicarbonate content of duodenal juice was reduced by 25% (p less than 0.05) and 24% (p less than 0.05) respectively, while protein and bilirubin concentrations were more markedly reduced by 68% (p less than 0.0005) and 67% (p less than 0.0005) respectively. PP, thus, may be an important inhibitory factor in the control of bilirubin and pancreatic enzyme secretion in man. PMID:761835

Polypeptides having laccase activity and polynucleotides encoding same

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Ye; Tang, Lan; Duan, Junxin

The present invention relates to isolated polypeptides having laccase activity and polynucleotides encoding the polypeptides and polynucleotides encoding the polypeptides. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.

Theoretical Modeling of Molecular and Electron Kinetic Processes. Volume I. Theoretical Formulation of Analysis and Description of Computer Program.

DTIC Science & Technology

1979-01-01

syn- thesis proceed s by ignoring unacceptable syntax or other errors , pro- tection against subsequent execution of a faulty reaction scheme can be...resulting TAPE9 . During subroutine syn thesis and reaction processing, a search is made (fo r each secondary electron collision encountered) to...program library, which can be cat- alogued and saved if any future specialized modifications (beyond the scope of the syn thesis capability of LASER

Evidence for large-scale imbrication during Eocene syn-orogenic exhumation of the Hellenic subduction channel (Cyclades, Greece)

NASA Astrophysics Data System (ADS)

Grasemann, Bernhard; Huet, Benjamin; Schneider, David; Rice, Hugh; Lemonnier, Nicolas; Tschegg, Cornelius

2017-04-01

In the Cyclades, Miocene post-orogenic back-arc extension overprinted the exhumed syn- orogenic Eocene subduction channel. Whereas the exact geometry and kinematics of the syn-orogenic exhumation are still controversial, but must have involved a floor thrust and an apparent normal fault at the roof, the post-orogenic extension, leading to the exhumation of Cordilleran-type metamorphic core complexes, is well constrained by several major detachment systems. On the island of Milos, which is part of the South Aegean Volcanic Arc, minor outcrops of schist occur. New data indicate that these witnessed Eocene blueschist facies metamorphism at 8.5 kbar and 400°C, but escaped the Miocene extensional overprint, as they lie in the hanging wall of the West Cycladic Detachment System. In contrast, eclogite pebbles in "Green Lahars" on Milos yield metamorphic conditions of 19.5 kbar at 550°C. Both high-pressure units belong to the Cycladic Blueschist Unit and can only have been juxtaposed by thrusting. This indicates that two nappes, the newly defined Cycladic Blueschist Nappe and the overlying Cycladic Eclogite Nappe, both comprising rocks of the Cycladic Blueschist Unit, exist on Milos. These nappes probably also form the other Cycladic islands, separated by a syn-orogenic thrust, which we name the Trans Cycladic Thrust. The Trans Cycladic Thrust, which traces the orientation of the syn-orogenic exhumation channel, is partly offset by the post-orogenic Miocene extensional detachment systems. As a result of the Mid- to Late Miocene clockwise crustal block rotation, the syn-orogenic channel, and hence the Trans Cycladic Thrust, bends through 90° at Milos, changing from a W-E trending to a N-S trending extrusion-related stretching lineation. Restoration of the Miocene block-rotation and extension results in syn-orogenic thrusting kinematics (top-SSW) in the Cycladic Blueschist Nappe and along the Trans Cycladic Thrust and syn-orogenic apparent normal faulting kinematics (top-NNE) at the roof of the Cycladic Eclogite Nappe, consistent with the Eocene extrusion of the high-pressure rocks in the Cyclades.

Dosimetric and workflow evaluation of first commercial synthetic CT software for clinical use in pelvis

NASA Astrophysics Data System (ADS)

Tyagi, Neelam; Fontenla, Sandra; Zhang, Jing; Cloutier, Michelle; Kadbi, Mo; Mechalakos, Jim; Zelefsky, Michael; Deasy, Joe; Hunt, Margie

2017-04-01

To evaluate a commercial synthetic CT (syn-CT) software for use in prostate radiotherapy. Twenty-five prostate patients underwent CT and MR simulation scans in treatment position on a 3T MR scanner. A commercially available MR protocol was used that included a T2w turbo spin-echo sequence for soft-tissue contrast and a dual echo 3D mDIXON fast field echo (FFE) sequence for generating syn-CT. A dual-echo 3D FFE B 0 map was used for patient-induced susceptibility distortion analysis and a new 3D balanced-FFE sequence was evaluated for identification of implanted gold fiducial markers and subsequent image-guidance during radiotherapy delivery. Tissues were classified as air, adipose, water, trabecular/spongy bone and compact/cortical bone and assigned bulk HU values. The accuracy of syn-CT for treatment planning was analyzed by transferring the structures and plan from planning CT to syn-CT and recalculating the dose. Accuracy of localization at the treatment machine was evaluated by comparing registration of kV radiographs to either digitally reconstructed radiographs (DRRs) generated from syn-CT or traditional DRRs generated from the planning CT. Similarly, accuracy of setup using CBCT and syn-CT was compared to that using the planning CT. Finally, a MR-only simulation workflow was established and end-to-end testing was completed on five patients undergoing MR-only simulation. Dosimetric comparison between the original CT and syn-CT plans was within 0.5% on average for all structures. The de-novo optimized plans on the syn-CT met institutional clinical objectives for target and normal structures. Patient-induced susceptibility distortion based on B 0 maps was within 1 mm and 0.5 mm in the body and prostate respectively. DRR and CBCT localization based on MR-localized fiducials showed a standard deviation of  <1 mm. End-to-end testing and MR simulation workflow was successfully validated. MRI derived synthetic CT can be successfully used for a MR-only planning and treatment for prostate radiotherapy.

Nomenclatural studies toward a world catalog of Diptera genus-group names. III. Christian Rudolph Wilhelm Wiedemann.

PubMed

Evenhuis, Neal L; Pont, Adrian C

2013-01-01

The Diptera genus-group names of Christian Rudolph Wilhelm Wiedemann are reviewed and annotated. A total of 50 available genus-group names in 25 families of Diptera are listed alphabetically for each name giving author, year and page of original publication, originally included species, type species and method of fixation, current status of the name, family placement, and a list of any emendations of it that have been found in the literature. Remarks are given to clarify nomenclatural or taxonomic information. A biography of Wiedemann is given with discussion of his works and his relationships with contemporaries. In addition, an index is given to all the species-group names of Diptera proposed by Wiedemann (1,775 of which 1,698 are available) with bibliographic reference to each original citation. An appendix gives a complete bibliography of all the known writings by Wiedemann, non-zoological as well as zoological.The following type species is designated herein: Eristalis chrysopygus Wiedemann, 1819 for Pachycephalus Wiedemann, 1830, by present designation [Syrphidae].Corrected or clarified type-species and methods of typification are given for: Colax Wiedemann, 1824 [Nemestrinidae]; Cyphomyia Wiedemann, 1819 [Stratiomyidae]; Philoliche Wiedemann, 1821 [Tabanidae]; Ropalomera Wiedemann, 1820 [Ropalomeridae]; Timia Wiedemann, 1824 [Ulidiidae].Acting as First Reviser, the following correct original spelling for multiple original spellings is selected: Maekistocera Wiedemann, 1820 [Tipulidae]. A previous First Reviser action for multiple original spellings missed by other workers is given for the following: Rhaphiorhynchus Wiedemann, 1821 [Pantophthalmidae].The following nominal genera enter into new synonymies: Ceratophyia Osten Sacken, 1858 of Ceratophya Wiedemann, 1824, n. syn. [Syrphidae]; Epopter Wiedemann, 1830 of Sphecomyia Le Peletier & Serville, 1825, n. syn. [Syrphidae]; Melophaga Wiedemann, 1830 of Melophagus Latreille, 1802, n. syn. [Hippoboscidae]; Midas Latreille, 1797 of Mydas Fabricius, 1794, n. syn. [Mydidae]; Nemestrina Latreille, 1809 of Nemestrinus Latreille, 1802, n. syn. [Nemestrinidae]; Pangonia Latreille, 1809 of Pangonius Latreille, 1802, n. syn. [Tabanidae]; Scatophaga Wiedemann, 1828 of Scathophaga Meigen, 1803, n. syn. [Scathophagidae]; Threneste Wiedemann, 1830 of Penthetria Meigen, 1803, n. syn. [Bibionidae].

The visible touch: in planta visualization of protein-protein interactions by fluorophore-based methods

PubMed Central

Bhat, Riyaz A; Lahaye, Thomas; Panstruga, Ralph

2006-01-01

Non-invasive fluorophore-based protein interaction assays like fluorescence resonance energy transfer (FRET) and bimolecular fluorescence complementation (BiFC, also referred to as "split YFP") have been proven invaluable tools to study protein-protein interactions in living cells. Both methods are now frequently used in the plant sciences and are likely to develop into standard techniques for the identification, verification and in-depth analysis of polypeptide interactions. In this review, we address the individual strengths and weaknesses of both approaches and provide an outlook about new directions and possible future developments for both techniques. PMID:16800872

Family-group names in Coleoptera (Insecta)

PubMed Central

Bouchard, Patrice; Bousquet, Yves; Davies, Anthony E.; Alonso-Zarazaga, Miguel A.; Lawrence, John F.; Lyal, Chris H. C.; Newton, Alfred F.; Reid, Chris A. M.; Schmitt, Michael; Ślipiński, S. Adam; Smith, Andrew B. T.

2011-01-01

Abstract We synthesize data on all known extant and fossil Coleoptera family-group names for the first time. A catalogue of 4887 family-group names (124 fossil, 4763 extant) based on 4707 distinct genera in Coleoptera is given. A total of 4492 names are available, 183 of which are permanently invalid because they are based on a preoccupied or a suppressed type genus. Names are listed in a classification framework. We recognize as valid 24 superfamilies, 211 families, 541 subfamilies, 1663 tribes and 740 subtribes. For each name, the original spelling, author, year of publication, page number, correct stem and type genus are included. The original spelling and availability of each name were checked from primary literature. A list of necessary changes due to Priority and Homonymy problems, and actions taken, is given. Current usage of names was conserved, whenever possible, to promote stability of the classification. New synonymies (family-group names followed by genus-group names): Agronomina Gistel, 1848 syn. nov. of Amarina Zimmermann, 1832 (Carabidae), Hylepnigalioini Gistel, 1856 syn. nov. of Melandryini Leach, 1815 (Melandryidae), Polycystophoridae Gistel, 1856 syn. nov. of Malachiinae Fleming, 1821 (Melyridae), Sclerasteinae Gistel, 1856 syn. nov. of Ptilininae Shuckard, 1839 (Ptinidae), Phloeonomini Ádám, 2001 syn. nov. of Omaliini MacLeay, 1825 (Staphylinidae), Sepedophilini Ádám, 2001 syn. nov. of Tachyporini MacLeay, 1825 (Staphylinidae), Phibalini Gistel, 1856 syn. nov. of Cteniopodini Solier, 1835 (Tenebrionidae); Agronoma Gistel 1848 (type species Carabus familiaris Duftschmid, 1812, designated herein) syn. nov. of Amara Bonelli, 1810 (Carabidae), Hylepnigalio Gistel, 1856 (type species Chrysomela caraboides Linnaeus, 1760, by monotypy) syn. nov. of Melandrya Fabricius, 1801 (Melandryidae), Polycystophorus Gistel, 1856 (type species Cantharis aeneus Linnaeus, 1758, designated herein) syn. nov. of Malachius Fabricius, 1775 (Melyridae), Sclerastes Gistel, 1856 (type species Ptilinus costatus Gyllenhal, 1827, designated herein) syn. nov. of Ptilinus Geoffroy, 1762 (Ptinidae), Paniscus Gistel, 1848 (type species Scarabaeus fasciatus Linnaeus, 1758, designated herein) syn. nov. of Trichius Fabricius, 1775 (Scarabaeidae), Phibalus Gistel, 1856 (type species Chrysomela pubescens Linnaeus, 1758, by monotypy) syn. nov. of Omophlus Dejean, 1834 (Tenebrionidae). The following new replacement name is proposed: Gompeliina Bouchard, 2011 nom. nov. for Olotelina Báguena Corella, 1948 (Aderidae). Reversal of Precedence (Article 23.9) is used to conserve usage of the following names (family-group names followed by genus-group names): Perigonini Horn, 1881 nom. protectum over Trechicini Bates, 1873 nom. oblitum (Carabidae), Anisodactylina Lacordaire, 1854 nom. protectum over Eurytrichina LeConte, 1848 nom. oblitum (Carabidae), Smicronychini Seidlitz, 1891 nom. protectum over Desmorini LeConte, 1876 nom. oblitum (Curculionidae), Bagoinae Thomson, 1859 nom. protectum over Lyprinae Gistel 1848 nom. oblitum (Curculionidae), Aterpina Lacordaire, 1863 nom. protectum over Heliomenina Gistel, 1848 nom. oblitum (Curculionidae), Naupactini Gistel, 1848 nom. protectum over Iphiini Schönherr, 1823 nom. oblitum (Curculionidae), Cleonini Schönherr, 1826 nom. protectum over Geomorini Schönherr, 1823 nom. oblitum (Curculionidae), Magdalidini Pascoe, 1870 nom. protectum over Scardamyctini Gistel, 1848 nom. oblitum (Curculionidae), Agrypninae/-ini Candèze, 1857 nom. protecta over Adelocerinae/-ini Gistel, 1848 nom. oblita and Pangaurinae/-ini Gistel, 1856 nom. oblita (Elateridae), Prosternini Gistel, 1856 nom. protectum over Diacanthini Gistel, 1848 nom. oblitum (Elateridae), Calopodinae Costa, 1852 nom. protectum over Sparedrinae Gistel, 1848 nom. oblitum (Oedemeridae), Adesmiini Lacordaire, 1859 nom. protectum over Macropodini Agassiz, 1846 nom. oblitum (Tenebrionidae), Bolitophagini Kirby, 1837 nom. protectum over Eledonini Billberg, 1820 nom. oblitum (Tenebrionidae), Throscidae Laporte, 1840 nom. protectum over Stereolidae Rafinesque, 1815 nom. oblitum (Throscidae) and Lophocaterini Crowson, 1964 over Lycoptini Casey, 1890 nom. oblitum (Trogossitidae); Monotoma Herbst, 1799 nom. protectum over Monotoma Panzer, 1792 nom. oblitum (Monotomidae); Pediacus Shuckard, 1839 nom. protectum over Biophloeus Dejean, 1835 nom. oblitum (Cucujidae), Pachypus Dejean, 1821 nom. protectum over Pachypus Billberg, 1820 nom. oblitum (Scarabaeidae), Sparrmannia Laporte, 1840 nom. protectum over Leocaeta Dejean, 1833 nom. oblitum and Cephalotrichia Hope, 1837 nom. oblitum (Scarabaeidae). PMID:21594053

