A comparison of the interaction of nitric oxide with the heteropolytungstic acids H{sub 3}PW{sub 12}O{sub 40}, H{sub 0.5}Cs{sub 2.5}PW{sub 12}O{sub 40}, HMgPW{sub 12}O{sub 40}, H{sub 8}SiW{sub 11}O{sub 38}, H{sub 4}SiW{sub 12}O{sub 40}, and H{sub 10}CoW{sub 12}O{sub 42}

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

Herring, A.M.; McCormick, R.L.; Boonrueng, S.R.

2000-05-18

The interaction between NO and the heteropolytungstic acids (HPAs) H{sub 3}PW{sub 12}O{sub 40} (HPW), H{sub 0.5}Cs{sub 2.5}PW{sub 12}O{sub 40} (HCsPW), HMgPW{sub 12}O{sub 40} (HMgPW), H{sub 8}SiW{sub 11}O{sub 38} (HSiW{sub 11}), H{sub 4}SiW{sub 12}O{sub 40} (HSiW), and H{sub 10}CoW{sub 12}{sub 42}(HCoW) in the presence of O{sub 2}(g) was investigated. The tools employed were in situ diffuse reflectance infrared spectroscopy, X-ray powder diffraction, and solid-state {sup 1}H NMR. It was determined that protons may either be present in the HPAs secondary structure as anhydrous protons or be bound to one or two water molecules as H{sub 3}O{sup +} or H{sub 5}O{sub 2}{supmore » +}, respectively. A previous investigation found that HPW sorbed NO into its bulk structure as NOH{sup +}, whereas the anhydrous potassium salt of HPW exhibited weak chemisorption of NO on its surface. In the present study, it was found that NO chemisorbed weakly on the surface of the anhydrous HCsPW. For HMgPW and HSiW, IR and NMR evidence suggests that water is present as H{sub 3}O{sup +}, and the formation of surface-bound NOH{sup +} was observed. Inclusion of NO into the secondary structure as NOH{sup +} was observed for HPW, HSiW{sub 11}, and HCoW. Literature data for HPW, as well as IR and NMR results reported here, indicate that these HPAs contain water as H{sub 5}O{sub 2}{sup +}. The presence of H{sub 5}O{sub 2}{sup +} is, therefore, a prerequisite for NO incorporation as NOH{sup +} in the HPA secondary structure. These HPAs exhibited two modes of NO bonding in the secondary structure: a strongly hydrogen-bound hydrated form, NOH{sup +}{center_dot}H{sub 2}O, and a more weakly bound anhydrous form, NOH{sup +}. Evidence suggests that it is the hydrated form that decomposes, yielding N{sub 2} upon rapid heating. Both NO-containing species interact with terminal and corner-sharing oxygen atoms of the Keggin ion. Anions that are held apart by terminal oxygen-hydrogen bonds have a larger lattice parameter than those held apart by terminal oxygen-hydrogen and corner-sharing oxygen-nitrogen bonds. The lattice contraction upon NO incorporation implies activation of NO by an intimate interaction with oxygen of the Keggin anion.« less

Prediction of Long Loops with Embedded Secondary Structure using the Protein Local Optimization Program

PubMed Central

Miller, Edward B.; Murrett, Colleen S.; Zhu, Kai; Zhao, Suwen; Goldfeld, Dahlia A.; Bylund, Joseph H.; Friesner, Richard A.

2013-01-01

Robust homology modeling to atomic-level accuracy requires in the general case successful prediction of protein loops containing small segments of secondary structure. Further, as loop prediction advances to success with larger loops, the exclusion of loops containing secondary structure becomes awkward. Here, we extend the applicability of the Protein Local Optimization Program (PLOP) to loops up to 17 residues in length that contain either helical or hairpin segments. In general, PLOP hierarchically samples conformational space and ranks candidate loops with a high-quality molecular mechanics force field. For loops identified to possess α-helical segments, we employ an alternative dihedral library composed of (ϕ,ψ) angles commonly found in helices. The alternative library is searched over a user-specified range of residues that define the helical bounds. The source of these helical bounds can be from popular secondary structure prediction software or from analysis of past loop predictions where a propensity to form a helix is observed. Due to the maturity of our energy model, the lowest energy loop across all experiments can be selected with an accuracy of sub-Ångström RMSD in 80% of cases, 1.0 to 1.5 Å RMSD in 14% of cases, and poorer than 1.5 Å RMSD in 6% of cases. The effectiveness of our current methods in predicting hairpin-containing loops is explored with hairpins up to 13 residues in length and again reaching an accuracy of sub-Ångström RMSD in 83% of cases, 1.0 to 1.5 Å RMSD in 10% of cases, and poorer than 1.5 Å RMSD in 7% of cases. Finally, we explore the effect of an imprecise surrounding environment, in which side chains, but not the backbone, are initially in perturbed geometries. In these cases, loops perturbed to 3Å RMSD from the native environment were restored to their native conformation with sub-Ångström RMSD. PMID:23814507

The secondary structure of the ets domain of human Fli-1 resembles that of the helix-turn-helix DNA-binding motif of the Escherichia coli catabolite gene activator protein.

PubMed Central

Liang, H; Olejniczak, E T; Mao, X; Nettesheim, D G; Yu, L; Thompson, C B; Fesik, S W

1994-01-01

The ets family of eukaryotic transcription factors is characterized by a conserved DNA-binding domain of approximately 85 amino acids for which the three-dimensional structure is not known. By using multidimensional NMR spectroscopy, we have determined the secondary structure of the ets domain of one member of this gene family, human Fli-1, both in the free form and in a complex with a 16-bp cognate DNA site. The secondary structure of the Fli-1 ets domain consists of three alpha-helices and a short four-stranded antiparallel beta-sheet. This secondary structure arrangement resembles that of the DNA-binding domain of the catabolite gene activator protein of Escherichia coli, as well as those of several eukaryotic DNA-binding proteins including histone H5, HNF-3/fork head, and the heat shock transcription factor. Differences in chemical shifts of backbone resonances and amide exchange rates between the DNA-bound and free forms of the Fli-1 ets domain suggest that the third helix is the DNA recognition helix, as in the catabolite gene activator protein and other structurally related proteins. These results suggest that the ets domain is structurally similar to the catabolite gene activator protein family of helix-turn-helix DNA-binding proteins. Images PMID:7972119

Influence of phosphocholine alkyl chain length on peptide-micelle interactions and micellar size and shape.

PubMed

Göbl, Christoph; Dulle, Martin; Hohlweg, Walter; Grossauer, Jörg; Falsone, S Fabio; Glatter, Otto; Zangger, Klaus

2010-04-08

The interaction with biological membranes is of functional importance for many peptides and proteins. Structural studies on such membrane-bound biomacromolecules are often carried out in solutions containing small membrane-mimetic assemblies of detergent molecules. To investigate the influence of the hydrophobic chain length on the structure, diffusional and dynamical behavior of a peptide bound to micelles, we studied the binding of three peptides to n-phosphocholines with n ranging from 8 to 16. The peptides studied are the 15 residue antimicrobial peptide CM15, the 25-residue transmembrane helix 7 of yeast V-ATPase (TM7), and the 35-residue bacterial toxin LdrD. To keep the dimension of the peptide-membrane-mimetic assembly small, micelles are typically used when studying membrane-bound peptides and proteins, for example, by solution NMR spectroscopy. Since they are readily available in deuterated form most often sodium-dodecylsulfate (SDS) and dodecylphosphocholine (DPC) are used as the micelle-forming detergent. Using NMR, CD, and SAXS, we found that all phosphocholines studied form spherical micelles in the presence and absence of small bound peptides and the diameters of the micelles are basically unchanged upon peptide binding. The size of the peptide relative to the micelle determines to what extent the secondary structure can form. For small peptides (up to approximately 25 residues) the use of shorter chain phosphocholines is recommended for solution NMR studies due to the favorable spectral quality and since they are as well-structured as in DPC. In contrast, larger peptides are better structured in micelles formed by detergents with chain lengths longer than DPC.

The crystal structure of the Sox4 HMG domain-DNA complex suggests a mechanism for positional interdependence in DNA recognition.

PubMed

Jauch, Ralf; Ng, Calista K L; Narasimhan, Kamesh; Kolatkar, Prasanna R

2012-04-01

It has recently been proposed that the sequence preferences of DNA-binding TFs (transcription factors) can be well described by models that include the positional interdependence of the nucleotides of the target sites. Such binding models allow for multiple motifs to be invoked, such as principal and secondary motifs differing at two or more nucleotide positions. However, the structural mechanisms underlying the accommodation of such variant motifs by TFs remain elusive. In the present study we examine the crystal structure of the HMG (high-mobility group) domain of Sox4 [Sry (sex-determining region on the Y chromosome)-related HMG box 4] bound to DNA. By comparing this structure with previously solved structures of Sox17 and Sox2, we observed subtle conformational differences at the DNA-binding interface. Furthermore, using quantitative electrophoretic mobility-shift assays we validated the positional interdependence of two nucleotides and the presence of a secondary Sox motif in the affinity landscape of Sox4. These results suggest that a concerted rearrangement of two interface amino acids enables Sox4 to accommodate primary and secondary motifs. The structural adaptations lead to altered dinucleotide preferences that mutually reinforce each other. These analyses underline the complexity of the DNA recognition by TFs and provide an experimental validation for the conceptual framework of positional interdependence and secondary binding motifs.

Solution structural ensembles of substrate-free cytochrome P450(cam).

PubMed

Asciutto, Eliana K; Young, Matthew J; Madura, Jeffry; Pochapsky, Susan Sondej; Pochapsky, Thomas C

2012-04-24

Removal of substrate (+)-camphor from the active site of cytochrome P450(cam) (CYP101A1) results in nuclear magnetic resonance-detected perturbations in multiple regions of the enzyme. The (1)H-(15)N correlation map of substrate-free diamagnetic Fe(II) CO-bound CYP101A permits these perturbations to be mapped onto the solution structure of the enzyme. Residual dipolar couplings (RDCs) were measured for (15)N-(1)H amide pairs in two independent alignment media for the substrate-free enzyme and used as restraints in solvated molecular dynamics (MD) simulations to generate an ensemble of best-fit structures of the substrate-free enzyme in solution. Nuclear magnetic resonance-detected chemical shift perturbations reflect changes in the electronic environment of the NH pairs, such as hydrogen bonding and ring current shifts, and are observed for residues in the active site as well as in hinge regions between secondary structural features. RDCs provide information about relative orientations of secondary structures, and RDC-restrained MD simulations indicate that portions of a β-rich region adjacent to the active site shift so as to partially occupy the vacancy left by removal of the substrate. The accessible volume of the active site is reduced in the substrate-free enzyme relative to the substrate-bound structure calculated using the same methods. Both symmetric and asymmetric broadening of multiple resonances observed upon substrate removal as well as localized increased errors in RDC fits suggest that an ensemble of enzyme conformations are present in the substrate-free form.

Entropic (de)stabilization of surface-bound peptides conjugated with polymers

NASA Astrophysics Data System (ADS)

Carmichael, Scott P.; Shell, M. Scott

2015-12-01

In many emerging biotechnologies, functional proteins must maintain their native structures on or near interfaces (e.g., tethered peptide arrays, protein coated nanoparticles, and amphiphilic peptide micelles). Because the presence of a surface is known to dramatically alter the thermostability of tethered proteins, strategies to stabilize surface-bound proteins are highly sought. Here, we show that polymer conjugation allows for significant control over the secondary structure and thermostability of a model surface-tethered peptide. We use molecular dynamics simulations to examine the folding behavior of a coarse-grained helical peptide that is conjugated to polymers of various lengths and at various conjugation sites. These polymer variations reveal surprisingly diverse behavior, with some stabilizing and some destabilizing the native helical fold. We show that ideal-chain polymer entropies explain these varied effects and can quantitatively predict shifts in folding temperature. We then develop a generic theoretical model, based on ideal-chain entropies, that predicts critical lengths for conjugated polymers to effect changes in the folding of a surface-bound protein. These results may inform new design strategies for the stabilization of surface-associated proteins important for a range technological applications.

Entropic (de)stabilization of surface-bound peptides conjugated with polymers.

PubMed

Carmichael, Scott P; Shell, M Scott

2015-12-28

In many emerging biotechnologies, functional proteins must maintain their native structures on or near interfaces (e.g., tethered peptide arrays, protein coated nanoparticles, and amphiphilic peptide micelles). Because the presence of a surface is known to dramatically alter the thermostability of tethered proteins, strategies to stabilize surface-bound proteins are highly sought. Here, we show that polymer conjugation allows for significant control over the secondary structure and thermostability of a model surface-tethered peptide. We use molecular dynamics simulations to examine the folding behavior of a coarse-grained helical peptide that is conjugated to polymers of various lengths and at various conjugation sites. These polymer variations reveal surprisingly diverse behavior, with some stabilizing and some destabilizing the native helical fold. We show that ideal-chain polymer entropies explain these varied effects and can quantitatively predict shifts in folding temperature. We then develop a generic theoretical model, based on ideal-chain entropies, that predicts critical lengths for conjugated polymers to effect changes in the folding of a surface-bound protein. These results may inform new design strategies for the stabilization of surface-associated proteins important for a range technological applications.

Hepatitis Delta Antigen Requires a Flexible Quasi-Double-Stranded RNA Structure To Bind and Condense Hepatitis Delta Virus RNA in a Ribonucleoprotein Complex

PubMed Central

Griffin, Brittany L.; Chasovskikh, Sergey; Dritschilo, Anatoly

2014-01-01

ABSTRACT The circular genome and antigenome RNAs of hepatitis delta virus (HDV) form characteristic unbranched, quasi-double-stranded RNA secondary structures in which short double-stranded helical segments are interspersed with internal loops and bulges. The ribonucleoprotein complexes (RNPs) formed by these RNAs with the virus-encoded protein hepatitis delta antigen (HDAg) perform essential roles in the viral life cycle, including viral replication and virion formation. Little is understood about the formation and structure of these complexes and how they function in these key processes. Here, the specific RNA features required for HDAg binding and the topology of the complexes formed were investigated. Selective 2′OH acylation analyzed by primer extension (SHAPE) applied to free and HDAg-bound HDV RNAs indicated that the characteristic secondary structure of the RNA is preserved when bound to HDAg. Notably, the analysis indicated that predicted unpaired positions in the RNA remained dynamic in the RNP. Analysis of the in vitro binding activity of RNAs in which internal loops and bulges were mutated and of synthetically designed RNAs demonstrated that the distinctive secondary structure, not the primary RNA sequence, is the major determinant of HDAg RNA binding specificity. Atomic force microscopy analysis of RNPs formed in vitro revealed complexes in which the HDV RNA is substantially condensed by bending or wrapping. Our results support a model in which the internal loops and bulges in HDV RNA contribute flexibility to the quasi-double-stranded structure that allows RNA bending and condensing by HDAg. IMPORTANCE RNA-protein complexes (RNPs) formed by the hepatitis delta virus RNAs and protein, HDAg, perform critical roles in virus replication. Neither the structures of these RNPs nor the RNA features required to form them have been characterized. HDV RNA is unusual in that it forms an unbranched quasi-double-stranded structure in which short base-paired segments are interspersed with internal loops and bulges. We analyzed the role of the HDV RNA sequence and secondary structure in the formation of a minimal RNP and visualized the structure of this RNP using atomic force microscopy. Our results indicate that HDAg does not recognize the primary sequence of the RNA; rather, the principle contribution of unpaired bases in HDV RNA to HDAg binding is to allow flexibility in the unbranched quasi-double-stranded RNA structure. Visualization of RNPs by atomic force microscopy indicated that the RNA is significantly bent or condensed in the complex. PMID:24741096

Intermixed adatom and surface-bound adsorbates in regular self-assembled monolayers of racemic 2-butanethiol on Au(111).

PubMed

Ouyang, Runhai; Yan, Jiawei; Jensen, Palle S; Ascic, Erhad; Gan, Shiyu; Tanner, David; Mao, Bingwei; Niu, Li; Zhang, Jingdong; Tang, Chunguang; Hush, Noel S; Reimers, Jeffrey R; Ulstrup, Jens

2015-04-07

In situ scanning tunneling microscopy combined with density functional theory molecular dynamics simulations reveal a complex structure for the self-assembled monolayer (SAM) of racemic 2-butanethiol on Au(111) in aqueous solution. Six adsorbate molecules occupy a (10×√3)R30° cell organized as two RSAuSR adatom-bound motifs plus two RS species bound directly to face-centered-cubic and hexagonally close-packed sites. This is the first time that these competing head-group arrangements have been observed in the same ordered SAM. Such unusual packing is favored as it facilitates SAMs with anomalously high coverage (30%), much larger than that for enantiomerically resolved 2-butanethiol or secondary-branched butanethiol (25%) and near that for linear-chain 1-butanethiol (33%). © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Analysis of sequencing data for probing RNA secondary structures and protein-RNA binding in studying posttranscriptional regulations.

PubMed

Hu, Xihao; Wu, Yang; Lu, Zhi John; Yip, Kevin Y

2016-11-01

High-throughput sequencing has been used to study posttranscriptional regulations, where the identification of protein-RNA binding is a major and fast-developing sub-area, which is in turn benefited by the sequencing methods for whole-transcriptome probing of RNA secondary structures. In the study of RNA secondary structures using high-throughput sequencing, bases are modified or cleaved according to their structural features, which alter the resulting composition of sequencing reads. In the study of protein-RNA binding, methods have been proposed to immuno-precipitate (IP) protein-bound RNA transcripts in vitro or in vivo By sequencing these transcripts, the protein-RNA interactions and the binding locations can be identified. For both types of data, read counts are affected by a combination of confounding factors, including expression levels of transcripts, sequence biases, mapping errors and the probing or IP efficiency of the experimental protocols. Careful processing of the sequencing data and proper extraction of important features are fundamentally important to a successful analysis. Here we review and compare different experimental methods for probing RNA secondary structures and binding sites of RNA-binding proteins (RBPs), and the computational methods proposed for analyzing the corresponding sequencing data. We suggest how these two types of data should be integrated to study the structural properties of RBP binding sites as a systematic way to better understand posttranscriptional regulations. © The Author 2015. Published by Oxford University Press. For Permissions, please email: journals.permissions@oup.com.

Identification and tracking of hairpin vortex auto-generation in turbulent wall-bounded flow

NASA Astrophysics Data System (ADS)

Huang, Yangzi; Green, Melissa

2016-11-01

Hairpin vortices have been widely accepted as component structures of turbulent boundary layers. Their properties (size, vorticity, energy) and dynamic phenomena (origin, growth, breakdown) have been shown to correlate to the complex, multi-scaled turbulent motions observed in both experiments and simulations. As established in the literature, the passage of a hairpin vortex creates a wall-normal ejection of fluid, which encounters the high-speed freestream resulting in near-wall shear and increased drag. A previously generated simulation of an isolated hairpin vortex is used to study the auto-generation of a secondary vortex structure. Eulerian methods such as the Q criterion and Γ2 function, as well as Lagrangian methods are used to visualize the three-dimensional hairpin vortices and the auto-generation process. The circulation development and wall-normal location of both primary and secondary hairpin heads are studied to determine if there is a correlation between the strength and height of the primary hairpin vortex with the secondary hairpin vortex auto-generation.

Substrate-bound outward-open state of the betaine transporter BetP provides insights into Na+ coupling

NASA Astrophysics Data System (ADS)

Perez, Camilo; Faust, Belinda; Mehdipour, Ahmad Reza; Francesconi, Kevin A.; Forrest, Lucy R.; Ziegler, Christine

2014-07-01

The Na+-coupled betaine symporter BetP shares a highly conserved fold with other sequence unrelated secondary transporters, for example, with neurotransmitter symporters. Recently, we obtained atomic structures of BetP in distinct conformational states, which elucidated parts of its alternating-access mechanism. Here, we report a structure of BetP in a new outward-open state in complex with an anomalous scattering substrate, adding a fundamental piece to an unprecedented set of structural snapshots for a secondary transporter. In combination with molecular dynamics simulations these structural data highlight important features of the sequential formation of the substrate and sodium-binding sites, in which coordinating water molecules play a crucial role. We observe a strictly interdependent binding of betaine and sodium ions during the coupling process. All three sites undergo progressive reshaping and dehydration during the alternating-access cycle, with the most optimal coordination of all substrates found in the closed state.

Conformational changes in the expression domain of the Escherichia coli thiM riboswitch

PubMed Central

Rentmeister, Andrea; Mayer, Günter; Kuhn, Nicole; Famulok, Michael

2007-01-01

The thiM riboswitch contains an aptamer domain that adaptively binds the coenzyme thiamine pyrophosphate (TPP). The binding of TPP to the aptamer domain induces structural rearrangements that are relayed to a second domain, the so-called expression domain, thereby interfering with gene expression. The recently solved crystal structures of the aptamer domains of the thiM riboswitches in complex with TPP revealed how TPP stabilizes secondary and tertiary structures in the RNA ligand complex. To understand the global modes of reorganization between the two domains upon metabolite binding the structure of the entire riboswitch in presence and absence of TPP needs to be determined. Here we report the secondary structure of the entire thiM riboswitch from Escherichia coli in its TPP-free form and its transition into the TPP-bound variant, thereby depicting domains of the riboswitch that serve as communication links between the aptamer and the expression domain. Furthermore, structural probing provides an explanation for the lack of genetic control exerted by a riboswitch variant with mutations in the expression domain that still binds TPP. PMID:17517779

34 CFR 645.1 - What is the Upward Bound Program?

Code of Federal Regulations, 2010 CFR

2010-07-01

... participants the skills and motivation necessary to complete a program of secondary education and to enter and... the following three types of projects: (1) Regular Upward Bound projects. (2) Upward Bound Math and... Education Regulations of the Offices of the Department of Education (Continued) OFFICE OF POSTSECONDARY...

Induced secondary structure and polymorphism in an intrinsically disordered structural linker of the CNS: solid-state NMR and FTIR spectroscopy of myelin basic protein bound to actin.