Key aromatic/hydrophobic amino acids controlling a cross-amyloid peptide interaction versus amyloid self-assembly.

PubMed

Bakou, Maria; Hille, Kathleen; Kracklauer, Michael; Spanopoulou, Anna; Frost, Christina V; Malideli, Eleni; Yan, Li-Mei; Caporale, Andrea; Zacharias, Martin; Kapurniotu, Aphrodite

2017-09-01

The interaction of the intrinsically disordered polypeptide islet amyloid polypeptide (IAPP), which is associated with type 2 diabetes (T2D), with the Alzheimer's disease amyloid-β (Aβ) peptide modulates their self-assembly into amyloid fibrils and may link the pathogeneses of these two cell-degenerative diseases. However, the molecular determinants of this interaction remain elusive. Using a systematic alanine scan approach, fluorescence spectroscopy, and other biophysical methods, including heterocomplex pulldown assays, far-UV CD spectroscopy, the thioflavin T binding assay, transmission EM, and molecular dynamics simulations, here we identified single aromatic/hydrophobic residues within the amyloid core IAPP region as hot spots or key residues of its cross-interaction with Aβ40(42) peptide. Importantly, we also find that none of these residues in isolation plays a key role in IAPP self-assembly, whereas simultaneous substitution of four aromatic/hydrophobic residues with Ala dramatically impairs both IAPP self-assembly and hetero-assembly with Aβ40(42). Furthermore, our experiments yielded several novel IAPP analogs, whose sequences are highly similar to that of IAPP but have distinct amyloid self- or cross-interaction potentials. The identified similarities and major differences controlling IAPP cross-peptide interaction with Aβ40(42) versus its amyloid self-assembly offer a molecular basis for understanding the underlying mechanisms. We propose that these insights will aid in designing intervention strategies and novel IAPP analogs for the management of type 2 diabetes, Alzheimer's disease, or other diseases related to IAPP dysfunction or cross-amyloid interactions. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Candide: An Interactive System for the Acquisition of Domain Specific Knowledge

DTIC Science & Technology

1988-05-01

wats coiirlltely’ overhlauiled duin g lie termi of thnis project. As a result it can directly use miorphlological rille., iii stead iif, precorri oiled...November). [Bresnan, 83] Bresnan, Joan, 1983: The mental representation of g . mmatical relations (ed.), Cambridge: MIT Press. [Gazdar and Pullum, 82] Gazdar... G . and G.K. Pullum, 1982: "GPSG: A l’heoretical Syn- opsis," Indiana University Linguistics Club, Bloomington, Indiana. * [Grosz, el al., 82] Grosz

Segmental allotetraploidy and allelic interactions in buffelgrass (Pennisetum ciliare (L.) Link syn. Cenchrus ciliaris L.) as revealed by genome mapping.

PubMed

Jessup, R W; Burson, B L; Burow, O; Wang, Y W; Chang, C; Li, Z; Paterson, A H; Hussey, M A

2003-04-01

Linkage analyses increasingly complement cytological and traditional plant breeding techniques by providing valuable information regarding genome organization and transmission genetics of complex polyploid species. This study reports a genome map of buffelgrass (Pennisetum ciliare (L.) Link syn. Cenchrus ciliaris L.). Maternal and paternal maps were constructed with restriction fragment length polymorphisms (RFLPs) segregating in 87 F1 progeny from an intraspecific cross between two heterozygous genotypes. A survey of 862 heterologous cDNAs and gDNAs from across the Poaceae, as well as 443 buffelgrass cDNAs, yielded 100 and 360 polymorphic probes, respectively. The maternal map included 322 RFLPs, 47 linkage groups, and 3464 cM, whereas the paternal map contained 245 RFLPs, 42 linkage groups, and 2757 cM. Approximately 70 to 80% of the buffelgrass genome was covered, and the average marker spacing was 10.8 and 11.3 cM on the respective maps. Preferential pairing was indicated between many linkage groups, which supports cytological reports that buffelgrass is a segmental allotetraploid. More preferential pairing (disomy) was found in the maternal than paternal parent across linkage groups (55 vs. 38%) and loci (48 vs. 15%). Comparison of interval lengths in 15 allelic bridges indicated significantly less meiotic recombination in paternal gametes. Allelic interactions were detected in four regions of the maternal map and were absent in the paternal map.

Acemetacin cocrystal structures by powder X-ray diffraction.

PubMed

Bolla, Geetha; Chernyshev, Vladimir; Nangia, Ashwini

2017-05-01

Cocrystals of acemetacin drug (ACM) with nicotinamide (NAM), p -aminobenzoic acid (PABA), valerolactam (VLM) and 2-pyridone (2HP) were prepared by melt crystallization and their X-ray crystal structures determined by high-resolution powder X-ray diffraction. The powerful technique of structure determination from powder data (SDPD) provided details of molecular packing and hydrogen bonding in pharmaceutical cocrystals of acemetacin. ACM-NAM occurs in anhydrate and hydrate forms, whereas the other structures crystallized in a single crystalline form. The carboxylic acid group of ACM forms theacid-amide dimer three-point synthon R 3 2 (9) R 2 2 (8) R 3 2 (9) with three different syn amides (VLM, 2HP and caprolactam). The conformations of the ACM molecule observed in the crystal structures differ mainly in the mutual orientation of chlorobenzene fragment and the neighboring methyl group, being anti (type I) or syn (type II). ACM hydrate, ACM-NAM, ACM-NAM-hydrate and the piperazine salt of ACM exhibit the type I conformation, whereas ACM polymorphs and other cocrystals adopt the ACM type II conformation. Hydrogen-bond interactions in all the crystal structures were quantified by calculating their molecular electrostatic potential (MEP) surfaces. Hirshfeld surface analysis of the cocrystal surfaces shows that about 50% of the contribution is due to a combination of strong and weak O⋯H, N⋯H, Cl⋯H and C⋯H interactions. The physicochemical properties of these cocrystals are under study.

Acemetacin cocrystal structures by powder X-ray diffraction

PubMed Central

Bolla, Geetha

2017-01-01

Cocrystals of acemetacin drug (ACM) with nicotinamide (NAM), p-aminobenzoic acid (PABA), valerolactam (VLM) and 2-pyridone (2HP) were prepared by melt crystallization and their X-ray crystal structures determined by high-resolution powder X-ray diffraction. The powerful technique of structure determination from powder data (SDPD) provided details of molecular packing and hydrogen bonding in pharmaceutical cocrystals of acemetacin. ACM–NAM occurs in anhydrate and hydrate forms, whereas the other structures crystallized in a single crystalline form. The carboxylic acid group of ACM forms theacid–amide dimer three-point synthon R 3 2(9)R 2 2(8)R 3 2(9) with three different syn amides (VLM, 2HP and caprolactam). The conformations of the ACM molecule observed in the crystal structures differ mainly in the mutual orientation of chlorobenzene fragment and the neighboring methyl group, being anti (type I) or syn (type II). ACM hydrate, ACM—NAM, ACM–NAM-hydrate and the piperazine salt of ACM exhibit the type I conformation, whereas ACM polymorphs and other cocrystals adopt the ACM type II conformation. Hydrogen-bond interactions in all the crystal structures were quantified by calculating their molecular electrostatic potential (MEP) surfaces. Hirshfeld surface analysis of the cocrystal surfaces shows that about 50% of the contribution is due to a combination of strong and weak O⋯H, N⋯H, Cl⋯H and C⋯H interactions. The physicochemical properties of these cocrystals are under study. PMID:28512568

Polypeptide having an amino acid replaced with N-benzylglycine

DOEpatents

Mitchell, Alexander R.; Young, Janis D.

1996-01-01

The present invention relates to one or more polypeptides having useful biological activity in a mammal, which comprise: a polypeptide related to bradykinin of four to ten amino acid residues wherein one or more specific amino acids in the polypeptide chain are replaced with achiral N-benzylglycine. These polypeptide analogues have useful potent agonist or antagonist pharmacological properties depending upon the structure. A preferred polypeptide is (N-benzylglycine.sup.7)-bradykinin.

Atomistic Picture for the Folding Pathway of a Hybrid-1 Type Human Telomeric DNA G-quadruplex

PubMed Central

Bian, Yunqiang; Tan, Cheng; Wang, Jun; Sheng, Yuebiao; Zhang, Jian; Wang, Wei

2014-01-01

In this work we studied the folding process of the hybrid-1 type human telomeric DNA G-quadruplex with solvent and ions explicitly modeled. Enabled by the powerful bias-exchange metadynamics and large-scale conventional molecular dynamic simulations, the free energy landscape of this G-DNA was obtained for the first time and four folding intermediates were identified, including a triplex and a basically formed quadruplex. The simulations also provided atomistic pictures for the structures and cation binding patterns of the intermediates. The results showed that the structure formation and cation binding are cooperative and mutually supporting each other. The syn/anti reorientation dynamics of the intermediates was also investigated. It was found that the nucleotides usually take correct syn/anti configurations when they form native and stable hydrogen bonds with the others, while fluctuating between two configurations when they do not. Misfolded intermediates with wrong syn/anti configurations were observed in the early intermediates but not in the later ones. Based on the simulations, we also discussed the roles of the non-native interactions. Besides, the formation process of the parallel conformation in the first two G-repeats and the associated reversal loop were studied. Based on the above results, we proposed a folding pathway for the hybrid-1 type G-quadruplex with atomistic details, which is new and more complete compared with previous ones. The knowledge gained for this type of G-DNA may provide a general insight for the folding of the other G-quadruplexes. PMID:24722458

Comparative testing of HPV L1 protein monoclonal antibody panel for the detection of HPV in cervical exfoliated cells.

PubMed

Wang, Le; Hu, Yu-Chang; Xiao, Chang-Yi; Wang, Fei; Liu, Yu-Fei; Tang, Li-Hua; Xiao, Rong-Shuang

2018-07-01

To determine the value of a monoclonal antibody panel against a C-terminal conserved sequence polypeptide of human papillomavirus (HPV) L1 (a major capsid protein) for the detection of HPV in cervical exfoliated cells, as well as the potential of this antibody panel to be developed into an assay kit for the clinical screening of cervical cancer. Cervical exfoliated cells were collected at a gynecology clinic. One part of each sample was sent to the Department of Pathology for HPV genotyping, and the other part was sent to the Department of Pathology for cytologic testing and then to the laboratory for immunological histological chemistry (IHC) assay in which an HPV L1 C-terminal conserved sequence polypeptide-induced mouse monoclonal antibody panel was used to detect HPV L1. Cervical cell samples were collected from 514 patients at the gynecology clinic; of these, 339 samples were sent for HPV genotyping, and 220 were HPV positive (64.90%, 220/339). Moreover, the duplicate samples from these 339 patients were sent for IHC assay, and 229 samples were positive (67.55%, 229/339). The IHC result was concordant with that obtained by HPV genotyping (Kappa = 0.743, p < 0.001). This study showed that use of the HPV L1 C-terminal conserved sequence polypeptide-induced mouse monoclonal antibody panel was of great value for the detection of HPV in cervical cells; the resulting detection rate was comparable to that obtained using the commercial HPV genotyping kit that is currently in use in clinical practice. Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.