PubMed

Ahmed, Mumdooh A M; Bamm, Vladimir V; Shi, Lichi; Steiner-Mosonyi, Marta; Dawson, John F; Brown, Leonid; Harauz, George; Ladizhansky, Vladimir

2009-01-01

The 18.5 kDa isoform of myelin basic protein (MBP) is a peripheral membrane protein that maintains the structural integrity of the myelin sheath of the central nervous system by conjoining the cytoplasmic leaflets of oligodendrocytes and by linking the myelin membrane to the underlying cytoskeleton whose assembly it strongly promotes. It is a multifunctional, intrinsically disordered protein that behaves primarily as a structural stabilizer, but with elements of a transient or induced secondary structure that represent binding sites for calmodulin or SH3-domain-containing proteins, inter alia. In this study we used solid-state NMR (SSNMR) and Fourier transform infrared (FTIR) spectroscopy to study the conformation of 18.5 kDa MBP in association with actin microfilaments and bundles. FTIR spectroscopy of fully (13)C,(15)N-labeled MBP complexed with unlabeled F-actin showed induced folding of both protein partners, viz., some increase in beta-sheet content in actin, and increases in both alpha-helix and beta-sheet content in MBP, albeit with considerable extended structure remaining. Solid-state NMR spectroscopy revealed that MBP in MBP-actin assemblies is structurally heterogeneous but gains ordered secondary structure elements (both alpha-helical and beta-sheet), particularly in the terminal fragments and in a central immunodominant epitope. The overall conformational polymorphism of MBP is consistent with its in vivo roles as both a linker (membranes and cytoskeleton) and a putative signaling hub.

Structural studies of Saccharomyces cerevesiae mitochondrial NADP-dependent isocitrate dehydrogenase in different enzymatic states reveal substantial conformational changes during the catalytic reaction.

PubMed

Peng, Yingjie; Zhong, Chen; Huang, Wei; Ding, Jianping

2008-09-01

Isocitrate dehydrogenases (IDHs) catalyze oxidative decarboxylation of isocitrate (ICT) into alpha-ketoglutarate (AKG). We report here the crystal structures of Saccharomyces cerevesiae mitochondrial NADP-IDH Idp1p in binary complexes with coenzyme NADP, or substrate ICT, or product AKG, and in a quaternary complex with NADPH, AKG, and Ca(2+), which represent different enzymatic states during the catalytic reaction. Analyses of these structures identify key residues involved in the binding of these ligands. Comparisons among these structures and with the previously reported structures of other NADP-IDHs reveal that eukaryotic NADP-IDHs undergo substantial conformational changes during the catalytic reaction. Binding or release of the ligands can cause significant conformational changes of the structural elements composing the active site, leading to rotation of the large domain relative to the small and clasp domains along two hinge regions (residues 118-124 and residues 284-287) while maintaining the integrity of its secondary structural elements, and thus, formation of at least three distinct overall conformations. Specifically, the enzyme adopts an open conformation when bound to NADP, a quasi-closed conformation when bound to ICT or AKG, and a fully closed conformation when bound to NADP, ICT, and Ca(2+) in the pseudo-Michaelis complex or with NADPH, AKG, and Ca(2+) in the product state. The conformational changes of eukaryotic NADP-IDHs are quite different from those of Escherichia coli NADP-IDH, for which significant conformational changes are observed only between two forms of the apo enzyme, suggesting that the catalytic mechanism of eukaryotic NADP-IDHs is more complex than that of EcIDH, and involves more fine-tuned conformational changes.

Structural studies of Saccharomyces cerevesiae mitochondrial NADP-dependent isocitrate dehydrogenase in different enzymatic states reveal substantial conformational changes during the catalytic reaction

PubMed Central

Peng, Yingjie; Zhong, Chen; Huang, Wei; Ding, Jianping

2008-01-01

Isocitrate dehydrogenases (IDHs) catalyze oxidative decarboxylation of isocitrate (ICT) into α-ketoglutarate (AKG). We report here the crystal structures of Saccharomyces cerevesiae mitochondrial NADP-IDH Idp1p in binary complexes with coenzyme NADP, or substrate ICT, or product AKG, and in a quaternary complex with NADPH, AKG, and Ca2+, which represent different enzymatic states during the catalytic reaction. Analyses of these structures identify key residues involved in the binding of these ligands. Comparisons among these structures and with the previously reported structures of other NADP-IDHs reveal that eukaryotic NADP-IDHs undergo substantial conformational changes during the catalytic reaction. Binding or release of the ligands can cause significant conformational changes of the structural elements composing the active site, leading to rotation of the large domain relative to the small and clasp domains along two hinge regions (residues 118–124 and residues 284–287) while maintaining the integrity of its secondary structural elements, and thus, formation of at least three distinct overall conformations. Specifically, the enzyme adopts an open conformation when bound to NADP, a quasi-closed conformation when bound to ICT or AKG, and a fully closed conformation when bound to NADP, ICT, and Ca2+ in the pseudo-Michaelis complex or with NADPH, AKG, and Ca2+ in the product state. The conformational changes of eukaryotic NADP-IDHs are quite different from those of Escherichia coli NADP-IDH, for which significant conformational changes are observed only between two forms of the apo enzyme, suggesting that the catalytic mechanism of eukaryotic NADP-IDHs is more complex than that of EcIDH, and involves more fine-tuned conformational changes. PMID:18552125

Conformational Analysis on structural perturbations of the zinc finger NEMO

NASA Astrophysics Data System (ADS)

Godwin, Ryan; Salsbury, Freddie; Salsbury Group Team

2014-03-01

The NEMO (NF-kB Essential Modulator) Zinc Finger protein (2jvx) is a functional Ubiquitin-binding domain, and plays a role in signaling pathways for immune/inflammatory responses, apoptosis, and oncogenesis [Cordier et al., 2008]. Characterized by 3 cysteines and 1 histidine residue at the active site, the biologically occurring, bound zinc configuration is a stable structural motif. Perturbations of the zinc binding residues suggest conformational changes in the 423-atom protein characterized via analysis of all-atom molecular dynamics simulations. Structural perturbations include simulations with and without a zinc ion and with and without de-protonated cysteines, resulting in four distinct configurations. Simulations of various time scales show consistent results, yet the longest, GPU driven, microsecond runs show more drastic structural and dynamic fluctuations when compared to shorter duration time-scales. The last cysteine residue (26 of 28) and the helix on which it resides exhibit a secondary, locally unfolded conformation in addition to its normal bound conformation. Combined analytics elucidate how the presence of zinc and/or protonated cysteines impact the dynamics and energetic fluctuations of NEMO. Comprehensive Cancer Center of Wake Forest University Computational Biosciences shared resource supported by NCI CCSG P30CA012197.

A Secondary Structural Transition in the C-helix Promotes Gating of Cyclic Nucleotide-regulated Ion Channels*

PubMed Central

Puljung, Michael C.; Zagotta, William N.

2013-01-01

Cyclic nucleotide-regulated ion channels bind second messengers like cAMP to a C-terminal domain, consisting of a β-roll, followed by two α-helices (B- and C-helices). We monitored the cAMP-dependent changes in the structure of the C-helix of a C-terminal fragment of HCN2 channels using transition metal ion FRET between fluorophores on the C-helix and metal ions bound between histidine pairs on the same helix. cAMP induced a change in the dimensions of the C-helix and an increase in the metal binding affinity of the histidine pair. cAMP also caused an increase in the distance between a fluorophore on the C-helix and metal ions bound to the B-helix. Stabilizing the C-helix of intact CNGA1 channels by metal binding to a pair of histidines promoted channel opening. These data suggest that ordering of the C-helix is part of the gating conformational change in cyclic nucleotide-regulated channels. PMID:23525108

Topology characterization of a benzodiazepine-binding beta-rich domain of the GABAA receptor alpha1 subunit.

PubMed

Xu, Zhiwen; Fang, Shisong; Shi, Haifeng; Li, Hoiming; Deng, Yiqun; Liao, Yinglei; Wu, Jiun-Ming; Zheng, Hui; Zhu, Huaimin; Chen, Hueih-Min; Tsang, Shui Ying; Xue, Hong

2005-10-01

Structural investigation of GABAA receptors has been limited by difficulties imposed by its trans-membrane-complex nature. In the present study, the topology of a membrane-proximal beta-rich (MPB) domain in the C139-L269 segment of the receptor alpha1 subunit was probed by mapping the benzodiazepine (BZ)-binding and epitopic sites, as well as fluorescence resonance energy transfer (FRET) analysis. Ala-scanning and semiconservative substitutions within this segment revealed the contribution of the phenyl rings of Y160 and Y210, the hydroxy group of S186 and the positive charge on R187 to BZ-binding. FRET with the bound BZ ligand indicated the proximity of Y160, S186, R187, and S206 to the BZ-binding site. On the other hand, epitope-mapping using the monoclonal antibodies (mAbs) against the MPB domain established a clustering of T172, R173, E174, Q196, and T197. Based on the lack of FRET between Trp substitutionally placed at R173 or V198 and bound BZ, this epitope-mapped cluster is located on a separate end of the folded protein from the BZ-binding site. Mutations of the five conserved Cys and Trp residues in the MPB domain gave rise to synergistic and rescuing effects on protein secondary structures and unfolding stability that point to a CCWCW-pentad, reminiscent to the CWC-triad "pin" of immunoglobulin (Ig)-like domains, important for the structural maintenance. These findings, together with secondary structure and fold predictions suggest an anti-parallel beta-strand topology with resemblance to Ig-like fold, having the BZ-binding and the epitopic residues being clustered at two different ends of the fold.

An Inserted α/β Subdomain Shapes the Catalytic Pocket of Lactobacillus johnsonii Cinnamoyl Esterase

PubMed Central

Vu, Clara; Xu, Xiaohui; Cui, Hong; Molloy, Sara; Savchenko, Alexei; Yakunin, Alexander; Gonzalez, Claudio F.

2011-01-01

Background Microbial enzymes produced in the gastrointestinal tract are primarily responsible for the release and biochemical transformation of absorbable bioactive monophenols. In the present work we described the crystal structure of LJ0536, a serine cinnamoyl esterase produced by the probiotic bacterium Lactobacillus johnsonii N6.2. Methodology/Principal Findings We crystallized LJ0536 in the apo form and in three substrate-bound complexes. The structure showed a canonical α/β fold characteristic of esterases, and the enzyme is dimeric. Two classical serine esterase motifs (GlyXSerXGly) can be recognized from the amino acid sequence, and the structure revealed that the catalytic triad of the enzyme is formed by Ser106, His225, and Asp197, while the other motif is non-functional. In all substrate-bound complexes, the aromatic acyl group of the ester compound was bound in the deepest part of the catalytic pocket. The binding pocket also contained an unoccupied area that could accommodate larger ligands. The structure revealed a prominent inserted α/β subdomain of 54 amino acids, from which multiple contacts to the aromatic acyl groups of the substrates are made. Inserts of this size are seen in other esterases, but the secondary structure topology of this subdomain of LJ0536 is unique to this enzyme and its closest homolog (Est1E) in the Protein Databank. Conclusions The binding mechanism characterized (involving the inserted α/β subdomain) clearly differentiates LJ0536 from enzymes with similar activity of a fungal origin. The structural features herein described together with the activity profile of LJ0536 suggest that this enzyme should be clustered in a new group of bacterial cinnamoyl esterases. PMID:21876742

An inserted α/β subdomain shapes the catalytic pocket of Lactobacillus johnsonii cinnamoyl esterase.