The two subunits of the human asialoglycoprotein receptor have different fates when expressed alone in fibroblasts

PubMed Central

Shia, Michael A.; Lodish, Harvey F.

1989-01-01

Two related polypeptides, H1 and H2, comprise the human asialoglycoprotein receptor (ASGP-R). Stable lines of murine NIH 3T3 fibroblasts expressing H1 alone or H2 alone do not bind or internalize the ligand asialoorosomucoid (ASOR), which contains triantennary oligosaccharides. In contrast, cells expressing H1 and H2 together bind and degrade ASOR with properties indistinguishable from those of the ASPG-R in human hepatoma HepG2 cells. Whether or not H2 is coexpressed, H1 is synthesized as a 40-kDa precursor bearing high-mannose oligosaccharides, processed to its mature 46-kDa form, and transported to the cell surface. In cells expressing only H1, homodimers and -trimers of H1 are formed. In contrast, when expressed in 3T3 cells without H1, H2 is synthesized as its 43-kDa precursor, bearing high-mannose oligosaccharides, but is rapidly degraded. When H1 and H2 are coexpressed in the same cell, the H1 polypeptide “rescues” the H2 polypeptide; H2 is processed to its characteristic 50-kDa mature form and is transported to the surface. We conclude that the human ASGP-R is a multichain heterooligomer, probably a trimer of H1 molecules in noncovalent association with one, two, or three H2 molecules, and that the two polypeptides normally interact early in biosynthesis. Images PMID:2919187

Randomized controlled trial of Syn-Ergel and an active placebo in the treatment of heartburn of pregnancy.

PubMed

Shaw, R W

1978-01-01

A randomized controlled trial was performed to study the efficacy of Syn-Ergel with an active placebo in the treatment of heartburn of pregnancy in ninety-two patients completing 7 days of therapy. Syn-Ergel was significantly better (p less than 0.001) in all groups of pre-treatment pain severity in relieving the symptoms, and had a longer duration of action, than the active placebo. Complete relief of pain was achieved in 79.5% of Syn-Ergel treatments with a further 10% of treatments resulting in marked easing of discomfort at 1 hour following administration. The corresponding figures for the 'active placebo' were 56% and 20%. The combination of an antacid and a protective mucosal coating agent would appear to be a useful approach in the treatment of heartburn of pregnacy.

Configurational and conformational preferences in oximes and oxime carbanions. Ab initio study of the syn effect in reactions of oxyimine enolate equivalents

DOE Office of Scientific and Technical Information (OSTI.GOV)

Glaser, R.; Streitwieser, A.

1989-09-13

Geometries and relative energies of stationary structures of several conformers of geometrical isomers of NO s-trans-configured acetaldoxime are reported. The calculated energies and geometries agree well with comparable experimental data. Effects of the theoretical model on the NO band lengths are discussed for formaldoxime. The theoretical results suggest that the regiochemistry of enolate equivalents of oxyimines in dissociating solvents is due to the thermodynamic syn preference of the anions. Syn/anti isomerization of the anions (E{sub a} < 26 kcal mol{sup {minus}1}) is rapid even at low temperatures. In contrast, the anti preference of the radicals of acetaldoxime indicates that themore » formation of the syn products in oxidative coupling reactions of the anions of oxime ethers is a kinetic effect.« less

Extracellular alpha-synuclein oligomers modulate synaptic transmission and impair LTP via NMDA-receptor activation.

PubMed

Diógenes, Maria José; Dias, Raquel B; Rombo, Diogo M; Vicente Miranda, Hugo; Maiolino, Francesca; Guerreiro, Patrícia; Näsström, Thomas; Franquelim, Henri G; Oliveira, Luís M A; Castanho, Miguel A R B; Lannfelt, Lars; Bergström, Joakim; Ingelsson, Martin; Quintas, Alexandre; Sebastião, Ana M; Lopes, Luísa V; Outeiro, Tiago Fleming

2012-08-22

Parkinson's disease (PD) is the most common representative of a group of disorders known as synucleinopathies, in which misfolding and aggregation of α-synuclein (a-syn) in various brain regions is the major pathological hallmark. Indeed, the motor symptoms in PD are caused by a heterogeneous degeneration of brain neurons not only in substantia nigra pars compacta but also in other extrastriatal areas of the brain. In addition to the well known motor dysfunction in PD patients, cognitive deficits and memory impairment are also an important part of the disorder, probably due to disruption of synaptic transmission and plasticity in extrastriatal areas, including the hippocampus. Here, we investigated the impact of a-syn aggregation on AMPA and NMDA receptor-mediated rat hippocampal (CA3-CA1) synaptic transmission and long-term potentiation (LTP), the neurophysiological basis for learning and memory. Our data show that prolonged exposure to a-syn oligomers, but not monomers or fibrils, increases basal synaptic transmission through NMDA receptor activation, triggering enhanced contribution of calcium-permeable AMPA receptors. Slices treated with a-syn oligomers were unable to respond with further potentiation to theta-burst stimulation, leading to impaired LTP. Prior delivery of a low-frequency train reinstated the ability to express LTP, implying that exposure to a-syn oligomers drives the increase of glutamatergic synaptic transmission, preventing further potentiation by physiological stimuli. Our novel findings provide mechanistic insight on how a-syn oligomers may trigger neuronal dysfunction and toxicity in PD and other synucleinopathies.

Synthesising acid mine drainage to maintain and exploit indigenous mining micro-algae and microbial assemblies for biotreatment investigations.

PubMed

Orandi, Sanaz; Lewis, David M

2013-02-01

The stringent regulations for discharging acid mine drainage (AMD) has led to increased attention on traditional or emerging treatment technologies to establish efficient and sustainable management for mine effluents. To assess new technologies, laboratory investigations on AMD treatment are necessary requiring a consistent supply of AMD with a stable composition, thus limiting environmental variability and uncertainty during controlled experiments. Additionally, biotreatment systems using live cells, particularly micro-algae, require appropriate nutrient availability. Synthetic AMD (Syn-AMD) meets these requirements. However, to date, most of the reported Syn-AMDs are composed of only a few selected heavy metals without considering the complexity of actual AMD. In this study, AMD was synthesised based on the typical AMD characteristics from a copper mine where biotreatment is being considered using indigenous AMD algal-microbes. Major cations (Ca, Na, Cu, Zn, Mg, Mn and Ni), trace metals (Al, Fe, Ag, Na, Co, Mo, Pb and Cr), essential nutrients (N, P and C) and high SO(4) were incorporated into the Syn-AMD. This paper presents the preparation of chemically complex Syn-AMD and the challenges associated with combining metal salts of varying solubility that is not restricted to one particular mine site. The general approach reported and the particular reagents used can produce alternative Syn-AMD with varying compositions. The successful growth of indigenous AMD algal-microbes in the Syn-AMD demonstrated its applicability as appropriate generic media for cultivation and maintenance of mining microorganisms for future biotreatment studies.

Enhanced motivation to alcohol in transgenic mice expressing human α-synuclein.

PubMed

Rotermund, Carola; Reolon, Gustavo K; Leixner, Sarah; Boden, Cindy; Bilbao, Ainhoa; Kahle, Philipp J

2017-11-01

α-Synuclein (αSYN) is the neuropathological hallmark protein of Parkinson's disease (PD) and related neurodegenerative disorders. Moreover, the gene encoding αSYN (SNCA) is a major genetic contributor to PD. Interestingly, independent genome-wide association studies also identified SNCA as the most important candidate gene for alcoholism. Furthermore, single-nucleotide-polymorphisms have been associated with alcohol-craving behavior and alcohol-craving patients showed augmented αSYN expression in blood. To investigate the effect of αSYN on the addictive properties of chronic alcohol use, we examined consumption, motivation, and seeking responses induced by environmental stimuli and relapse behavior in transgenic mice expressing the human mutant [A30P]αSYN throughout the brain. The primary reinforcing effects of alcohol under operant self-administration conditions were increased, while consumption and the alcohol deprivation effect were not altered in the transgenic mice. The same mice were subjected to immunohistochemical measurements of immediate-early gene inductions in brain regions involved in addiction-related behaviors. Acute ethanol injection enhanced immunostaining for the phosphorylated form of cAMP response element binding protein in both amygdala and nucleus accumbens of αSYN transgenic mice, while in wild-type mice no effect was visible. However, at the same time, levels of cFos remain unchanged in both genotypes. These results provide experimental confirmation of SNCA as a candidate gene for alcoholism in addition to its known link to PD. © 2017 International Society for Neurochemistry.

SU-F-303-12: Implementation of MR-Only Simulation for Brain Cancer: A Virtual Clinical Trial

DOE Office of Scientific and Technical Information (OSTI.GOV)

Glide-Hurst, C; Zheng, W; Kim, J

2015-06-15

Purpose: To perform a retrospective virtual clinical trial using an MR-only workflow for a variety of brain cancer cases by incorporating novel imaging sequences, tissue segmentation using phase images, and an innovative synthetic CT (synCT) solution. Methods: Ten patients (16 lesions) were evaluated using a 1.0T MR-SIM including UTE-DIXON imaging (TE = 0.144/3.4/6.9ms). Bone-enhanced images were generated from DIXON-water/fat and inverted UTE. Automated air segmentation was performed using unwrapped UTE phase maps. Segmentation accuracy was assessed by calculating intersection and Dice similarity coefficients (DSC) using CT-SIM as ground truth. SynCTs were generated using voxel-based weighted summation incorporating T2, FLAIR, UTE1,more » and bone-enhanced images. Mean absolute error (MAE) characterized HU differences between synCT and CT-SIM. Dose was recalculated on synCTs; differences were quantified using planar gamma analysis (2%/2 mm dose difference/distance to agreement) at isocenter. Digitally reconstructed radiographs (DRRs) were compared. Results: On average, air maps intersected 80.8 ±5.5% (range: 71.8–88.8%) between MR-SIM and CT-SIM yielding DSCs of 0.78 ± 0.04 (range: 0.70–0.83). Whole-brain MAE between synCT and CT-SIM was 160.7±8.8 HU, with the largest uncertainty arising from bone (MAE = 423.3±33.2 HU). Gamma analysis revealed pass rates of 99.4 ± 0.04% between synCT and CT-SIM for the cohort. Dose volume histogram analysis revealed that synCT tended to yield slightly higher doses. Organs at risk such as the chiasm and optic nerves were most sensitive due to their proximities to air/bone interfaces. DRRs generated via synCT and CT-SIM were within clinical tolerances. Conclusion: Our approach for MR-only simulation for brain cancer treatment planning yielded clinically acceptable results relative to the CT-based benchmark. While slight dose differences were observed, reoptimization of treatment plans and improved image registration can address this limitation. Future work will incorporate automated registration between setup images (cone-beam CT and kilovoltage images) for synCT and CT-SIM. Submitting institution holds research agreements with Philips HealthCare, Best, Netherlands and Varian Medical Systems, Palo Alto, CA. Research partially sponsored via an Internal Mentored Research Grant.« less

moxFG region encodes four polypeptides in the methanol-oxidizing bacterium Methylobacterium sp. strain AM1

DOE Office of Scientific and Technical Information (OSTI.GOV)

Anderson, D.J.; Lidstrom, M.E.

The polypeptides encoded by a putative methanol oxidation (mox) operon of Methylobacterium sp. strain AM1 were expressed in Escherichia coli, using a coupled in vivo T7 RNA polymerase/promoter gene expression system. Two mox genes had been previously mapped to this region: moxF, the gene encoding the methanol dehydrogenase (MeDH) polypeptide; and moxG, a gene believed to encode a soluble type c cytochrome, cytochrome c/sub L/. In this study, four polypeptides of M/sub r/, 60,000, 30,000, 20,000, and 12,000 were found to be encoded by the moxFG region and were tentatively designated moxF, -J, -G, and -I, respectively. The arrangement ofmore » the genes (5' to 3') was found to be moxFJGI. The identities of three of the four polypeptides were determined by protein immunoblot analysis. The product of moxF, the M/sub r/-60,000 polypeptide, was confirmed to be the MeDH polypeptide. The product of moxG, the M/sub r/-20,000 polypeptide, was identified as mature cytochrome c/sub L/, and the product of moxI, the M/sub r/-12,000 polypeptide, was identified as a MeDH-associated polypeptide that copurifies with the holoenzyme. The identity of the M/sub r/-30,000 polypeptide (the moxJ gene product) could not be determined. The function of the M/sub r/-12,000 MeDH-associated polypeptide is not yet clear. However, it is not present in mutants that lack the M/sub r/-60,000 MeDH subunit, and it appears that the stability of the MeDH-associated polypeptide is dependent on the presence of the M/sub r/-60,000 MeDH polypeptide. Our data suggest that both the M/sub r/-30,000 and -12,000 polypeptides are involved in methanol oxidation, which would bring to 12 the number of mox genes in Methylobacterium sp. strain AM1.« less

“Gate-keeper” Residues and Active-Site Rearrangements in DNA Polymerase μ Help Discriminate Non-cognate Nucleotides

PubMed Central

Li, Yunlang; Schlick, Tamar

2013-01-01

Incorporating the cognate instead of non-cognate substrates is crucial for DNA polymerase function. Here we analyze molecular dynamics simulations of DNA polymerase μ (pol μ) bound to different non-cognate incoming nucleotides including A:dCTP, A:dGTP, A(syn):dGTP, A:dATP, A(syn):dATP, T:dCTP, and T:dGTP to study the structure-function relationships involved with aberrant base pairs in the conformational pathway; while a pol μ complex with the A:dTTP base pair is available, no solved non-cognate structures are available. We observe distinct differences of the non-cognate systems compared to the cognate system. Specifically, the motions of active-site residue His329 and Asp330 distort the active site, and Trp436, Gln440, Glu443 and Arg444 tend to tighten the nucleotide-binding pocket when non-cognate nucleotides are bound; the latter effect may further lead to an altered electrostatic potential within the active site. That most of these “gate-keeper” residues are located farther apart from the upstream primer in pol μ, compared to other X family members, also suggests an interesting relation to pol μ's ability to incorporate nucleotides when the upstream primer is not paired. By examining the correlated motions within pol μ complexes, we also observe different patterns of correlations between non-cognate systems and the cognate system, especially decreased interactions between the incoming nucleotides and the nucleotide-binding pocket. Altered correlated motions in non-cognate systems agree with our recently proposed hybrid conformational selection/induced-fit models. Taken together, our studies propose the following order for difficulty of non-cognate system insertions by pol μ: T:dGTP

A new species of Voria Robineau-Desvoidy (Diptera: Tachinidae) from Area de Conservación Guanacaste in northwestern Costa Rica.

PubMed

Fleming, A J; Wood, D Monty; Smith, M Alex; Dapkey, Tanya; Hallwachs, Winnie; Janzen, Daniel

2017-01-01

We describe a new species in the genus Voria Robineau-Desvoidy, 1830 (Diptera: Tachinidae: Voriini) from Area de Conservación Guanacaste (ACG) in northwestern Costa Rica. It was reared as part of an ongoing inventory of wild-caught caterpillars spanning a variety of moth and butterfly families (Lepidoptera). Our study provides a concise description of the new species using morphology, life history, molecular data, and photographic documentation. In addition to the new species, we provide a diagnosis of the genus as well as new data relating to host use. The following new species of Voria is described: Voria erasmocoronadoi Fleming & Wood sp. n. The following are proposed by Fleming & Wood as new synonyms of Voria : Xenoplagia Townsend, 1914 syn. n. , Hystricovoria Townsend, 1928 syn. n. , Afrovoria Curran, 1938 syn. n. , and Anavoria Mesnil, 1953 syn. n. , and Itavoria Townsend, 1931 syn. n. The following new combinations are proposed as a result of the new synonymies: Voria bakeri (Townsend, 1928), comb. n. and Voria setosa (Townsend, 1914), comb. n. The authors also propose Voria pollyclari (Rocha-e-Silva, Lopes & Della Lucia, 1999), comb. n. based on the morphology of the holotype.

Ice recrystallization kinetics in the presence of synthetic antifreeze glycoprotein analogues using the framework of LSW theory.

PubMed

Budke, C; Heggemann, C; Koch, M; Sewald, N; Koop, T

2009-03-05

The Ostwald ripening of polycrystalline ice in aqueous sucrose solutions was investigated experimentally. The kinetics of this ice recrystallization process was studied at temperatures between -6 and -10 degrees C and varying ice volume fractions. Using the theory of Lifshitz, Slyozov, and Wagner (LSW), the diffusion-limited rate constant for ice recrystallization was determined. Also, the effects of synthetic analogues of natural antifreeze glycoproteins (AFGP) were studied. These analogues synAFGPmi (i = 3-5) contained monosaccharide side groups instead of disaccharide side groups that occur in natural AFGP. In order to account for the inhibition effect of the synAFGPmi, we have modified classical LSW theory, allowing for the derivation of inhibition rate constants. It was found that the investigated synAFGPmi inhibit ice recrystallization at concentrations down to approximately 3 microg mL(-1) or, equivalently, approximately 1 micromol L(-1) for the largest synAFGPmi investigated: synAFGPm5. Hence, our new method is capable of quantitatively assessing the efficiency of very similar AFGP with a sensitivity that is at least 2 orders of magnitude larger than that typical for quantitative thermal hysteresis measurements.

Synthetic biology: ensuring the greatest global value.

PubMed

Hollis, Aidan

2013-09-01

Synthetic biology (SynBio) has tremendous, transformative potential. Like other technologies, it can be used for good or ill. Currently, the structure of the allocation of potential benefits and risks is biased in favor of richer countries. The underlying problem is simple: most risks from SynBio are universal and affect both the rich and the poor with equal force; but benefits from SynBio can be expected to accrue chiefly to the rich. The risk/benefit balance is therefore skewed in a way that may lead to inefficient and unfair decisions. One potential solution is presented in this paper, using the principles that underlie the Health Impact Fund (HIF). The HIF is designed to reward companies based on assessed health impact, no matter where it occurs in the world, so that extending the life of a poor person is as profitable as extending the life of a rich person. This paper considers both the potential benefits and costs of SynBio; examines how the current global pharmaceutical industry is structured; introduces the HIF proposal; and finally explores how the principles underlying the HIF could be used productively with SynBio for global health.

The Interplay between Alpha-Synuclein Clearance and Spreading

PubMed Central

Lopes da Fonseca, Tomás; Villar-Piqué, Anna; Outeiro, Tiago Fleming

2015-01-01

Parkinson’s Disease (PD) is a complex neurodegenerative disorder classically characterized by movement impairment. Pathologically, the most striking features of PD are the loss of dopaminergic neurons and the presence of intraneuronal protein inclusions primarily composed of alpha-synuclein (α-syn) that are known as Lewy bodies and Lewy neurites in surviving neurons. Though the mechanisms underlying the progression of PD pathology are unclear, accumulating evidence suggests a prion-like spreading of α-syn pathology. The intracellular homeostasis of α-syn requires the proper degradation of the protein by three mechanisms: chaperone-mediated autophagy, macroautophagy and ubiquitin-proteasome. Impairment of these pathways might drive the system towards an alternative clearance mechanism that could involve its release from the cell. This increased release to the extracellular space could be the basis for α-syn propagation to different brain areas and, ultimately, for the spreading of pathology and disease progression. Here, we review the interplay between α-syn degradation pathways and its intercellular spreading. The understanding of this interplay is indispensable for obtaining a better knowledge of the molecular basis of PD and, consequently, for the design of novel avenues for therapeutic intervention. PMID:25874605

Chronic Caffeine Treatment Protects Against α-Synucleinopathy by Reestablishing Autophagy Activity in the Mouse Striatum.

PubMed

Luan, Yanan; Ren, Xiangpeng; Zheng, Wu; Zeng, Zhenhai; Guo, Yingzi; Hou, Zhidong; Guo, Wei; Chen, Xingjun; Li, Fei; Chen, Jiang-Fan

2018-01-01

Despite converging epidemiological evidence for the inverse relationship of regular caffeine consumption and risk of developing Parkinson's disease (PD) with animal studies demonstrating protective effect of caffeine in various neurotoxin models of PD, whether caffeine can protect against mutant α-synuclein (α-Syn) A53T-induced neurotoxicity in intact animals has not been examined. Here, we determined the effect of chronic caffeine treatment using the α-Syn fibril model of PD by intra-striatal injection of preformed A53T α-Syn fibrils. We demonstrated that chronic caffeine treatment blunted a cascade of pathological events leading to α-synucleinopathy, including pSer129α-Syn-rich aggregates, apoptotic neuronal cell death, microglia, and astroglia reactivation. Importantly, chronic caffeine treatment did not affect autophagy processes in the normal striatum, but selectively reversed α-Syn-induced defects in macroautophagy (by enhancing microtubule-associated protein 1 light chain 3, and reducing the receptor protein sequestosome 1, SQSTM1/p62) and chaperone-mediated autophagy (CMA, by enhancing LAMP2A). These findings support that caffeine-a strongly protective environment factor as suggested by epidemiological evidence-may represent a novel pharmacological therapy for PD by targeting autophagy pathway.

Syn-deformational features of Carlin-type Au deposits

USGS Publications Warehouse

Peters, S.G.

2004-01-01

Syn-deformational ore deposition played an important role in some Carlin-type Au deposits according to field and laboratory evidence, which indicates that flow of Au-bearing fluids was synchronous with regional-scale deformation events. Gold-related deformation events linked to ore genesis were distinct from high-level, brittle deformation that is typical of many epithermal deposits. Carlin-type Au deposits, with brittle-ductile features, most likely formed during tectonic events that were accompanied by significant fluid flow. Interactive deformation-fluid processes involved brittle-ductile folding, faulting, shearing, and gouge development that were focused along illite-clay and dissolution zones caused by hydrothermal alteration. Alteration along these deformation zones resulted in increased porosity and enhancement of fluid flow, which resulted in decarbonated, significant dissolution, collapse, and volume and mass reduction. Carlin-type Au deposits commonly are hosted in Paleozoic and Mesozoic sedimentary rocks (limestone, siltstone, argillite, shale, and quartzite) on the margins of cratons. The sedimentary basins containing the host rocks underwent tectonic events that influenced the development of stratabound, structurally controlled orebodies. Published by Elsevier Ltd.

Synthon preference in the cocrystal of 3,4,5-trifluorophenylboronic acid with urea.

PubMed

Kopczyńska, Karolina; Marek, Paulina H; Banaś, Bartłomiej; Madura, Izabela D

2017-11-01

The comprehensive description of the crystal structure of a novel 1:1 cocrystal of 3,4,5-trifluorophenylboronic acid with urea, C 6 H 4 BF 3 O 2 ·CH 4 N 2 O, is presented. Both components are good candidates for crystal engineering as they can create a variety of supramolecular synthons. The preference for the formation of different hetrosynthons is verified based on theoretical calculations. The syn-anti conformation of boronic acid has been found to be the most favourable in the formation of intermolecular interactions with urea. Moreover, the distortions present in the boron coordination sphere have been described quantitatively based on experimental data according to bond-valence vector model calculations. The results revealed that the deformation of the sphere is typical for a syn-anti conformation of boronic acids. The supramolecular structure of the cocrystal is composed of large synthons in the form of layers made up of O-H...O and N-H...O hydrogen bonds. The layers are joined via N-H...F hydrogen bonds which are unusual for urea cocrystal structures.

1D helical cadmium coordination polymers containing hydrazide ligand: The role of solvent and molar ratio

NASA Astrophysics Data System (ADS)

Notash, Behrouz

2018-03-01

Three new cadmium coordination polymers, [Cd(L)(NO3)2CH3OH]n, 1, {[Cd(L)2(NO3)]NO3}n, 2 and {[Cd(L)2(NO3)]NO3.H2O}n3, which L is nicotinohydrazide have been synthesized and characterized by spectroscopic methods as well as single crystal X-ray diffraction. Compounds 1-3 have been synthesized by changing solvent and metal-to-ligand ratio. X-ray crystallography showed that compounds 1-3 have different 1D helical structural motif. Semi-flexible nature of L ligand causes to syn-syn conformation which leading to form 1D helical chains coordination polymers. Compounds 2 and 3 were synthesized under the same reaction conditions with similar molar ratio, but using different solvent system. These compounds are pseudopolymorph which differs in the presence or absence of water molecule in their crystal packing. Hirshfeld surface analysis of the structures 1-3 have been performed and find the percent of participation of intermolecular interactions in the crystal packing of compounds.

Elastomeric Polypeptides

PubMed Central

van Eldijk, Mark B.; McGann, Christopher L.

2013-01-01

Elastomeric polypeptides are very interesting biopolymers and are characterized by rubber-like elasticity, large extensibility before rupture, reversible deformation without loss of energy, and high resilience upon stretching. Their useful properties have motivated their use in a wide variety of materials and biological applications. This chapter focuses on elastin and resilin – two elastomeric biopolymers – and the recombinant polypeptides derived from them (elastin-like polypeptides and resilin-like polypeptides). This chapter also discusses the applications of these recombinant polypeptides in the fields of purification, drug delivery, and tissue engineering. PMID:21826606

The effect of grapefruit juice on drug disposition

PubMed Central

Hanley, Michael J.; Cancalon, Paul; Widmer, Wilbur W.; Greenblatt, David J.