PubMed

Lai, Kin-Kwan; Stogios, Peter J; Vu, Clara; Xu, Xiaohui; Cui, Hong; Molloy, Sara; Savchenko, Alexei; Yakunin, Alexander; Gonzalez, Claudio F

2011-01-01

Microbial enzymes produced in the gastrointestinal tract are primarily responsible for the release and biochemical transformation of absorbable bioactive monophenols. In the present work we described the crystal structure of LJ0536, a serine cinnamoyl esterase produced by the probiotic bacterium Lactobacillus johnsonii N6.2. We crystallized LJ0536 in the apo form and in three substrate-bound complexes. The structure showed a canonical α/β fold characteristic of esterases, and the enzyme is dimeric. Two classical serine esterase motifs (GlyXSerXGly) can be recognized from the amino acid sequence, and the structure revealed that the catalytic triad of the enzyme is formed by Ser(106), His(225), and Asp(197), while the other motif is non-functional. In all substrate-bound complexes, the aromatic acyl group of the ester compound was bound in the deepest part of the catalytic pocket. The binding pocket also contained an unoccupied area that could accommodate larger ligands. The structure revealed a prominent inserted α/β subdomain of 54 amino acids, from which multiple contacts to the aromatic acyl groups of the substrates are made. Inserts of this size are seen in other esterases, but the secondary structure topology of this subdomain of LJ0536 is unique to this enzyme and its closest homolog (Est1E) in the Protein Databank. The binding mechanism characterized (involving the inserted α/β subdomain) clearly differentiates LJ0536 from enzymes with similar activity of a fungal origin. The structural features herein described together with the activity profile of LJ0536 suggest that this enzyme should be clustered in a new group of bacterial cinnamoyl esterases.

Multiple-probe analysis of folding and unfolding pathways of human serum albumin. Evidence for a framework mechanism of folding.

PubMed

Santra, Manas Kumar; Banerjee, Abhijit; Krishnakumar, Shyam Sundar; Rahaman, Obaidur; Panda, Dulal

2004-05-01

The changes in the far-UV CD signal, intrinsic tryptophan fluorescence and bilirubin absorbance showed that the guanidine hydrochloride (GdnHCl)-induced unfolding of a multidomain protein, human serum albumin (HSA), followed a two-state process. However, using environment sensitive Nile red fluorescence, the unfolding and folding pathways of HSA were found to follow a three-state process and an intermediate was detected in the range 0.25-1.5 m GdnHCl. The intermediate state displayed 45% higher fluorescence intensity than that of the native state. The increase in the Nile red fluorescence was found to be due to an increase in the quantum yield of the HSA-bound Nile red. Low concentrations of GdnHCl neither altered the binding affinity of Nile red to HSA nor induced the aggregation of HSA. In addition, the secondary structure of HSA was not perturbed during the first unfolding transition (<1.5 m GdnHCl); however, the secondary structure was completely lost during the second transition. The data together showed that the half maximal loss of the tertiary structure occurred at a lower GdnHCl concentration than the loss of the secondary structure. Further kinetic studies of the refolding process of HSA using multiple spectroscopic techniques showed that the folding occurred in two phases, a burst phase followed by a slow phase. An intermediate with native-like secondary structure but only a partial tertiary structure was found to form in the burst phase of refolding. Then, the intermediate slowly folded into the native state. An analysis of the refolding data suggested that the folding of HSA could be best explained by the framework model.

Structural Analysis of HMGD-DNA Complexes Reveal Influence of Intercalation on Sequence Selectivity and DNA Bending

PubMed Central

Churchill, Mair E.A.; Klass, Janet; Zoetewey, David L.

2010-01-01

The ubiquitous eukaryotic High-Mobility-Group-Box (HMGB) chromosomal proteins promote many chromatin-mediated cellular activities through their non-sequence-specific binding and bending of DNA. Minor groove DNA binding by the HMG box results in substantial DNA bending toward the major groove owing to electrostatic interactions, shape complementarity and DNA intercalation that occurs at two sites. Here, the structures of the complexes formed with DNA by a partially DNA intercalation-deficient mutant of Drosophila melanogaster HMGD have been determined by X-ray crystallography at a resolution of 2.85 Å. The six proteins and fifty base pairs of DNA in the crystal structure revealed a variety of bound conformations. All of the proteins bound in the minor groove, bridging DNA molecules, presumably because these DNA regions are easily deformed. The loss of the primary site of DNA intercalation decreased overall DNA bending and shape complementarity. However, DNA bending at the secondary site of intercalation was retained and most protein-DNA contacts were preserved. The mode of binding resembles the HMGB1-boxA-cisplatin-DNA complex, which also lacks a primary intercalating residue. This study provides new insights into the binding mechanisms used by HMG boxes to recognize varied DNA structures and sequences as well as modulate DNA structure and DNA bending. PMID:20800069

Monte Carlo modeling of single-molecule cytoplasmic dynein.

PubMed

Singh, Manoranjan P; Mallik, Roop; Gross, Steven P; Yu, Clare C

2005-08-23

Molecular motors are responsible for active transport and organization in the cell, underlying an enormous number of crucial biological processes. Dynein is more complicated in its structure and function than other motors. Recent experiments have found that, unlike other motors, dynein can take different size steps along microtubules depending on load and ATP concentration. We use Monte Carlo simulations to model the molecular motor function of cytoplasmic dynein at the single-molecule level. The theory relates dynein's enzymatic properties to its mechanical force production. Our simulations reproduce the main features of recent single-molecule experiments that found a discrete distribution of dynein step sizes, depending on load and ATP concentration. The model reproduces the large steps found experimentally under high ATP and no load by assuming that the ATP binding affinities at the secondary sites decrease as the number of ATP bound to these sites increases. Additionally, to capture the essential features of the step-size distribution at very low ATP concentration and no load, the ATP hydrolysis of the primary site must be dramatically reduced when none of the secondary sites have ATP bound to them. We make testable predictions that should guide future experiments related to dynein function.

Enhanced Shear-induced Platelet Aggregation Due to Low-temperature Storage

DTIC Science & Technology

2013-07-01

conjugated sheep polyclonal anti-VWF antibody , goat polyclonal anti-mouse FITC and PE secondary antibodies , and immunoglobulin G isotype control antibodies ...of fresh or stored PRP was incubated with 25 mg/mL FITC- conjugated AK2 or unconjugated SZ2 or VM16d antibodies for 20 minutes. In case of SZ2 and...VM16d, the primary antibody -bound PLTs were incubated with a PE- conjugated secondary antibody . The antibody - bound PLTs were then fixed in 400 mL of 1

Some Surprising Implications of NMR-directed Simulations of Substrate Recognition and Binding by Cytochrome P450cam (CYP101A1).

PubMed

Asciutto, Eliana K; Pochapsky, Thomas C

2018-04-27

Cytochrome P450 cam (CYP101A1) catalyzes the stereospecific 5-exo hydroxylation of d-camphor by molecular oxygen. Previously, residual dipolar couplings measured for backbone amide 1 H- 15 N correlations in both substrate-free and bound forms of CYP101A1 were used as restraints in soft annealing molecular dynamic simulations in order to identify average conformations of the enzyme with and without substrate bound. Multiple substrate-dependent conformational changes remote from the enzyme active site were identified, and site-directed mutagenesis and activity assays confirmed the importance of these changes in substrate recognition. The current work makes use of perturbation response scanning (PRS) and umbrella sampling molecular dynamic of the residual dipolar coupling-derived CYP101A1 structures to probe the roles of remote structural features in enforcing the regio- and stereospecific nature of the hydroxylation reaction catalyzed by CYP101A1. An improper dihedral angle Ψ was defined and used to maintain substrate orientation in the CYP101A1 active site, and it was observed that different values of Ψ result in different PRS response maps. Umbrella sampling methods show that the free energy of the system is sensitive to Ψ, and bound substrate forms an important mechanical link in the transmission of mechanical coupling through the enzyme structure. Finally, a qualitative approach to interpreting PRS maps in terms of the roles of secondary structural features is proposed. Copyright © 2018 Elsevier Ltd. All rights reserved.

Backbone and side-chain resonance assignments of (Ca2+)4-calmodulin bound to beta calcineurin A CaMBD peptide.

PubMed

Fowler, C Andrew; Núñez Hernandez, Maria F; O'Donnell, Susan E; Yu, Liping; Shea, Madeline A

2017-10-01

Calcineurin (CaN) is a heterodimeric and highly conserved serine/threonine phosphatase (PP2B) that plays a critical role in coupling calcium signals to physiological processes including embryonic cardiac development, NF-AT-regulated gene expression in immune responses, and apoptosis. The catalytic subunit (CaN A ) has three isoforms (α, β, and γ,) in humans and seven isoforms in Paramecium. In all eukaryotes, the EF-hand protein calmodulin (CaM) regulates CaN activity in a calcium-dependent manner. The N- and C-domains of CaM (CaM N and CaM C ) recognize a CaM-binding domain (CaMBD) within an intrinsically disordered region of CaN A that precedes the auto-inhibitory domain (AID) of CaN A . Here we present nearly complete 1 H, 13 C, and 15 N resonance assignments of (Ca 2+ ) 4 -CaM bound to a peptide containing the CaMBD sequence in the beta isoform of CaN A (βCaN A -CaMBDp). Its secondary structure elements predicted from the assigned chemical shifts were in good agreement with those observed in the high-resolution structures of (Ca 2+ ) 4 -CaM bound to CaMBDs of multiple enzymes. Based on the reported literature, the CaMBD of the α isoform of CaN A can bind to CaM in two opposing orientations which may influence the regulatory function of CaM. Because a high resolution structure of (Ca 2+ ) 4 -CaM bound to βCaN A -CaMBDp has not been reported, our studies serve as a starting point for determining the solution structure of this complex. This will demonstrate the preferred orientation of (Ca 2+ ) 4 -CaM on the CaMBD as well as the orientations of CaM N and CaM C relative to each other and to the AID of βCaN A .

Computational prediction and biochemical characterization of novel RNA aptamers to Rift Valley fever virus nucleocapsid protein.

PubMed

Ellenbecker, Mary; St Goddard, Jeremy; Sundet, Alec; Lanchy, Jean-Marc; Raiford, Douglas; Lodmell, J Stephen

2015-10-01

Rift Valley fever virus (RVFV) is a potent human and livestock pathogen endemic to sub-Saharan Africa and the Arabian Peninsula that has potential to spread to other parts of the world. Although there is no proven effective and safe treatment for RVFV infections, a potential therapeutic target is the virally encoded nucleocapsid protein (N). During the course of infection, N binds to viral RNA, and perturbation of this interaction can inhibit viral replication. To gain insight into how N recognizes viral RNA specifically, we designed an algorithm that uses a distance matrix and multidimensional scaling to compare the predicted secondary structures of known N-binding RNAs, or aptamers, that were isolated and characterized in previous in vitro evolution experiment. These aptamers did not exhibit overt sequence or predicted structure similarity, so we employed bioinformatic methods to propose novel aptamers based on analysis and clustering of secondary structures. We screened and scored the predicted secondary structures of novel randomly generated RNA sequences in silico and selected several of these putative N-binding RNAs whose secondary structures were similar to those of known N-binding RNAs. We found that overall the in silico generated RNA sequences bound well to N in vitro. Furthermore, introduction of these RNAs into cells prior to infection with RVFV inhibited viral replication in cell culture. This proof of concept study demonstrates how the predictive power of bioinformatics and the empirical power of biochemistry can be jointly harnessed to discover, synthesize, and test new RNA sequences that bind tightly to RVFV N protein. The approach would be easily generalizable to other applications. Copyright © 2015 Elsevier Ltd. All rights reserved.

Using Neutron Reflectometry to Discern the Structure of Fibrinogen Adsorption at the Stainless Steel/Aqueous Interface.