2011-01-01

Introduction Since their initial discovery in 1989, grapefruit juice-drug interactions have received extensive interest from the scientific, medical, regulatory, and lay communities. Although knowledge regarding the effects of grapefruit juice on drug disposition continues to expand, the list of drugs studied in the clinical setting remains relatively limited. Areas covered This article reviews the in vitro effects of grapefruit juice and its constituents on the activity of cytochrome P450 enzymes, organic anion-transporting polypeptides, P-glycoprotein, esterases and sulfotransferases. The translational applicability of the in vitro findings to the clinical setting is discussed for each drug metabolizing enzyme and transporter. Reported area under the plasma concentration-time curve ratios for available grapefruit juice-drug interaction studies are also provided. Relevant investigations were identified by searching the Pubmed electronic database from 1989 to 2010. Expert opinion Grapefruit juice increases the bioavailability of some orally-administered drugs that are metabolized by CYP3A and normally undergo extensive presystemic extraction. In addition, grapefruit juice can decrease the oral absorption of a few drugs that rely on organic anion-transporting polypeptides in the gastrointestinal tract for their uptake. The number of drugs shown to interact with grapefruit juice in vitro is far greater than the number of clinically relevant grapefruit juice-drug interactions. For the majority of patients, complete avoidance of grapefruit juice is unwarranted. PMID:21254874

Methods for engineering polypeptide variants via somatic hypermutation and polypeptide made thereby

DOEpatents

Tsien, Roger Y; Wang, Lei

2015-01-13

Methods using somatic hypermutation (SHM) for producing polypeptide and nucleic acid variants, and nucleic acids encoding such polypeptide variants are disclosed. Such variants may have desired properties. Also disclosed are novel polypeptides, such as improved fluorescent proteins, produced by the novel methods, and nucleic acids, vectors, and host cells comprising such vectors.

Accumulation of the Cyclobutane Thymine Dimer in Defined Sequences of Free and Nucleosomal DNA

DTIC Science & Technology

2013-08-01

cyclobutane dimer in a single-stranded vector , Proc. Natl. Acad. Sci. U. S. A., 1988, 85, 8141–8145. 11 C. A. Smith, M. Wang, N. Jiang, L. Che, X. Zhao and...J.-S. Taylor, Mutation spectra of M13 vectors containing site-specific cis–syn, trans–syn-I, (6-4), and Dewar pyrimi- done photoproducts of thymidylyl...Bypass of a site-specific cis–syn thymine dimer in a SV40 vector during in vitro replication by HeLa and XPV cell-free extracts, Biochemistry, 1998

MedSynDiKATe--design considerations for an ontology-based medical text understanding system.

PubMed Central

Hahn, U.; Romacker, M.; Schulz, S.

2000-01-01

MedSynDiKATe is a natural language processor for automatically acquiring knowledge from medical finding reports. The content of these documents is transferred to formal representation structures which constitute a corresponding text knowledge base. The general system architecture we present integrates requirements from the analysis of single sentences, as well as those of referentially linked sentences forming cohesive texts. The strong demands MedSynDiKATe poses to the availability of expressive knowledge sources are accounted for by two alternative approaches to (semi)automatic ontology engineering. PMID:11079899

Analysis using fluorescence labeling and mass spectrometry of disulfide-mediated interactions of soy protein when heated.

PubMed

Ruan, Qijun; Chen, Yeming; Kong, Xiangzhen; Hua, Yufei

2015-04-08

It is well-known that disulfide-mediated interactions are important when soy protein is heated, in which soy proteins are dissociated and rearranged to some new forms. In this study, the disulfide bond (SS) linked polymer, which was composed of the acidic (A) polypeptides of glycinin, basic (B) polypeptides of glycinin, and a small amount of α' and α of β-conglycinin, was formed as the major product, accompanied by the formation of SS-linked dimer of B and monomer of A as minor products. The role of sulfhydryl (SH) of different subunits/polypeptides for forming intermolecular SS was investigated. The SH of B in glycinin (Cys298 of G1, Cys289 of G2, Cys440 of G4) was transformed into SS in polymer identified by mass spectrometry analysis. The SH content of polymer was lower than that of glycinin and β-conglycinin subunits when heated. The SH content of B in polymer was lower than that in glycinin subunit, and both of them were decreased by heating. The SH content of A in polymer was increased and higher than that of B in polymer and A in glycinin subunit when heated. These results indicated that the SH of B in glycinin subunit was subjected to not only SH oxidation but also SH-SS exchange (with SS of A) for forming intermolecular SS of polymer. The SH oxidation and SH-SS exchange were proven by the change of SH content for the first time. The SH of B was suggested to be reactive for forming intermolecular SS of polymer.

A taxonomic monograph of the assassin bug genus Zelus Fabricius (Hemiptera: Reduviidae): 71 species based on 10,000 specimens

PubMed Central

Hart, Elwood R; Weirauch, Christiane

2016-01-01

Abstract The New World assassin bug genus Zelus Fabricius, 1803 (Insecta: Hemiptera: Heteroptera: Reduviidae: Harpactorinae: Harpactorini) is revised based on more than 10,000 specimens. Seventy-one species are recognized and twenty-four described as new: Zelus aithaleos sp. n., Zelus amblycephalus sp. n., Zelus antiguensis sp. n., Zelus auralanus sp. n., Zelus bahiaensis sp. n., Zelus banksi sp. n., Zelus casii sp. n., Zelus championi sp. n., Zelus cordazulus sp. n., Zelus fuliginatus sp. n., Zelus gilboventris sp. n., Zelus gracilipes sp. n., Zelus grandoculus sp. n., Zelus kartaboides sp. n., Zelus lewisi sp. n., Zelus panamensis sp. n., Zelus paracephalus sp. n., Zelus rosulentus sp. n., Zelus russulumus sp. n., Zelus spatulosus sp. n., Zelus truxali sp. n., Zelus umbraculoides sp. n., Zelus umbraculus sp. n., and Zelus xouthos sp. n. Five species, Zelus araneiformis Haviland, 1931, Zelus gradarius Bergroth, 1905, Zelus modestus (Stål, 1862), Zelus subfasciatus Stål, 1860 and Zelus vittaticeps Stål, 1866, are removed from Zelus and placed incertae sedis within Harpactorini. Nine new synonyms are recognized (senior synonym in parentheses): Zelus atripes Champion, 1898 syn. nov. (=Zelus conjungens [Stål, 1860]), Zelus dispar Fabricius, 1803 syn. nov. (=Zelus pedestris Fabricius, 1803), Zelus formosus Haviland, 1931 syn. nov. (=Zelus laticornis Herrich-Schaeffer, 1853), Zelus obscuridorsis (Stål, 1860) syn. nov. (=Zelus pedestris), Zelus pallidinervus Haviland, 1931 syn. nov. (=Zelus kartabensis Haviland, 1931), Zelus personatus Berg, 1879 syn. nov. (=Zelus versicolor Herrich-Schaeffer, 1848), Zelus trimaculatus Champion, 1898 syn. nov. (=Zelus means Fabricius, 1803), Zelus trimaculicollis (Stål, 1855) syn. nov. (=Zelus means), and Zelus tristis Haviland, 1931 syn. nov. (=Zelus laticornis). Zelus conjungens (Stål, 1860) stat. rev. Is resurrected from junior synonymy with zealous armillatus (Lepeletier & Seville, 1825). Zelus ambulans Stål, 1862 stat. rev. and Zelus cognatus (Costa, 1862) stat. rev. are resurrected from synonymy with Zelus exsanguis Stål, 1862. Iquitozelus Bérenger syn. nov. is synonymized with Zelus and its only species transferred to Zelus, hence resulting in a new combination, Zelus couturieri (Bérenger, 2003) comb. nov. Lectotypes, paralectotypes or neotypes are designated for a number of species. Habitus images, illustrations of male genitalia, distribution maps and measurements are provided for nearly all species. The three previously recognized subgenera of Zelus are found to be based upon superficial characters and these divisions do not reflect natural groupings. Using sets of characters, especially those of the male genitalia, eleven species groups are proposed. It is also hypothesized that Zelus is closely related to three other New World genera: Atopozelus Elkins, Ischnoclopius Stål and an undescribed genus "Hartzelus" [manuscript name]. Zelus is endemic to the New World, occurring naturally in the Caribbean and all but one of the continental countries, with introductions to Pacific islands, Europe and Chile. PMID:27651730

A taxonomic monograph of the assassin bug genus Zelus Fabricius (Hemiptera: Reduviidae): 71 species based on 10,000 specimens.

PubMed

Zhang, Guanyang; Hart, Elwood R; Weirauch, Christiane

2016-01-01

The New World assassin bug genus Zelus Fabricius, 1803 (Insecta: Hemiptera: Heteroptera: Reduviidae: Harpactorinae: Harpactorini) is revised based on more than 10,000 specimens. Seventy-one species are recognized and twenty-four described as new: Zelus aithaleos sp. n., Zelus amblycephalus sp. n., Zelus antiguensis sp. n., Zelus auralanus sp. n., Zelus bahiaensis sp. n., Zelus banksi sp. n., Zelus casii sp. n., Zelus championi sp. n., Zelus cordazulus sp. n., Zelus fuliginatus sp. n., Zelus gilboventris sp. n., Zelus gracilipes sp. n., Zelus grandoculus sp. n., Zelus kartaboides sp. n., Zelus lewisi sp. n., Zelus panamensis sp. n., Zelus paracephalus sp. n., Zelus rosulentus sp. n., Zelus russulumus sp. n., Zelus spatulosus sp. n., Zelus truxali sp. n., Zelus umbraculoides sp. n., Zelus umbraculus sp. n., and Zelus xouthos sp. n. Five species, Zelus araneiformis Haviland, 1931, Zelus gradarius Bergroth, 1905, Zelus modestus (Stål, 1862), Zelus subfasciatus Stål, 1860 and Zelus vittaticeps Stål, 1866, are removed from Zelus and placed incertae sedis within Harpactorini. Nine new synonyms are recognized (senior synonym in parentheses): Zelus atripes Champion, 1898 syn. nov. (=Zelus conjungens [Stål, 1860]), Zelus dispar Fabricius, 1803 syn. nov. (=Zelus pedestris Fabricius, 1803), Zelus formosus Haviland, 1931 syn. nov. (=Zelus laticornis Herrich-Schaeffer, 1853), Zelus obscuridorsis (Stål, 1860) syn. nov. (=Zelus pedestris), Zelus pallidinervus Haviland, 1931 syn. nov. (=Zelus kartabensis Haviland, 1931), Zelus personatus Berg, 1879 syn. nov. (=Zelus versicolor Herrich-Schaeffer, 1848), Zelus trimaculatus Champion, 1898 syn. nov. (=Zelus means Fabricius, 1803), Zelus trimaculicollis (Stål, 1855) syn. nov. (=Zelus means), and Zelus tristis Haviland, 1931 syn. nov. (=Zelus laticornis). Zelus conjungens (Stål, 1860) stat. rev. Is resurrected from junior synonymy with zealous armillatus (Lepeletier & Seville, 1825). Zelus ambulans Stål, 1862 stat. rev. and Zelus cognatus (Costa, 1862) stat. rev. are resurrected from synonymy with Zelus exsanguis Stål, 1862. Iquitozelus Bérenger syn. nov. is synonymized with Zelus and its only species transferred to Zelus, hence resulting in a new combination, Zelus couturieri (Bérenger, 2003) comb. nov. Lectotypes, paralectotypes or neotypes are designated for a number of species. Habitus images, illustrations of male genitalia, distribution maps and measurements are provided for nearly all species. The three previously recognized subgenera of Zelus are found to be based upon superficial characters and these divisions do not reflect natural groupings. Using sets of characters, especially those of the male genitalia, eleven species groups are proposed. It is also hypothesized that Zelus is closely related to three other New World genera: Atopozelus Elkins, Ischnoclopius Stål and an undescribed genus "Hartzelus" [manuscript name]. Zelus is endemic to the New World, occurring naturally in the Caribbean and all but one of the continental countries, with introductions to Pacific islands, Europe and Chile.

Reduction of Phosphorylated Synapsin I (Ser-553) Leads to Spatial Memory Impairment by Attenuating GABA Release after Microwave Exposure in Wistar Rats

PubMed Central

Qiao, Simo; Peng, Ruiyun; Yan, Haitao; Gao, Yabing; Wang, Changzhen; Wang, Shuiming; Zou, Yong; Xu, Xinping; Zhao, Li; Dong, Ji; Su, Zhentao; Feng, Xinxin; Wang, Lifeng; Hu, Xiangjun

2014-01-01

Background Abnormal release of neurotransmitters after microwave exposure can cause learning and memory deficits. This study investigated the mechanism of this effect by exploring the potential role of phosphorylated synapsin I (p-Syn I). Methods Wistar rats, rat hippocampal synaptosomes, and differentiated (neuronal) PC12 cells were exposed to microwave radiation for 5 min at a mean power density of 30 mW/cm2. Sham group rats, synaptosomes, and cells were otherwise identically treated and acted as controls for all of the following post-exposure analyses. Spatial learning and memory in rats was assessed using the Morris Water Maze (MWM) navigation task. The protein expression and presynaptic distribution of p-Syn I and neurotransmitter transporters were examined via western blotting and immunoelectron microscopy, respectively. Levels amino acid neurotransmitter release from rat hippocampal synaptosomes and PC12 cells were measured using high performance liquid chromatograph (HPLC) at 6 hours after exposure, with or without synapsin I silencing via shRNA transfection. Results In the rat experiments, there was a decrease in spatial memory performance after microwave exposure. The expression of p-Syn I (ser-553) was decreased at 3 days post-exposure and elevated at later time points. Vesicular GABA transporter (VGAT) was significantly elevated after exposure. The GABA release from synaptosomes was attenuated and p-Syn I (ser-553) and VGAT were both enriched in small clear synaptic vesicles, which abnormally assembled in the presynaptic terminal after exposure. In the PC12 cell experiments, the expression of p-Syn I (ser-553) and GABA release were both attenuated at 6 hours after exposure. Both microwave exposure and p-Syn I silencing reduced GABA release and maximal reduction was found for the combination of the two, indicating a synergetic effect. Conclusion p-Syn I (ser-553) was found to play a key role in the impaired GABA release and cognitive dysfunction that was induced by microwave exposure. PMID:24743689

Reduction of phosphorylated synapsin I (ser-553) leads to spatial memory impairment by attenuating GABA release after microwave exposure in Wistar rats.