PubMed

Wood, Mary H; Browning, Kathryn L; Barker, Robert D; Clarke, Stuart M

2016-06-23

Neutron reflectometry has been successfully used to study adsorption on a stainless steel surface by means of depositing a thin steel film on silicon. The film was characterized using XPS (X-ray photoelectron spectroscopy), TOF-SIMS (time-of-flight secondary ion mass spectrometry), and GIXRD (grazing incidence X-ray diffraction), demonstrating the retention both of the austenitic phase and of the required composition for 316L stainless steel. The adsorption of fibrinogen from a physiologically-relevant solution onto the steel surface was studied using neutron reflectometry and QCM (quartz crystal microbalance) and compared to that on a deposited chromium oxide surface. It was found that the protein forms an irreversibly bound layer at low concentrations, with maximum protein concentration a distance of around 20 Å from the surface. Evidence for a further diffuse reversibly-bound layer forming at higher concentrations was also observed. Both the structure of the layer revealed by the neutron reflectometry data and the high water retention predicted by the QCM data suggest that there is a significant extent of protein unfolding upon adsorption. A lower extent of adsorption was seen on the chromium surfaces, although the adsorbed layer structures were similar, suggesting comparable adsorption mechanisms.

Dynamics of biomolecules, ligand binding & biological functions

NASA Astrophysics Data System (ADS)

Yi, Myunggi

Proteins are flexible and dynamic. One static structure alone does not often completely explain biological functions of the protein, and some proteins do not even have high resolution structures. In order to provide better understanding to the biological functions of nicotinic acetylcholine receptor, Diphtheria toxin repressor and M2 proton channel, the dynamics of these proteins are investigated using molecular modeling and molecular dynamics (MD) simulations. With absence of high resolution structure of alpha7 receptor, the homology models of apo and cobra toxin bound forms have been built. From the MD simulations of these model structures, we observed one subunit of apo simulation moved away from other four subunits. With local movement of flexible loop regions, the whole subunit tilted clockwise. These conformational changes occurred spontaneously, and were strongly correlated with the conformational change when the channel is activated by agonists. Unlike other computational studies, we directly compared our model of open conformation with the experimental data. However, the subunits of toxin bound form were stable, and conformational change is restricted by the bound cobra toxin. These results provide activation and inhibition mechanisms of alpha7 receptors and a possible explanation for intermediate conductance of the channel. Intramolecular complex of SH3-like domain with a proline-rich (Pr) peptide segment in Diphtheria toxin repressor (DtxR) is stabilized in inactive state. Upon activation of DtxR by transition metal binding, this intramolecular complex should be dissociated. The dynamics of this intramolecular complex is investigated using MD simulations and NMR spectroscopy. We observed spontaneous opening and closing motions of the Pr segment binding pockets in both Pr-SH3 and SH3 simulations. The MD simulation results and NMR relaxation data suggest that the Pr segment exhibits a binding ↔ unbinding equilibrium. Despite a wealth of experimental validation of Gouy-Chapman (GC) theory to charged lipid membranes, a test of GC theory by MD simulations has been elusive. Here we demonstrate that the ion distributions at different salt concentrations in anionic lipid bilayer systems agree well with GC predictions using MD simulations. Na+ ions are adsorbed to the bilayer through favorable interactions with carbonyls and hydroxyls, reducing the surface charge density by 72.5%. The interactions of amantadine, an antiinfluenza A drug, with DMPC bilayers are investigated by an MD simulation and by solid-state NMR. The MD simulation results and NMR data demonstrate that amantadine is located within the interfacial region with upward orientation and interacts with the lipid headgroup and glycerol backbone, while the adamantane group of amantadine interacts with the glycerol backbone and much of fatty acyl chain, as it wraps underneath of the drug. The lipid headgroup orientation is influenced by the drug as well. The recent prevalence of amantadine-resistant mutants makes a development of new drug urgent. The mechanism of inhibition of M2 proton channel in influenza virus A by amantadine is investigated. In the absence of high resolution structure, we model the apo and drug bound forms based on NMR structures. MD simulations demonstrate that channel pore is blocked by a primary gate formed by Trp41 helped by His37 and a secondary gate formed by Val27. The blockage by the secondary gate is extended by the drug bound just below the gate, resulting in a broken water wire throughout the simulation, suggesting a novel role of Val27 in the inhibition by amantadine. Recent X-ray structure validates the simulation results.

Molecular Dynamics of "Fuzzy" Transcriptional Activator-Coactivator Interactions

PubMed Central

Scholes, Natalie S.; Weinzierl, Robert O. J.

2016-01-01

Transcriptional activation domains (ADs) are generally thought to be intrinsically unstructured, but capable of adopting limited secondary structure upon interaction with a coactivator surface. The indeterminate nature of this interface made it hitherto difficult to study structure/function relationships of such contacts. Here we used atomistic accelerated molecular dynamics (aMD) simulations to study the conformational changes of the GCN4 AD and variants thereof, either free in solution, or bound to the GAL11 coactivator surface. We show that the AD-coactivator interactions are highly dynamic while obeying distinct rules. The data provide insights into the constant and variable aspects of orientation of ADs relative to the coactivator, changes in secondary structure and energetic contributions stabilizing the various conformers at different time points. We also demonstrate that a prediction of α-helical propensity correlates directly with the experimentally measured transactivation potential of a large set of mutagenized ADs. The link between α-helical propensity and the stimulatory activity of ADs has fundamental practical and theoretical implications concerning the recruitment of ADs to coactivators. PMID:27175900

Two-dimensional cross correlation analysis of protein unfolding: Portrayal of the thermal denaturation of CMP kinases in the absence and presence of substrates

NASA Astrophysics Data System (ADS)

Schultz, Christian P.; Bârzu, Octavian; Mantsch, Henry H.

2000-03-01

The functional role of CMP kinases is to regenerate mono-phosphate nucleotides in cells by transferring phosphate residues from tri-phosphorylated nucleotides to monophosphorylated nucleotides. These enzymes possess two binding sites and maintain a highly conserved secondary structure. They are essential for cell survival. Herein we compare the infrared spectra of two similar, but not identical enzymes, the CMP kinases from Escherichia coli and Bacillus subtilis. A two-dimensional cross correlation analysis of the infrared spectra reveals differences in the denaturation behavior of the two proteins. Different secondary structure elements show different time-delayed or advanced unfolding events in the two enzymes. When bound to the active sites, the two nucleotide-substrates CMP and ATP exert a stabilizing effect on the structure of both proteins. The changes observed upon thermal denaturation are different for the two enzymes. Model 2D correlations are used to simulate the different denaturation of the two enzymes. Thermal denaturation and aggregation can be distinguished as two processes separated in time.

On topological RNA interaction structures.

PubMed

Qin, Jing; Reidys, Christian M

2013-07-01

Recently a folding algorithm of topological RNA pseudoknot structures was presented in Reidys et al. (2011). This algorithm folds single-stranded γ-structures, that is, RNA structures composed by distinct motifs of bounded topological genus. In this article, we set the theoretical foundations for the folding of the two backbone analogues of γ structures: the RNA γ-interaction structures. These are RNA-RNA interaction structures that are constructed by a finite number of building blocks over two backbones having genus at most γ. Combinatorial properties of γ-interaction structures are of practical interest since they have direct implications for the folding of topological interaction structures. We compute the generating function of γ-interaction structures and show that it is algebraic, which implies that the numbers of interaction structures can be computed recursively. We obtain simple asymptotic formulas for 0- and 1-interaction structures. The simplest class of interaction structures are the 0-interaction structures, which represent the two backbone analogues of secondary structures.

Reducing the worst case running times of a family of RNA and CFG problems, using Valiant's approach.

PubMed

Zakov, Shay; Tsur, Dekel; Ziv-Ukelson, Michal

2011-08-18

RNA secondary structure prediction is a mainstream bioinformatic domain, and is key to computational analysis of functional RNA. In more than 30 years, much research has been devoted to defining different variants of RNA structure prediction problems, and to developing techniques for improving prediction quality. Nevertheless, most of the algorithms in this field follow a similar dynamic programming approach as that presented by Nussinov and Jacobson in the late 70's, which typically yields cubic worst case running time algorithms. Recently, some algorithmic approaches were applied to improve the complexity of these algorithms, motivated by new discoveries in the RNA domain and by the need to efficiently analyze the increasing amount of accumulated genome-wide data. We study Valiant's classical algorithm for Context Free Grammar recognition in sub-cubic time, and extract features that are common to problems on which Valiant's approach can be applied. Based on this, we describe several problem templates, and formulate generic algorithms that use Valiant's technique and can be applied to all problems which abide by these templates, including many problems within the world of RNA Secondary Structures and Context Free Grammars. The algorithms presented in this paper improve the theoretical asymptotic worst case running time bounds for a large family of important problems. It is also possible that the suggested techniques could be applied to yield a practical speedup for these problems. For some of the problems (such as computing the RNA partition function and base-pair binding probabilities), the presented techniques are the only ones which are currently known for reducing the asymptotic running time bounds of the standard algorithms.

Reducing the worst case running times of a family of RNA and CFG problems, using Valiant's approach

PubMed Central

2011-01-01

Background RNA secondary structure prediction is a mainstream bioinformatic domain, and is key to computational analysis of functional RNA. In more than 30 years, much research has been devoted to defining different variants of RNA structure prediction problems, and to developing techniques for improving prediction quality. Nevertheless, most of the algorithms in this field follow a similar dynamic programming approach as that presented by Nussinov and Jacobson in the late 70's, which typically yields cubic worst case running time algorithms. Recently, some algorithmic approaches were applied to improve the complexity of these algorithms, motivated by new discoveries in the RNA domain and by the need to efficiently analyze the increasing amount of accumulated genome-wide data. Results We study Valiant's classical algorithm for Context Free Grammar recognition in sub-cubic time, and extract features that are common to problems on which Valiant's approach can be applied. Based on this, we describe several problem templates, and formulate generic algorithms that use Valiant's technique and can be applied to all problems which abide by these templates, including many problems within the world of RNA Secondary Structures and Context Free Grammars. Conclusions The algorithms presented in this paper improve the theoretical asymptotic worst case running time bounds for a large family of important problems. It is also possible that the suggested techniques could be applied to yield a practical speedup for these problems. For some of the problems (such as computing the RNA partition function and base-pair binding probabilities), the presented techniques are the only ones which are currently known for reducing the asymptotic running time bounds of the standard algorithms. PMID:21851589

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.

Intrinsic disorder in the partitioning protein KorB persists after co-operative complex formation with operator DNA and KorA.

PubMed

Hyde, Eva I; Callow, Philip; Rajasekar, Karthik V; Timmins, Peter; Patel, Trushar R; Siligardi, Giuliano; Hussain, Rohanah; White, Scott A; Thomas, Christopher M; Scott, David J

2017-08-30

The ParB protein, KorB, from the RK2 plasmid is required for DNA partitioning and transcriptional repression. It acts co-operatively with other proteins, including the repressor KorA. Like many multifunctional proteins, KorB contains regions of intrinsically disordered structure, existing in a large ensemble of interconverting conformations. Using NMR spectroscopy, circular dichroism and small-angle neutron scattering, we studied KorB selectively within its binary complexes with KorA and DNA, and within the ternary KorA/KorB/DNA complex. The bound KorB protein remains disordered with a mobile C-terminal domain and no changes in the secondary structure, but increases in the radius of gyration on complex formation. Comparison of wild-type KorB with an N-terminal deletion mutant allows a model of the ensemble average distances between the domains when bound to DNA. We propose that the positive co-operativity between KorB, KorA and DNA results from conformational restriction of KorB on binding each partner, while maintaining disorder. © 2017 The Author(s).

Thermal and Structural Analysis of a Hollow Core Space Shuttle Main Engine (SSME) Turbine Blade

NASA Technical Reports Server (NTRS)

Abdul-Aziz, Ali; Kalluri, Sreeramesh; McGaw, Michael A.

1995-01-01

The influence of primary and secondary orientations on the elastic response of a hollow core, (001)-oriented nickel base single-crystal superalloy turbine blade, was investigated under combined thermal and mechanical conditions. Finite element techniques is employed through MARC finite element code to conduct the analyses on a hollow core SSME turbine blade made out of PWA 1480 single crystal material. Primary orientation of the single crystal superalloy was varied in increments of 2 deg, from 0 to 10 deg, from the (001) direction. Two secondary orientations (0 and 45 deg) were considered with respect to the global coordinate system, as the primary orientation angle was varied. The stresses developed within the single crystal blade were determined for different orientations of the blade. The influence of angular offsets such as the single crystal's primary and secondary orientations and the loading conditions on the elastic stress response of the PWA 1480 hollow blade are summarized. The influence of he primary orientation angle, when constrained between the bounds considered, was not found to be as significant as the influence of the secondary orientation angle.

Folding-unfolding transitions of Rv3221c on the pressure-temperature plane

NASA Astrophysics Data System (ADS)

Somkuti, Judit; Jain, Sriyans; Ramachandran, Srinivasan; ászló Smeller, L.

2013-06-01

Rv3221c is a biotin-binding protein found in Mycobacterium tuberculosis. It has been reported that an elevated temperature is needed for it to adopt a folded conformation. We determined the complete pressure-temperature phase diagram, and determined the thermodynamical parameters of the denaturation. The phase diagram follows well the Hawley theory. The secondary structure of the protein was found to contain predominantly beta sheet. The pressure unfolding was partially reversible, resulting in pressure-sensitive aggregates, besides the correctly refolded and biotin-bound fraction of proteins.

Protein Secondary Structures (alpha-helix and beta-sheet) at a Cellular Levle and Protein Fractions in Relation to Rumen Degradation Behaviours of Protein: A New Approach

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

Yu,P.

2007-01-01

Studying the secondary structure of proteins leads to an understanding of the components that make up a whole protein, and such an understanding of the structure of the whole protein is often vital to understanding its digestive behaviour and nutritive value in animals. The main protein secondary structures are the {alpha}-helix and {beta}-sheet. The percentage of these two structures in protein secondary structures influences protein nutritive value, quality and digestive behaviour. A high percentage of {beta}-sheet structure may partly cause a low access to gastrointestinal digestive enzymes, which results in a low protein value. The objectives of the present studymore » were to use advanced synchrotron-based Fourier transform IR (S-FTIR) microspectroscopy as a new approach to reveal the molecular chemistry of the protein secondary structures of feed tissues affected by heat-processing within intact tissue at a cellular level, and to quantify protein secondary structures using multicomponent peak modelling Gaussian and Lorentzian methods, in relation to protein digestive behaviours and nutritive value in the rumen, which was determined using the Cornell Net Carbohydrate Protein System. The synchrotron-based molecular chemistry research experiment was performed at the National Synchrotron Light Source at Brookhaven National Laboratory, US Department of Energy. The results showed that, with S-FTIR microspectroscopy, the molecular chemistry, ultrastructural chemical make-up and nutritive characteristics could be revealed at a high ultraspatial resolution ({approx}10 {mu}m). S-FTIR microspectroscopy revealed that the secondary structure of protein differed between raw and roasted golden flaxseeds in terms of the percentages and ratio of {alpha}-helixes and {beta}-sheets in the mid-IR range at the cellular level. By using multicomponent peak modelling, the results show that the roasting reduced (P <0.05) the percentage of {alpha}-helixes (from 47.1% to 36.1%: S-FTIR absorption intensity), increased the percentage of {beta}-sheets (from 37.2% to 49.8%: S-FTIR absorption intensity) and reduced the {alpha}-helix to {beta}-sheet ratio (from 0.3 to 0.7) in the golden flaxseeds, which indicated a negative effect of the roasting on protein values, utilisation and bioavailability. These results were proved by the Cornell Net Carbohydrate Protein System in situ animal trial, which also revealed that roasting increased the amount of protein bound to lignin, and well as of the Maillard reaction protein (both of which are poorly used by ruminants), and increased the level of indigestible and undegradable protein in ruminants. The present results demonstrate the potential of highly spatially resolved synchrotron-based infrared microspectroscopy to locate 'pure' protein in feed tissues, and reveal protein secondary structures and digestive behaviour, making a significant step forward in and an important contribution to protein nutritional research. Further study is needed to determine the sensitivities of protein secondary structures to various heat-processing conditions, and to quantify the relationship between protein secondary structures and the nutrient availability and digestive behaviour of various protein sources. Information from the present study arising from the synchrotron-based IR probing of the protein secondary structures of protein sources at the cellular level will be valuable as a guide to maintaining protein quality and predicting digestive behaviours.« less

High-resolution structures of adenylate kinase from yeast ligated with inhibitor Ap5A, showing the pathway of phosphoryl transfer.

PubMed Central

Abele, U.; Schulz, G. E.

1995-01-01

The structure of adenylate kinase from yeast ligated with the two-substrate-mimicking inhibitor Ap5A and Mg2+ has been refined to 1.96 A resolution. In addition, the refined structure of the same complex with a bound imidazole molecule replacing Mg2+ has been determined at 1.63 A. These structures indicate that replacing Mg2+ by imidazole disturbs the water structure and thus the complex. A comparison with the G-proteins shows that Mg2+ is exactly at the same position with respect to the phosphates. However, although the Mg2+ ligand sphere of the G-proteins is a regular octahedron containing peptide ligands, the reported adenylate kinase has no such ligands and an open octahedron leaving space for the Mg2+ to accompany the transferred phosphoryl group. A superposition of the known crystalline and therefore perturbed phosphoryl transfer geometries in the adenylate kinases demonstrates that all of them are close to the start of the forward reaction with bound ATP and AMP. Averaging all observed perturbed structures gives rise to a close approximation of the transition state, indicating in general how to establish an elusive transition state geometry. The average shows that the in-line phosphoryl transfer is associative, because there is no space for a dissociative metaphosphate intermediate. As a side result, the secondary dipole interaction in the alpha-helices of both protein structures has been quantified. PMID:7670369

Exploration of electrostatic interaction in the hydrophobic pocket of lysozyme: Importance of ligand-induced perturbation of the secondary structure on the mode of binding of exogenous ligand and possible consequences.

PubMed

Panja, Sudipta; Halder, Mintu

2016-08-01

Exogenous ligand binding can be adequate to alter the secondary structure of biomolecules besides other external stimuli. In such cases, structural alterations can complicate on the nature of interaction with the exogenous molecules. In order to accommodate the exogenous ligand, the biomolecule has to unfold resulting in a considerable change to its properties. If the bound ligand can be unbound, the biomolecule gets the opportunity to refold back and return to its native state. Keeping this in mind, we have purposely investigated the interaction of tartrazine (TZ), a well abundant azo food colorant, with two homologous lysozymes, namely, human lysozyme (HLZ) and chicken egg white lysozyme (CEWLZ) in physiological pH condition. The binding of TZ with lysozymes has been identified to accompany a ligand-induced secondary structure alteration as indicated by the circular dichroism spectra, and the reduction of α-helical content is more with HLZ than CEWLZ. Interestingly, the binding is identified to occur in the electronic ground state of TZ with lysozyme in its hydrophobic cavity, containing excess of positive charge, predominantly via electrostatic interaction. With increase of salinity of the medium the protein tends to refold back due to wakening of electrostatic forces and consequent reduction of strength of ligand interaction and unbinding. The entropy enthalpy compensation (EEC) has been probed to understand the binding features and it is found that CEWLZ-TZ shows better compensation than HLZ-TZ complex. This is presumably due to the fact that with CEWLZ the binding does not accompany substantial change in the protein secondary structure and hence ineffective to scramble the EEC. The present study initiates the importance of ligand-perturbed structural alteration of biomolecule in controlling the thermodynamics of binding. If there is a considerable alteration of the protein secondary structure due to binding, it is indicative that such changes should bring in the overall loss of activity of protein. Copyright © 2016 Elsevier B.V. All rights reserved.

34 CFR 645.11 - What services do all Upward Bound projects provide?

Code of Federal Regulations, 2010 CFR

2010-07-01

... programs involving elementary or secondary school teachers, faculty members at institutions of higher....11 Section 645.11 Education Regulations of the Offices of the Department of Education (Continued) OFFICE OF POSTSECONDARY EDUCATION, DEPARTMENT OF EDUCATION UPWARD BOUND PROGRAM What Kinds of Projects...

Sensational Studies in Marine Science.

ERIC Educational Resources Information Center

Keller, E. C., Jr.; Schroyer, Fred C.

1981-01-01

Presents a description of a five-week summer course in marine biology and oceanography offered to college-bound, secondary students with varied physical handicaps. Summarizes insights gained after four summer sessions related to communication problems, physical arrangements for the wheelchair-bound, and handicap-proof maps; evaluates the course's…

Deciphering complex dynamics of water counteraction around secondary structural elements of allosteric protein complex: Case study of SAP-SLAM system in signal transduction cascade

NASA Astrophysics Data System (ADS)

Samanta, Sudipta; Mukherjee, Sanchita

2018-01-01

The first hydration shell of a protein exhibits heterogeneous behavior owing to several attributes, majorly local polarity and structural flexibility as revealed by solvation dynamics of secondary structural elements. We attempt to recognize the change in complex water counteraction generated due to substantial alteration in flexibility during protein complex formation. The investigation is carried out with the signaling lymphocytic activation molecule (SLAM) family of receptors, expressed by an array of immune cells, and interacting with SLAM-associated protein (SAP), composed of one SH2 domain. All atom molecular dynamics simulations are employed to the aqueous solutions of free SAP and SLAM-peptide bound SAP. We observed that water dynamics around different secondary structural elements became highly affected as well as nicely correlated with the SLAM-peptide induced change in structural rigidity obtained by thermodynamic quantification. A few instances of contradictory dynamic features of water to the change in structural flexibility are explained by means of occluded polar residues by the peptide. For βD, EFloop, and BGloop, both structural flexibility and solvent accessibility of the residues confirm the obvious contribution. Most importantly, we have quantified enhanced restriction in water dynamics around the second Fyn-binding site of the SAP due to SAP-SLAM complexation, even prior to the presence of Fyn. This observation leads to a novel argument that SLAM induced more restricted water molecules could offer more water entropic contribution during the subsequent Fyn binding and provide enhanced stability to the SAP-Fyn complex in the signaling cascade. Finally, SLAM induced water counteraction around the second binding site of the SAP sheds light on the allosteric property of the SAP, which becomes an integral part of the underlying signal transduction mechanism.

Deciphering complex dynamics of water counteraction around secondary structural elements of allosteric protein complex: Case study of SAP-SLAM system in signal transduction cascade.