PubMed

Qiao, Simo; Peng, Ruiyun; Yan, Haitao; Gao, Yabing; Wang, Changzhen; Wang, Shuiming; Zou, Yong; Xu, Xinping; Zhao, Li; Dong, Ji; Su, Zhentao; Feng, Xinxin; Wang, Lifeng; Hu, Xiangjun

2014-01-01

Abnormal release of neurotransmitters after microwave exposure can cause learning and memory deficits. This study investigated the mechanism of this effect by exploring the potential role of phosphorylated synapsin I (p-Syn I). Wistar rats, rat hippocampal synaptosomes, and differentiated (neuronal) PC12 cells were exposed to microwave radiation for 5 min at a mean power density of 30 mW/cm2. Sham group rats, synaptosomes, and cells were otherwise identically treated and acted as controls for all of the following post-exposure analyses. Spatial learning and memory in rats was assessed using the Morris Water Maze (MWM) navigation task. The protein expression and presynaptic distribution of p-Syn I and neurotransmitter transporters were examined via western blotting and immunoelectron microscopy, respectively. Levels amino acid neurotransmitter release from rat hippocampal synaptosomes and PC12 cells were measured using high performance liquid chromatograph (HPLC) at 6 hours after exposure, with or without synapsin I silencing via shRNA transfection. In the rat experiments, there was a decrease in spatial memory performance after microwave exposure. The expression of p-Syn I (ser-553) was decreased at 3 days post-exposure and elevated at later time points. Vesicular GABA transporter (VGAT) was significantly elevated after exposure. The GABA release from synaptosomes was attenuated and p-Syn I (ser-553) and VGAT were both enriched in small clear synaptic vesicles, which abnormally assembled in the presynaptic terminal after exposure. In the PC12 cell experiments, the expression of p-Syn I (ser-553) and GABA release were both attenuated at 6 hours after exposure. Both microwave exposure and p-Syn I silencing reduced GABA release and maximal reduction was found for the combination of the two, indicating a synergetic effect. p-Syn I (ser-553) was found to play a key role in the impaired GABA release and cognitive dysfunction that was induced by microwave exposure.

One-Pot Catalytic Enantio- and Diastereoselective Syntheses of anti-, syn-cis-Disubstituted, and syn-Vinyl Cyclopropyl Alcohols

PubMed Central

Kim, Hun Young; Salvi, Luca; Carroll, Patrick J.; Walsh, Patrick J.

2009-01-01

Highly enantio- and diastereoselective methods for the synthesis of a variety of cyclopropyl alcohols are reported. These methods represent the first one-pot approaches to syn-vinyl cyclopropyl alcohols, syn-cis-disubstituted cyclopropyl alcohols, and anti-cyclopropyl alcohols from achiral precursors. The methods begin with enantioselective C–C bond formations promoted by a MIB-based zinc catalyst to generate allylic alkoxide intermediates. The intermediates are then subjected to in situ alkoxide-directed cyclopropanation to provide cyclopropyl alcohols. In the synthesis of vinyl cyclopropyl alcohols, hydroboration of enynes is followed by transmetalation of the resulting dienylborane to zinc to provide dienylzinc reagents. Enantioselective addition to aldehydes generates the requisite dienyl zinc alkoxides, which are then subjected to in situ cyclopropanation to furnish vinyl cyclopropyl alcohols. Cyclopropanation occurs at the double bond allylic to the alkoxide. Using this method, syn-vinylcyclopropyl alcohols are obtained in 65–85% yield, 76–93% ee, and >19:1 dr. To prepare anti-cyclopropanols, enantioselective addition of alkylzinc reagents to conjugated enals provides allylic zinc alkoxides. Because direct cyclopropanation provides syn-cyclopropyl alcohols, the intermediate allylic alkoxides were treated with TMSCl/Et3N to generate intermediate silyl ethers. In situ cyclopropanation of the allylic silyl ether resulted in cyclopropanation to form the anti-cyclopropyl silyl ether. Workup with TBAF affords the anti-cyclopropyl alcohols in one-pot in 60–82% yield, 89–99% ee, and ≥10:1 dr. For the synthesis of cis-disubstituted cyclopropyl alcohols, in situ generated (Z)-vinyl zinc reagents were employed in asymmetric addition to aldehydes to generate (Z)-allylic zinc alkoxides. In situ cyclopropanation provides syn-cis-disubstituted cyclopropyl alcohols in 42–70% yield, 88–97% ee, and >19:1 dr. These one-pot procedures enable the synthesis of a diverse array of cyclopropyl alcohol building blocks with high enantio- and diastereoselectivities. PMID:19954146

Deep functional analysis of synII, a 770-kilobase synthetic yeast chromosome.

PubMed

Shen, Yue; Wang, Yun; Chen, Tai; Gao, Feng; Gong, Jianhui; Abramczyk, Dariusz; Walker, Roy; Zhao, Hongcui; Chen, Shihong; Liu, Wei; Luo, Yisha; Müller, Carolin A; Paul-Dubois-Taine, Adrien; Alver, Bonnie; Stracquadanio, Giovanni; Mitchell, Leslie A; Luo, Zhouqing; Fan, Yanqun; Zhou, Baojin; Wen, Bo; Tan, Fengji; Wang, Yujia; Zi, Jin; Xie, Zexiong; Li, Bingzhi; Yang, Kun; Richardson, Sarah M; Jiang, Hui; French, Christopher E; Nieduszynski, Conrad A; Koszul, Romain; Marston, Adele L; Yuan, Yingjin; Wang, Jian; Bader, Joel S; Dai, Junbiao; Boeke, Jef D; Xu, Xun; Cai, Yizhi; Yang, Huanming

2017-03-10

Here, we report the successful design, construction, and characterization of a 770-kilobase synthetic yeast chromosome II (synII). Our study incorporates characterization at multiple levels-including phenomics, transcriptomics, proteomics, chromosome segregation, and replication analysis-to provide a thorough and comprehensive analysis of a synthetic chromosome. Our Trans-Omics analyses reveal a modest but potentially relevant pervasive up-regulation of translational machinery observed in synII, mainly caused by the deletion of 13 transfer RNAs. By both complementation assays and SCRaMbLE (synthetic chromosome rearrangement and modification by loxP -mediated evolution), we targeted and debugged the origin of a growth defect at 37°C in glycerol medium, which is related to misregulation of the high-osmolarity glycerol response. Despite the subtle differences, the synII strain shows highly consistent biological processes comparable to the native strain. Copyright © 2017, American Association for the Advancement of Science.

Multi-functional roles for the polypeptide transport associated domains of Toc75 in chloroplast protein import

PubMed Central

Paila, Yamuna D; Richardson, Lynn GL; Inoue, Hitoshi; Parks, Elizabeth S; McMahon, James; Inoue, Kentaro; Schnell, Danny J

2016-01-01

Toc75 plays a central role in chloroplast biogenesis in plants as the membrane channel of the protein import translocon at the outer envelope of chloroplasts (TOC). Toc75 is a member of the Omp85 family of bacterial and organellar membrane insertases, characterized by N-terminal POTRA (polypeptide-transport associated) domains and C-terminal membrane-integrated β-barrels. We demonstrate that the Toc75 POTRA domains are essential for protein import and contribute to interactions with TOC receptors, thereby coupling preprotein recognition at the chloroplast surface with membrane translocation. The POTRA domains also interact with preproteins and mediate the recruitment of molecular chaperones in the intermembrane space to facilitate membrane transport. Our studies are consistent with the multi-functional roles of POTRA domains observed in other Omp85 family members and demonstrate that the domains of Toc75 have evolved unique properties specific to the acquisition of protein import during endosymbiotic evolution of the TOC system in plastids. DOI: http://dx.doi.org/10.7554/eLife.12631.001 PMID:26999824

A split motor domain in a cytoplasmic dynein

PubMed Central

Straube, Anne; Enard, Wolfgang; Berner, Alexandra; Wedlich-Söldner, Roland; Kahmann, Regine; Steinberg, Gero

2001-01-01

The heavy chain of dynein forms a globular motor domain that tightly couples the ATP-cleavage region and the microtubule-binding site to transform chemical energy into motion along the cytoskeleton. Here we show that, in the fungus Ustilago maydis, two genes, dyn1 and dyn2, encode the dynein heavy chain. The putative ATPase region is provided by dyn1, while dyn2 includes the predicted microtubule-binding site. Both genes are located on different chromosomes, are transcribed into independent mRNAs and are translated into separate polypeptides. Both Dyn1 and Dyn2 co-immunoprecipitated and co-localized within growing cells, and Dyn1–Dyn2 fusion proteins partially rescued mutant phenotypes, suggesting that both polypeptides interact to form a complex. In cell extracts the Dyn1–Dyn2 complex dissociated, and microtubule affinity purification indicated that Dyn1 or associated polypeptides bind microtubules independently of Dyn2. Both Dyn1 and Dyn2 were essential for cell survival, and conditional mutants revealed a common role in nuclear migration, cell morphogenesis and microtubule organization, indicating that the Dyn1–Dyn2 complex serves multiple cellular functions. PMID:11566874

Characterization of a novel isoform of alpha-nascent polypeptide-associated complex as IgE-defined autoantigen.

PubMed

Mossabeb, Roschanak; Seiberler, Susanne; Mittermann, Irene; Reininger, Renate; Spitzauer, Susanne; Natter, Susanne; Verdino, Petra; Keller, Walter; Kraft, Dietrich; Valenta, Rudolf

2002-10-01

The nascent polypeptide-associated complex is required for intracellular translocation of newly synthesized polypeptides in eukaryotic cells. It may also act as a transcriptional coactivator in humans and various eukaryotic organisms and binds to nucleic acids. Recently, we provided evidence that a component of nascent polypeptide-associated complex, alpha-nascent polypeptide-associated complex, represents an IgE-reactive autoantigen for atopic dermatitis patients. By oligonucleotide screening we isolated a complete cDNA coding for a so far unknown alpha-nascent polypeptide-associated complex isoform from a human epithelial cDNA library. Southern blot hybridization experiments provided further evidence that alpha-nascent polypeptide-associated complex is encoded by a gene family. Recombinant alpha-nascent polypeptide-associated complex was expressed in Escherichia coli as a soluble, His-tagged protein, and purified via nickel affinity chromatography. By circular dichroism analysis it is demonstrated that purified recombinant alpha-nascent polypeptide-associated complex represents a folded protein of mixed alpha-helical and beta-sheet conformation with unusual high thermal stability and remarkable refolding capacity. Complete recombinant alpha-nascent polypeptide-associated complex (215 amino acids) and its 86 amino acid C-terminal fragment specifically bound IgE autoantibodies. Recombinant alpha-nascent polypeptide-associated complex also inhibited IgE binding to natural alpha-nascent polypeptide-associated complex, demonstrating the presence of common IgE epitopes between the recombinant and natural protein. Furthermore, recombinant alpha-nascent polypeptide-associated complex induced specific lymphoproliferative responses in peripheral blood mononuclear cells of a sensitized atopic dermatitis patient. As has been proposed for environmental allergens it is possible that T cell responses to IgE-defined autoantigens may contribute to the chronic skin manifestations in atopic dermatitis.

Polypeptides having catalase activity and polynucleotides encoding same

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Ye; Duan, Junxin; Zhang, Yu

Provided are isolated polypeptides having catalase activity and polynucleotides encoding the polypeptides. Also provided are nucleic acid constructs, vectors and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.

Evolution of enzymatic activity in the enolase superfamily: structural and mutagenic studies of the mechanism of the reaction catalyzed by o-succinylbenzoate synthase from Escherichia coli.