PubMed

Samanta, Sudipta; Mukherjee, Sanchita

2018-01-28

The first hydration shell of a protein exhibits heterogeneous behavior owing to several attributes, majorly local polarity and structural flexibility as revealed by solvation dynamics of secondary structural elements. We attempt to recognize the change in complex water counteraction generated due to substantial alteration in flexibility during protein complex formation. The investigation is carried out with the signaling lymphocytic activation molecule (SLAM) family of receptors, expressed by an array of immune cells, and interacting with SLAM-associated protein (SAP), composed of one SH2 domain. All atom molecular dynamics simulations are employed to the aqueous solutions of free SAP and SLAM-peptide bound SAP. We observed that water dynamics around different secondary structural elements became highly affected as well as nicely correlated with the SLAM-peptide induced change in structural rigidity obtained by thermodynamic quantification. A few instances of contradictory dynamic features of water to the change in structural flexibility are explained by means of occluded polar residues by the peptide. For βD, EFloop, and BGloop, both structural flexibility and solvent accessibility of the residues confirm the obvious contribution. Most importantly, we have quantified enhanced restriction in water dynamics around the second Fyn-binding site of the SAP due to SAP-SLAM complexation, even prior to the presence of Fyn. This observation leads to a novel argument that SLAM induced more restricted water molecules could offer more water entropic contribution during the subsequent Fyn binding and provide enhanced stability to the SAP-Fyn complex in the signaling cascade. Finally, SLAM induced water counteraction around the second binding site of the SAP sheds light on the allosteric property of the SAP, which becomes an integral part of the underlying signal transduction mechanism.

Mentoring College Bound High School Seniors.

ERIC Educational Resources Information Center

Mowrer-Popiel, Elizabeth

This article examines causes of the high rate of attrition of college freshmen during the first few weeks of school and describes a plan for mentorships between successful college students and college-bound secondary seniors prior to entrance into college. In discussing the challenges facing freshmen, the article suggests that they suffer stress…

College Bound Program; Summer 1975.

ERIC Educational Resources Information Center

Woloshin, Gerald W.

The principal objectives of the College Bound Summer Program, funded under the Elementary Secondary Education Act Title I, were to improve student's reading and mathematics, increase their ability to do college work, and make the students' transition from junior high to high school easier. Program participants were selected on the basis of either…

Analysis of nucleotides and oligonucleotides immobilized as self-assembled monolayers by static secondary ion mass spectrometry.

PubMed

Patrick, J S; Cooks, R G; Pachuta, S J

1994-11-01

Nucleic acid constituents can be bound to a metal surface in the form of self-assembled monolayers. Binding is achieved either through ionic interactions with a self-assembled 2-aminoethanethiol monolayer or by direct covalent binding of a dithiophosphate oligonucleotide to a metal surface through a sulfur-metal bond. Nucleotides, polynucleotides (both normal and a dithiophosphate analog) and double-stranded DNA have all been bound to surfaces. When the surfaces are interrogated using static secondary ion mass spectrometry (SIMS), the surface-bound nucleic acid constituents are observed in the form of the characteristic protonated nucleic acid base ions (BH2+). While a silver foil substrate was found to provide the highest absolute signal, vapor-deposited gold yields the best signal-to-noise ratio for ionically bound deoxyguanosine monophosphate. Under comparable conditions, a Cs+ projectile produces a 10-fold increase in the secondary ion signal relative to a Ga+ projectile. The experiment has been extended to a triple-quadrupole instrument where tandem mass spectrometric experiments on ionically immobilized dGMP showed the characteristic loss of ammonia from the released BH2+ ion. When a 'biomimetic' surface formed by ionically immobilizing double-stranded DNA is exposed to a solution containing ethidium bromide, ions corresponding to the non-covalent adduct are readily detectable using SIMS. This adduct and the nucleic acid constituents can be monitored at levels below 10 fmol.

NMR structural and dynamical investigation of the isolated voltage-sensing domain of the potassium channel KvAP: implications for voltage gating.

PubMed

Shenkarev, Zakhar O; Paramonov, Alexander S; Lyukmanova, Ekaterina N; Shingarova, Lyudmila N; Yakimov, Sergei A; Dubinnyi, Maxim A; Chupin, Vladimir V; Kirpichnikov, Mikhail P; Blommers, Marcel J J; Arseniev, Alexander S

2010-04-28

The structure and dynamics of the isolated voltage-sensing domain (VSD) of the archaeal potassium channel KvAP was studied by high-resolution NMR. The almost complete backbone resonance assignment and partial side-chain assignment of the (2)H,(13)C,(15)N-labeled VSD were obtained for the protein domain solubilized in DPC/LDAO (2:1) mixed micelles. Secondary and tertiary structures of the VSD were characterized using secondary chemical shifts and NOE contacts. These data indicate that the spatial structure of the VSD solubilized in micelles corresponds to the structure of the domain in an open state of the channel. NOE contacts and secondary chemical shifts of amide protons indicate the presence of tightly bound water molecule as well as hydrogen bond formation involving an interhelical salt bridge (Asp62-R133) that stabilizes the overall structure of the domain. The backbone dynamics of the VSD was studied using (15)N relaxation measurements. The loop regions S1-S2 and S2-S3 were found mobile, while the S3-S4 loop (voltage-sensor paddle) was found stable at the ps-ns time scale. The moieties of S1, S2, S3, and S4 helices sharing interhelical contacts (at the level of the Asp62-R133 salt bridge) were observed in conformational exchange on the micros-ms time scale. Similar exchange-induced broadening of characteristic resonances was observed for the VSD solubilized in the membrane of lipid-protein nanodiscs composed of DMPC, DMPG, and POPC/DOPG lipids. Apparently, the observed interhelical motions represent an inherent property of the VSD of the KvAP channel and can play an important role in the voltage gating.

Summer Upward Bound, Terre Haute, Indiana. Secondary Program in Compensatory Education, 4.

ERIC Educational Resources Information Center

American Institutes for Research in the Behavioral Sciences, Palo Alto, CA.

Upward Bound was a precollege program geared for high school students with potential who had been handicapped by economic, cultural, and educational deprivation. It involved a full-time summer program and follow-up programs (counseling, cultural activities, and physical education) during the academic year. Students stayed in the program for three…

34 CFR 645.11 - What services do all Upward Bound projects provide?

Code of Federal Regulations, 2011 CFR

2011-07-01

..., study skills, mathematics, science, and other subjects; (2) Advice and assistance in secondary and... 34 Education 3 2011-07-01 2011-07-01 false What services do all Upward Bound projects provide? 645.11 Section 645.11 Education Regulations of the Offices of the Department of Education (Continued...

Structural stability of myoglobin and glycomyoglobin: a comparative molecular dynamics simulation study.

PubMed

Alizadeh-Rahrovi, Joulia; Shayesteh, Alireza; Ebrahim-Habibi, Azadeh

2015-09-01

Glycoproteins are formed as the result of enzymatic glycosylation or chemical glycation in the body, and produced in vitro in industrial processes. The covalently attached carbohydrate molecule(s) confer new properties to the protein, including modified stability. In the present study, the structural stability of a glycoprotein form of myoglobin, bearing a glucose unit in the N-terminus, has been compared with its native form by the use of molecular dynamics simulation. Both structures were subjected to temperatures of 300 and 500 K in an aqueous environment for 10 ns. Changes in secondary structures and RMSD were then assessed. An overall higher stability was detected for glycomyoglobin, for which the most stable segments/residues were highlighted and compared with the native form. The simple addition of a covalently bound glucose is suggested to exert its stabilizing effect via increased contacts with surrounding water molecules, as well as a different pattern of interactions with neighbor residues.

Structures of holo wild-type human cellular retinol-binding protein II (hCRBPII) bound to retinol and retinal

PubMed Central

Nossoni, Zahra; Assar, Zahra; Yapici, Ipek; Nosrati, Meisam; Wang, Wenjing; Berbasova, Tetyana; Vasileiou, Chrysoula; Borhan, Babak; Geiger, James

2014-01-01

Cellular retinol-binding proteins (CRBPs) I and II, which are members of the intracellular lipid-binding protein (iLBP) family, are retinoid chaperones that are responsible for the intracellular transport and delivery of both retinol and retinal. Although structures of retinol-bound CRBPI and CRBPII are known, no structure of a retinal-bound CRBP has been reported. In addition, the retinol-bound human CRBPII (hCRBPII) structure shows partial occupancy of a non­canonical conformation of retinol in the binding pocket. Here, the structure of retinal-bound hCRBPII and the structure of retinol-bound hCRBPII with retinol fully occupying the binding pocket are reported. It is further shown that the retinoid derivative seen in both the zebrafish CRBP and the hCRBPII structures is likely to be the product of flux-dependent and wavelength-dependent X-ray damage during data collection. The structures of retinoid-bound CRBPs are compared and contrasted, and rationales for the differences in binding affinities for retinal and retinol are provided. PMID:25478840

Structures of holo wild-type human cellular retinol-binding protein II (hCRBPII) bound to retinol and retinal.

PubMed

Nossoni, Zahra; Assar, Zahra; Yapici, Ipek; Nosrati, Meisam; Wang, Wenjing; Berbasova, Tetyana; Vasileiou, Chrysoula; Borhan, Babak; Geiger, James

2014-12-01

Cellular retinol-binding proteins (CRBPs) I and II, which are members of the intracellular lipid-binding protein (iLBP) family, are retinoid chaperones that are responsible for the intracellular transport and delivery of both retinol and retinal. Although structures of retinol-bound CRBPI and CRBPII are known, no structure of a retinal-bound CRBP has been reported. In addition, the retinol-bound human CRBPII (hCRBPII) structure shows partial occupancy of a noncanonical conformation of retinol in the binding pocket. Here, the structure of retinal-bound hCRBPII and the structure of retinol-bound hCRBPII with retinol fully occupying the binding pocket are reported. It is further shown that the retinoid derivative seen in both the zebrafish CRBP and the hCRBPII structures is likely to be the product of flux-dependent and wavelength-dependent X-ray damage during data collection. The structures of retinoid-bound CRBPs are compared and contrasted, and rationales for the differences in binding affinities for retinal and retinol are provided.

Programmed folding of DNA origami structures through single-molecule force control.

PubMed

Bae, Wooli; Kim, Kipom; Min, Duyoung; Ryu, Je-Kyung; Hyeon, Changbong; Yoon, Tae-Young

2014-12-03

Despite the recent development in the design of DNA origami, its folding yet relies on thermal or chemical annealing methods. We here demonstrate mechanical folding of the DNA origami structure via a pathway that has not been accessible to thermal annealing. Using magnetic tweezers, we stretch a single scaffold DNA with mechanical tension to remove its secondary structures, followed by base pairing of the stretched DNA with staple strands. When the force is subsequently quenched, folding of the DNA nanostructure is completed through displacement between the bound staple strands. Each process in the mechanical folding is well defined and free from kinetic traps, enabling us to complete folding within 10 min. We also demonstrate parallel folding of DNA nanostructures through multiplexed manipulation of the scaffold DNAs. Our results suggest a path towards programmability of the folding pathway of DNA nanostructures.