PubMed

Klenchin, Vadim A; Taylor Ringia, Erika A; Gerlt, John A; Rayment, Ivan

2003-12-16

o-Succinylbenzoate synthase (OSBS) from Escherichia coli, a member of the enolase superfamily, catalyzes an exergonic dehydration reaction in the menaquinone biosynthetic pathway in which 2-succinyl-6-hydroxy-2,4-cyclohexadiene-1-carboxylate (SHCHC) is converted to 4-(2'-carboxyphenyl)-4-oxobutyrate (o-succinylbenzoate or OSB). Our previous structural studies of the Mg(2+).OSB complex established that OSBS is a member of the muconate lactonizing enzyme subgroup of the superfamily: the essential Mg(2+) is coordinated to carboxylate ligands at the ends of the third, fourth, and fifth beta-strands of the (beta/alpha)(7)beta-barrel catalytic domain, and the OSB product is located between the Lys 133 at the end of the second beta-strand and the Lys 235 at the end of the sixth beta-strand [Thompson, T. B., Garrett, J. B., Taylor, E. A, Meganathan, R., Gerlt, J. A., and Rayment, I. (2000) Biochemistry 39, 10662-76]. Both Lys 133 and Lys 235 were separately replaced with Ala, Ser, and Arg residues; all six mutants displayed no detectable catalytic activity. The structure of the Mg(2+).SHCHC complex of the K133R mutant has been solved at 1.62 A resolution by molecular replacement starting from the structure of the Mg(2+).OSB complex. This establishes the absolute configuration of SHCHC: the C1-carboxylate and the C6-OH leaving group are in a trans orientation, requiring that the dehydration proceed via a syn stereochemical course. The side chain of Arg 133 is pointed out of the active site so that it cannot function as a general base, whereas in the wild-type enzyme complexed with Mg(2+).OSB, the side chain of Lys 133 is appropriately positioned to function as the only acid/base catalyst in the syn dehydration. The epsilon-ammonium group of Lys 235 forms a cation-pi interaction with the cyclohexadienyl moiety of SHCHC, suggesting that Lys 235 also stabilizes the enediolate anion intermediate in the syn dehydration via a similar interaction.

Rosebud SynCoal Partnership, SynCoal{reg_sign} demonstration technology update

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sheldon, R.W.

1997-12-31

An Advanced Coal Conversion Process (ACCP) technology being demonstrated in eastern Montana (USA) at the heart of one of the world`s largest coal deposits is providing evidence that the molecular structure of low-rank coals can be altered successfully to produce a unique product for a variety of utility and industrial applications. The product is called SynCoal{reg_sign} and the process has been developed by the Rosebud SynCoal Partnership (RSCP) through the US Department of Energy`s multi-million dollar Clean Coal Technology Program. The ACCP demonstration process uses low-pressure, superheated gases to process coal in vibrating fluidized beds. Two vibratory fluidized processing stagesmore » are used to heat and convert the coal. This is followed by a water spray quench and a vibratory fluidized stage to cool the coal. Pneumatic separators remove the solid impurities from the dried coal. There are three major steps to the SynCoal{reg_sign} process: (1) thermal treatment of the coal in an inert atmosphere, (2) inert gas cooling of the hot coal, and (3) removal of ash minerals. When operated continuously, the demonstration plant produces over 1,000 tons per day (up to 300,000 tons per year) of SynCoal{reg_sign} with a 2% moisture content, approximately 11,800b Btu/lb and less than 1.0 pound of SO{sub 2} per million Btu. This product is obtained from Rosebud Mine sub-bituminous coal which starts with 25% moisture, 8,600 Btu/lb and approximately 1.6 pounds of SO{sub 2} per million Btu.« less

Secretion of pancreatic polypeptide in patients with pancreatic endocrine tumors.

PubMed

Adrian, T E; Uttenthal, L O; Williams, S J; Bloom, S R

1986-07-31

Pancreatic polypeptide is often secreted by pancreatic endocrine tumors and is considered a marker for such tumors. To investigate the diagnostic value of this marker, we studied 323 patients with proved pancreatic endocrine tumors. We found plasma concentrations of pancreatic polypeptide to be elevated (more than 300 pmol per liter) in 144 patients (diagnostic sensitivity, 45 percent). However, plasma levels of pancreatic polypeptide can also be elevated in the absence of a pancreatic tumor. To ascertain whether the administration of atropine could distinguish between normal and tumor-associated polypeptide secretion, we studied 30 patients with pancreatic tumors and high plasma levels of pancreatic polypeptide, 18 patients without tumors who had elevated levels of pancreatic polypeptide, and eight normal controls. Polypeptide levels in the 18 patients without tumors were substantially lower than in the 30 patients with tumors. Atropine (1 mg intramuscularly) did not suppress polypeptide levels in patients with tumors, but did suppress plasma levels by more than 50 percent in all subjects without tumors. Thus, although its diagnostic sensitivity is low, pancreatic polypeptide appears to be a useful adjunctive marker of many pancreatic endocrine tumors, and the atropine suppression test can be used to distinguish normal from tumor-related secretion of the polypeptide. Identification of the type of pancreatic endocrine tumor still requires measurement of the hormone that is specific for the tumor.

Polypeptides having beta-glucosidase activity and polynucleotides encoding same

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Ye; Duan, Junxin; Zhang, Yu

Provided are isolated polypeptides having beta-glucosidase activity and polynucleotides encoding the polypeptides. Also provided are nucleic acid constructs, vectors and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.

Polypeptides having endoglucanase activity and polynucleotides encoding same

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhang, Yu; Liu, Ye; Duan, Junxin

Provided are isolated polypeptides having endoglucanase activity and isolated polynucleotides encoding the polypeptides. Also provided are nucleic acid constructs, vectors and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.

Polypeptides having beta-xylosidase activity and polynucleotides encoding same

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Ye; Tang, Lan; Zhang, Yu

Provided are isolated polypeptides having beta-xylosidase activity and polynucleotides encoding the polypeptides. Also provided are nucleic acid constructs, vectors and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.

Hybrid polypeptides having cellobiohydrolase activity and polynucleotides encoding same

DOEpatents

Liu, Ye; Shaghasi, Tarana

2016-11-01

The present invention provides hybrid polypeptides having cellobiohydrolase activity. The present invention also provides polynucleotides encoding the hybrid polypeptides; nucleic acid constructs, vectors and host cells comprising the polynucleotides; and processes of using the hybrid polypeptides.

Combinatorial discovery of enzymes with utility in biomass transformation

DOEpatents

Fox, Brian G; Elsen, Nathaniel L

2015-02-03

Methods for the cell-free identification of polypeptide and polypeptide combinations with utility in biomass transformation, as well as specific novel polypeptides and cell-free systems containing polypeptide combinations discovered by such methods are disclosed.

In situ targeting TEM8 via immune response and polypeptide recognition by wavelength-modulated surface plasmon resonance biosensor

PubMed Central

Wang, Yimin; Luo, Zewei; Liu, Kunping; Wang, Jie; Duan, Yixiang

2016-01-01

There is an increasing interest in real-time and in situ monitoring of living cell activities in life science and medicine. This paper reports a whole cell sensing protocol over the interface of Au film coupled in a wavelength-modulated surface plasmon resonance (WMSPR) biosensor. With dual parabolic mirrors integrated in the sensor, the compact and miniaturized instrument shows satisfactory refractive index sensitivity (2220 nm/RIU) and a high resolution of resonance wavelength shift of 0.3 nm to liquid samples. The affinity interactions between the biomarker of human tumor endothelial marker 8 (TEM8) and antibody (Ab) or specific polypeptide (PEP) were firstly introduced to WMSPR biosensor analysis. Both the interaction events of Ab-cell and PEP-cell over the Au film interface can be recognized by the sensor and the balance time of interactions is about 20 min. The concentration range of Ab for quantitative monitoring of the TEM8 expression on human colon carcinoma SW620 cells was investigated. The present low-cost and time-saving method provides a time resolution of binding specificity between Ab/PEP and TEM8 for real-time analysis of antigen on living tumor cell surface. PMID:26822761

Registration of Azov meadow fescue

USDA-ARS?s Scientific Manuscript database

'Azov' meadow fescue [Schedonorus pratensis (Huds.) P. Beauv.; syn. Festuca pratensis Huds.; syn. Lolium pratense (Huds.) Darbysh.] is a synthetic population originating from 1000 parental genotypes. The parents of Azov were selected from ten Russian plant introductions, mostly originating from the...

Assessment of brain metabolite correlates of adeno-associated virus-mediated over-expression of human alpha-synuclein in cortical neurons by in vivo (1) H-MR spectroscopy at 9.4 T.

PubMed

Cuellar-Baena, Sandra; Landeck, Natalie; Sonnay, Sarah; Buck, Kerstin; Mlynarik, Vladimir; In 't Zandt, René; Kirik, Deniz

2016-06-01

In this study, we used proton-localized spectroscopy ((1) H-MRS) for the acquisition of the neurochemical profile longitudinally in a novel rat model of human wild-type alpha-synuclein (α-syn) over-expression. Our goal was to find out if the increased α-syn load in this model could be linked to changes in metabolites in the frontal cortex. Animals injected with AAV vectors encoding for human α-syn formed the experimental group, whereas green fluorescent protein expressing animals were used as the vector-treated control group and a third group of uninjected animals were used as naïve controls. Data were acquired at 2, 4, and 8 month time points. Nineteen metabolites were quantified in the MR spectra using LCModel software. On the basis of 92 spectra, we evaluated any potential gender effect and found that lactate (Lac) levels were lower in males compared to females, while the opposite was observed for ascorbate (Asc). Next, we assessed the effect of age and found increased levels of GABA, Tau, and GPC+PCho. Finally, we analyzed the effect of treatment and found that Lac levels (p = 0.005) were specifically lower in the α-syn group compared to the green fluorescent protein and control groups. In addition, Asc levels (p = 0.05) were increased in the vector-injected groups, whereas glucose levels remained unchanged. This study indicates that the metabolic switch between glucose-lactate could be detectable in vivo and might be modulated by Asc. No concomitant changes were found in markers of neuronal integrity (e.g., N-acetylaspartate) consistent with the fact that α-syn over-expression in cortical neurons did not result in neurodegeneration in this model. We acquired the neurochemical profile longitudinally in a rat model of human wild-type alpha-synuclein (α-syn) over-expression in cortical neurons. We found that Lactate levels were reduced in the α-syn group compared to the control groups and Ascorbate levels were increased in the vector-injected groups. No changes were found in markers of neuronal integrity consistent with the fact that α-syn over-expression did not result in frank neurodegeneration. © 2016 International Society for Neurochemistry.

An energy function for dynamics simulations of polypeptides in torsion angle space

NASA Astrophysics Data System (ADS)

Sartori, F.; Melchers, B.; Böttcher, H.; Knapp, E. W.

1998-05-01

Conventional simulation techniques to model the dynamics of proteins in atomic detail are restricted to short time scales. A simplified molecular description, in which high frequency motions with small amplitudes are ignored, can overcome this problem. In this protein model only the backbone dihedrals φ and ψ and the χi of the side chains serve as degrees of freedom. Bond angles and lengths are fixed at ideal geometry values provided by the standard molecular dynamics (MD) energy function CHARMM. In this work a Monte Carlo (MC) algorithm is used, whose elementary moves employ cooperative rotations in a small window of consecutive amide planes, leaving the polypeptide conformation outside of this window invariant. A single of these window MC moves generates local conformational changes only. But, the application of many such moves at different parts of the polypeptide backbone leads to global conformational changes. To account for the lack of flexibility in the protein model employed, the energy function used to evaluate conformational energies is split into sequentially neighbored and sequentially distant contributions. The sequentially neighbored part is represented by an effective (φ,ψ)-torsion potential. It is derived from MD simulations of a flexible model dipeptide using a conventional MD energy function. To avoid exaggeration of hydrogen bonding strengths, the electrostatic interactions involving hydrogen atoms are scaled down at short distances. With these adjustments of the energy function, the rigid polypeptide model exhibits the same equilibrium distributions as obtained by conventional MD simulation with a fully flexible molecular model. Also, the same temperature dependence of the stability and build-up of α helices of 18-alanine as found in MD simulations is observed using the adapted energy function for MC simulations. Analyses of transition frequencies demonstrate that also dynamical aspects of MD trajectories are faithfully reproduced. Finally, it is demonstrated that even for high temperature unfolded polypeptides the MC simulation is more efficient by a factor of 10 than conventional MD simulations.

Tracking polypeptide folds on the free energy surface: effects of the chain length and sequence.

PubMed

Brukhno, Andrey V; Ricchiuto, Piero; Auer, Stefan

2012-07-26

Characterization of the folding transition in polypeptides and assessing the thermodynamic stability of their structured folds are of primary importance for approaching the problem of protein folding. We use molecular dynamics simulations for a coarse grained polypeptide model in order to (1) obtain the equilibrium conformation diagram of homopolypeptides in a broad range of the chain lengths, N = 10, ..., 100, and temperatures, T (in a multicanonical ensemble), and (2) determine free energy profiles (FEPs) projected onto an optimal, so-called "natural", reaction coordinate that preserves the height of barriers and the diffusion coefficients on the underlying free energy hyper-surface. We then address the following fundamental questions. (i) How well does a kinetically determined free energy landscape of a single chain represent the polypeptide equilibrium (ensemble) behavior? In particular, under which conditions might the correspondence be lost, and what are the possible implications for the folding processes? (ii) How does the free energy landscape depend on the chain length (homopolypeptides) and the monomer interaction sequence (heteropolypeptides)? Our data reveal that at low T values equilibrium structures adopted by relatively short homopolypeptides (N < 60) are dominated by α-helical folds which correspond to the primary and secondary minima of the FEP. In contrast, longer homopolypeptides (N > 70), upon quasi-equilibrium cooling, fold preferentially in β-bundles with small helical portions, while the FEPs exhibit no distinct global minima. Moreover, subject to the choice of the initial configuration, at sufficiently low T, essentially metastable structures can be found and prevail far from the true thermodynamic equilibrium. We also show that, by sequence-enabling the polypeptide model, it is possible to restrict the chain to a very specific part of the configuration space, which results in substantial simplification and smoothing of the free energy landscape as compared to the case of the corresponding homopolypeptide.

Interaction of fluvastatin with the liver-specific Na+ -dependent taurocholate cotransporting polypeptide (NTCP).

PubMed

Greupink, Rick; Dillen, Lieve; Monshouwer, Mario; Huisman, Maarten T; Russel, Frans G M

2011-11-20

It has been reported that polymorphisms in the organic anion transporting polypeptide 1B1 (OATP1B1, SLCO1B1) result in decreased hepatic uptake of simvastatin carboxy acid, the active metabolite of simvastatin. This is not the case for fluvastatin and it has been hypothesized that for this drug other hepatic uptake pathways exist. Here, we studied whether Na(+)-dependent taurocholate co-transporting polypeptide (NTCP, SLC10A1) can be an alternative hepatic uptake route for fluvastatin. Chinese Hamster Ovary cells transfected with human NTCP (CHO-NTCP) were used to investigate the inhibitory effect of fluvastatin and other statins on [(3)H]-taurocholic acid uptake ([(3)H]-TCA). Statin uptake by CHO-NTCP and cryopreserved human hepatocytes was assessed via LC-MS/MS. Fluvastatin appeared to be a potent and competitive inhibitor of [(3)H]-TCA uptake (IC(50) of 40μM), pointing to an interaction at the level of the bile acid binding pocket of NTCP. The inhibitory action of other statins was also studied, which revealed that statin inhibitory potency increased with molecular descriptors of lipophilicity: calculated logP (r(2)=0.82, p=0.034), logD(7.4) (r(2)=0.77, p=0.0001). Studies in CHO-NTCP cells showed that fluvastatin was indeed an NTCP substrate (K(m) 250±30μM, V(max) 1340±50ng/mg total cell protein/min). However, subsequent studies revealed that at clinically relevant plasma concentrations, NTCP contributed minimally to overall accumulation in human hepatocytes. In conclusion, fluvastatin interacts with NTCP at the level of the bile acid binding pocket and is an NTCP substrate. However, under normal conditions, NTCP-mediated uptake of this drug seems not to be a significant hepatocellular uptake pathway. Copyright © 2011 Elsevier B.V. All rights reserved.

Hybrid polypeptides having cellobiohydrolase activity and polynucleotides encoding same

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Ye; Shaghasi, Tarana

The present invention relates to hybrid polypeptides having cellobiohydrolase activity. The present invention also relates to polynucleotides encoding the hybrid polypeptides; nucleic acid constructs, vectors, and host cells comprising the polynucleotides; and processes of using the hybrid polypeptides.

Polypeptides having cellulolytic enhancing activity and polynucleotides encoding same

DOEpatents

Zhang, Yu; Duan, Junxin; Tang, Lan; Wu, Wenping

2015-06-09

Provided are isolated polypeptides having cellulolytic enhancing activity and isolated polynucleotides encoding the polypeptides. Also provided are nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.

Polypeptides having endoglucanase activity and polynucleotides encoding same

DOEpatents

Liu, Ye; Duan, Junxin; Tang, Lan

2015-09-22

The present invention provides isolated polypeptides having endoglucanase activity and isolated polynucleotides encoding the polypeptides. The invention also provides nucleic acid constructs, vectors, and host cell comprising the polynucleotides as well as methods of producing and using the polypeptides.

Polypeptides having cellobiohydrolase activitiy and polynucleotides encoding same

DOEpatents

Liu, Ye; Tang, Lan; Duan, Junxin

2015-12-15

The present invention provides isolated polypeptides having cellobiohydrolase activity and isolated polynucleotides encoding the polypeptides. The invention also provides nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.

Isolation of Polypeptide Sample and Measurement of Its Concentration.

ERIC Educational Resources Information Center

Beanan, Maureen J.

2000-01-01

Introduces a laboratory experiment that isolates a bacterial polypeptide sample and measures the concentration of polypeptides in the sample. Uses Escherichia coli strain MM294 and performs a bio-rad assay to determine the concentration of polypeptides. (YDS)

Polypeptides having cellobiohydrolase activity and polynucleotides encoding same

DOEpatents

Liu, Ye; Tang, Lan

2015-07-14

The present invention provides isolated polypeptides having cellobiohydrolase activity and isolated polynucleotides encoding the polypeptides. The invention also provides nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.

Screening the Biosphere: The Fungicolous Fungus Trichoderma phellinicola, a Prolific Source of Hypophellins, New 17-, 18-, 19-, and 20-Residue Peptaibiotics1)

PubMed Central

Röhrich, Christian René; Iversen, Anita; Jaklitsch, Walter Michael; Voglmayr, Hermann; Vilcinskas, Andreas; Nielsen, Kristian Fog; Thrane, Ulf; von Döhren, Hans; Brückner, Hans; Degenkolb, Thomas

2013-01-01

To investigate the significance of antibiotics for the producing organism(s) in the natural habitat, we screened a specimen of the fungicolous fungus Trichoderma phellinicola (syn. Hypocrea phellinicola) growing on its natural host Phellinus ferruginosus. Results revealed that a particular group of non-ribosomal antibiotic polypeptides, peptaibiotics, which contain the non-proteinogenic marker amino acid, α-aminoisobutyric acid, was biosynthesized in the natural habitat by the fungicolous producer and, consequently, released into the host. By means of liquid chromatography coupled to electrospray high-resolution time-of-flight mass spectrometry, we detected ten 20-residue peptaibols in the specimen. Sequences of peptaibiotics found in vivo were independently confirmed by analyzing the peptaibiome of an agar plate culture of T. phellinicola CBS 119283 (ex-type) grown under laboratory conditions. Notably, this strain could be identified as a potent producer of 39 new 17-, 18-, and 19-residue peptaibiotics, which display the same building scheme as the 20-residue peptaibols found in the specimen. Two of the 19-residue peptaibols are tentatively assigned to carry tyrosinol, a novel C-terminal residue, as deduced from high-resolution tandem mass-spectrometry data. For the new peptaibiotics produced by T. phellinicola, the name ‘hypophellin(s)’, based on the teleomorph name, is introduced. PMID:23681726

Experience with the SynCardia total artificial heart in a Canadian centre

PubMed Central

Nguyen, Anthony; Pellerin, Michel; Perrault, Louis P.; White, Michel; Ducharme, Anique; Racine, Normand; Carrier, Michel

2017-01-01

Background The SynCardia total artificial heart (TAH) provides complete circulatory support by replacing both native ventricles. Accepted indications include bridge to transplantation and destination therapy. We review our experience with TAH implantation during a period when axial flow pump became available. Methods We retrospectively analyzed the demographics, clinical characteristics and survival of all patients receiving the TAH. Results From September 2004 to November 2016, 13 patients (12 men, mean age 45 ± 13 yr) received the TAH for refractory cardiogenic shock secondary to idiopathic (56%) or ischemic (17%) cardiomyopathy and to other various causes (33%). Before implantation, mean ejection fraction was 14% ± 4%, 7 (54%) patients had previous cardiac surgery, 4 (31%) were on mechanical ventilation, and 3 (23%) patients were on dialysis. The mean duration of TAH support was 46 ± 40 days. Three (23%) patients died while on support after a mean of 15 days. Actuarial survival on support was 77% ± 12% at 30 days after implantation. Complications on support included stroke (n = 1, 8%), acute respiratory distress syndrome requiring prolonged intubation (n = 5, 38%) and acute renal failure requiring temporary dialysis (n = 5, 38%). Ten (77%) patients survived to be transplanted after a mean of 52 ± 42 days of support. Actuarial survival rates after transplant were 67% ± 16% at 1 month and 56% ± 17% at 1 year after transplantation. Conclusion The TAH provides an alternative with low incidence of neurologic events in extremely fragile and complex patients waiting for heart transplantation. Complex and unusual anatomic conditions explained the current use of TAH. PMID:28930049

Experience with the SynCardia total artificial heart in a Canadian centre.

PubMed

Nguyen, Anthony; Pellerin, Michel; Perrault, Louis P; White, Michel; Ducharme, Anique; Racine, Normand; Carrier, Michel

2017-12-01

The SynCardia total artificial heart (TAH) provides complete circulatory support by replacing both native ventricles. Accepted indications include bridge to transplantation and destination therapy. We review our experience with TAH implantation during a period when axial flow pump became available. We retrospectively analyzed the demographics, clinical characteristics and survival of all patients receiving the TAH. From September 2004 to November 2016, 13 patients (12 men, mean age 45 ± 13 yr) received the TAH for refractory cardiogenic shock secondary to idiopathic (56%) or ischemic (17%) cardiomyopathy and to other various causes (33%). Before implantation, mean ejection fraction was 14% ± 4%, 7 (54%) patients had previous cardiac surgery, 4 (31%) were on mechanical ventilation, and 3 (23%) patients were on dialysis. The mean duration of TAH support was 46 ± 40 days. Three (23%) patients died while on support after a mean of 15 days. Actuarial survival on support was 77% ± 12% at 30 days after implantation. Complications on support included stroke ( n = 1, 8%), acute respiratory distress syndrome requiring prolonged intubation ( n = 5, 38%) and acute renal failure requiring temporary dialysis ( n = 5, 38%). Ten (77%) patients survived to be transplanted after a mean of 52 ± 42 days of support. Actuarial survival rates after transplant were 67% ± 16% at 1 month and 56% ± 17% at 1 year after transplantation. The TAH provides an alternative with low incidence of neurologic events in extremely fragile and complex patients waiting for heart transplantation. Complex and unusual anatomic conditions explained the current use of TAH.

Polypeptides having cellobiohydrolase activity and polynucleotides encoding same

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Ye; Tang, Lan; Duan, Junxin

The present invention relates to isolated polypeptides having cellobiohydrolase activity and isolated polynucleotides encoding the polypeptides. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.

Polypeptides having xylanase activity and polynucleotides encoding same

DOE Office of Scientific and Technical Information (OSTI.GOV)

Spodsberg, Nikolaj

The present invention relates to isolated polypeptides having xylanase activity and polynucleotides encoding the polypeptides. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.

Polypeptides having xylanase activity and polynucleotides encoding same

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lopez de Leon, Alfredo; Rey, Michael

The present invention relates to isolated polypeptides having xylanase activity and isolated polynucleotides encoding the polypeptides. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.

Polypeptides having cellobiohydrolase activity and polynucleotides encoding same

DOE Office of Scientific and Technical Information (OSTI.GOV)

Spodsberg, Nikolaj

The present invention relates to isolated polypeptides having cellobiohydrolase activity and polynucleotides encoding the polypeptides. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.

Polypeptides having endoglucanase activity and polynucleotides encoding same

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhang, Yu; Liu, Ye; Duan, Junxin

The present invention relates to isolated polypeptides having endoglucanase activity and isolated polynucleotides encoding the polypeptides. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.

Polypeptides having endoglucanase activity and polynucleotides encoding same

DOEpatents

Lopez de Leon, Alfredo; Rey, Michael

2012-09-18

The present invention relates to isolated polypeptides having endoglucanase activity and isolated polynucleotides encoding the polypeptides. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.

Polypeptides having xylanase activity and polynucleotides encoding same

DOEpatents

Lopez de Leon, Alfredo; Rey, Michael

2010-12-14

The present invention relates to isolated polypeptides having xylanase activity and isolated polynucleotides encoding the polypeptides. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.

Polypeptides having endoglucanase activity and polynucleotides encoding same

DOEpatents

Harris, Paul [Carnation, WA; Lopez de Leon, Alfredo [Davis, CA; Rey, Micheal [Davis, CA; Ding, Hanshu [Davis, CA; Vlasenko, Elena [Davis, CA

2012-02-21

The present invention relates to isolated polypeptides having endoglucanase activity and isolated polynucleotides encoding the polypeptides. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods for producing and using the polypeptides.

Polypeptides having cellobiohydrolase activity and polynucleotides encoding same

DOEpatents

Spodsberg, Nikolaj

2016-06-28

The present invention relates to isolated polypeptides having cellobiohydrolase activity and polynucleotides encoding the polypeptides. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.