The effect of wall geometry in particle-laden turbulent flow

NASA Astrophysics Data System (ADS)

Abdehkakha, Hoora; Iaccarino, Gianluca

2016-11-01

Particle-laden turbulent flow plays a significant role in various industrial applications, as turbulence alters the exchange of momentum and energy between particles and fluid flow. In wall-bounded flows, inhomogeneity in turbulent properties is the primary cause of turbophoresis that leads the particles toward the walls. Conversely, shear-induced lift force on the particles can become important if large scale vortical structures are present. The objective of this study is to understand the effects of geometry on fluid flows and consequently on particles transport and concentration. Direct numerical simulations combined with point particle Lagrangian tracking are performed for several geometries such as a pipe, channel, square duct, and squircle (rounded-corners duct). In non-circular ducts, anisotropic and inhomogeneous Reynolds stresses are the most influential phenomena that produce the secondary flows. It has been shown that these motions can have a significant impact on transporting momentum, vorticity, and energy from the core of the duct to the corners. The main focus of the present study is to explore the effects of near the wall structures and secondary flows on turbophoresis, lift, and particle concentration.

Thermal Design and Analysis of the Optical Telescope Assembly for the Gondola for High Altitude Planetary Science

NASA Technical Reports Server (NTRS)

O'Connor, Brian; Brooks, Thomas

2017-01-01

The NASA Gondola for High Altitude Planetary Science (GHAPS) project is an effort to design, build, and fly a balloon-borne platform for planetary science missions. GHAPS observations will be in the 300 nm to 5 micron wavelength region covering UV, visible, and near-mid IR. The primary element of the project is the Optical Telescope Assembly (OTA). It is a one meter aperture narrow-field-of-view telescope that contains the primary and secondary mirrors, the support system/metering structure, a secondary mirror focusing system, baffles, and insulation. This paper presents the thermal design and analysis that has been done to support the design of the OTA. A major part of the thermal analysis was bounding the flight environment for the six potential Columbia Scientific Balloon Facility launch sites. These analyses were used to give input into the Structural Thermal Optical Performance (STOP) analysis of the telescope. Also the analysis was used to select heater sizes for the few OTA associated electronic components. Currently the telescope is scheduled to have its first flight in 2019.

Multiscale Modelling for investigating single molecule effects on the mechanics of actin filaments

NASA Astrophysics Data System (ADS)

A, Deriu Marco; C, Bidone Tamara; Laura, Carbone; Cristina, Bignardi; M, Montevecchi Franco; Umberto, Morbiducci

2011-12-01

This work presents a preliminary multiscale computational investigation of the effects of nucleotides and cations on the mechanics of actin filaments (F-actin). At the molecular level, Molecular Dynamics (MD) simulations are employed to characterize the rearrangements of the actin monomers (G-actin) in terms of secondary structures evolution in physiological conditions. At the mesoscale level, a coarse grain (CG) procedure is adopted where each monomer is represented by means of Elastic Network Modeling (ENM) technique. At the macroscale level, actin filaments up to hundreds of nanometers are assumed as isotropic and elastic beams and characterized via Rotation Translation Block (RTB) analysis. F-actin bound to adenosine triphosphate (ATP) shows a persistence length around 5 μm, while actin filaments bound to adenosine diphosphate (ADP) have a persistence length of about 3 μm. With magnesium bound to the high affinity binding site of G-actin, the persistence length of F-actin decreases to about 2 μm only in the ADP-bound form of the filament, while the same ion has no effects, in terms of stiffness variation, on the ATP-bound form of F-actin. The molecular mechanisms behind these changes in flexibility are herein elucidated. Thus, this study allows to analyze how the local binding of cations and nucleotides on G-actin induce molecular rearrangements that transmit to the overall F-actin, characterizing shifts of mechanical properties, that can be related with physiological and pathological cellular phenomena, as cell migration and spreading. Further, this study provides the basis for upcoming investigating of network and cellular remodelling at higher length scales.

Covalent Bonding of Chlorogenic Acid Induces Structural Modifications on Sunflower Proteins.

PubMed

Karefyllakis, Dimitris; Salakou, Stavroula; Bitter, J Harry; van der Goot, Atze J; Nikiforidis, Constantinos V

2018-02-19

Proteins and phenols coexist in the confined space of plant cells leading to reactions between them, which result in new covalently bonded complex molecules. This kind of reactions has been widely observed during storage and processing of plant materials. However, the nature of the new complex molecules and their physicochemical properties are largely unknown. Therefore, we investigated the structural characteristics of covalently bonded complexes between sunflower protein isolate (SFPI, protein content 85 wt %) and the dominant phenol in the confined space of a sunflower seed cell (chlorogenic acid, CGA). It was shown that the efficiency of bond formation goes through a maximum as a function of the SFPI:CGA ratio. Moreover, the bonding of CGA with proteins resulted in changes in the secondary and tertiary structure of the protein. It was also shown that the phenol bound strongly to the protein, which resulted in new crosslinks between the polypeptide chains. As a result, secondary structures like α-helices and β-sheets diminished, which in turn resulted in more disordered domains and a subsequent modification of the tertiary structure of the proteins. These findings are relevant for establishing future protocols for extraction of high-quality proteins and phenols when utilizing plant material and offer insight into the impact of processing that these ingredients endure. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Intrinsic flexibility of West Nile virus protease in solution characterized using small-angle X-ray scattering.

PubMed

Garces, Andrea P; Watowich, Stanley J

2013-10-01

West Nile virus (WNV) is a mosquito-borne flavivirus with a rapidly expanding global distribution. Infection can cause severe neurological disease and fatality in humans. Efforts are ongoing to develop antiviral drugs that inhibit the WNV protease, a viral enzyme required for polyprotein processing. Unfortunately, little is known about the solution structure of recombinant WNV protease (NS2B-NS3pro) used for antiviral drug discovery and development, although X-ray crystal structures and nuclear magnetic resonance (NMR) studies have provided valuable insights into the interactions between NS2B-NS3pro and peptide-based inhibitors. We completed small-angle X-ray scattering and Fourier transform infrared spectroscopy experiments to determine the solution structure and dynamics of WNV NS2B-NS3pro in the absence of a bound substrate or inhibitor. Importantly, these solution studies suggested that all or most of the NS2B cofactor was highly flexible and formed an ensemble of structures, in contrast to the NS2B tertiary structures observed in crystallographic and NMR studies. The secondary structure of NS2B-NS3pro in solution had high β-content, similar to the secondary structure observed in crystallographic studies. This work provided evidence of the intrinsic flexibility and conformational heterogeneity of the NS2B chain of the WNV protease in the absence of substratelike ligands, which should be considered during antiviral drug discovery and development efforts.

C-terminal, endoplasmic reticulum-lumenal domain of prosurfactant protein C - structural features and membrane interactions.

PubMed

Casals, Cristina; Johansson, Hanna; Saenz, Alejandra; Gustafsson, Magnus; Alfonso, Carlos; Nordling, Kerstin; Johansson, Jan

2008-02-01

Surfactant protein C (SP-C) constitutes the transmembrane part of prosurfactant protein C (proSP-C) and is alpha-helical in its native state. The C-terminal part of proSP-C (CTC) is localized in the endoplasmic reticulum lumen and binds to misfolded (beta-strand) SP-C, thereby preventing its aggregation and amyloid fibril formation. In this study, we investigated the structure of recombinant human CTC and the effects of CTC-membrane interaction on protein structure. CTC forms noncovalent trimers and supratrimeric oligomers. It contains two intrachain disulfide bridges, and its secondary structure is significantly affected by urea or heat only after disulfide reduction. The postulated Brichos domain of CTC, with homologs found in proteins associated with amyloid and proliferative disease, is up to 1000-fold more protected from limited proteolysis than the rest of CTC. The protein exposes hydrophobic surfaces, as determined by CTC binding to the environment-sensitive fluorescent probe 1,1'-bis(4-anilino-5,5'-naphthalenesulfonate). Fluorescence energy transfer experiments further reveal close proximity between bound 1,1'-bis(4-anilino-5,5'-naphthalenesulfonate) and tyrosine residues in CTC, some of which are conserved in all Brichos domains. CTC binds to unilamellar phospholipid vesicles with low micromolar dissociation constants, and differential scanning calorimetry and CD analyses indicate that membrane-bound CTC is less structurally ordered than the unbound protein. The exposed hydrophobic surfaces and the structural disordering that result from interactions with phospholipid membranes suggest a mechanism whereby CTC binds to misfolded SP-C in the endoplasmic reticulum membrane.

ATP interacts with the CPVT mutation-associated central domain of the cardiac ryanodine receptor.

PubMed

Blayney, Lynda; Beck, Konrad; MacDonald, Ewan; D'Cruz, Leon; Nomikos, Michail; Griffiths, Julia; Thanassoulas, Angelos; Nounesis, George; Lai, F Anthony

2013-10-01

This study was designed to determine whether the cardiac ryanodine receptor (RyR2) central domain, a region associated with catecholamine polymorphic ventricular tachycardia (CPVT) mutations, interacts with the RyR2 regulators, ATP and the FK506-binding protein 12.6 (FKBP12.6). Wild-type (WT) RyR2 central domain constructs (G(2236)to G(2491)) and those containing the CPVT mutations P2328S and N2386I, were expressed as recombinant proteins. Folding and stability of the proteins were examined by circular dichroism (CD) spectroscopy and guanidine hydrochloride chemical denaturation. The far-UV CD spectra showed a soluble stably-folded protein with WT and mutant proteins exhibiting a similar secondary structure. Chemical denaturation analysis also confirmed a stable protein for both WT and mutant constructs with similar two-state unfolding. ATP and caffeine binding was measured by fluorescence spectroscopy. Both ATP and caffeine bound with an EC50 of ~200-400μM, and the affinity was the same for WT and mutant constructs. Sequence alignment with other ATP binding proteins indicated the RyR2 central domain contains the signature of an ATP binding pocket. Interaction of the central domain with FKBP12.6 was tested by glutaraldehyde cross-linking and no association was found. The RyR2 central domain, expressed as a 'correctly' folded recombinant protein, bound ATP in accord with bioinformatics evidence of conserved ATP binding sequence motifs. An interaction with FKBP12.6 was not evident. CPVT mutations did not disrupt the secondary structure nor binding to ATP. Part of the RyR2 central domain CPVT mutation cluster, can be expressed independently with retention of ATP binding. Copyright © 2013 Elsevier B.V. All rights reserved.

Specific Amyloid Binding of Polybasic Peptides In Vivo Is Retained by β-Sheet Conformers but Lost in the Disrupted Coil and All D-Amino Acid Variants.

PubMed

Wall, Jonathan S; Williams, Angela; Richey, Tina; Stuckey, Alan; Wooliver, Craig; Christopher Scott, J; Donnell, Robert; Martin, Emily B; Kennel, Stephen J

2017-10-01

The heparin-reactive, helical peptide p5 is an effective amyloid imaging agent in mice with systemic amyloidosis. Analogs of p5 with modified secondary structure characteristics exhibited altered binding to heparin, synthetic amyloid fibrils, and amyloid extracts in vitro. Herein, we further study the effects of peptide helicity and chirality on specific amyloid binding using a mouse model of systemic inflammation-associated (AA) amyloidosis. Peptides with disrupted helical structure [p5 (coil) and p5 (Pro3) ], with an extended sheet conformation [p5 (sheet) ] or an all-D enantiomer [p5 (D) ], were chemically synthesized, radioiodinated, and their biodistribution studied in WT mice as well as transgenic animals with severe systemic AA amyloidosis. Peptide binding was assessed qualitatively by using small animal single-photon emission computed tomography/x-ray computed tomography imaging and microautoradiography and quantitatively using tissue counting. Peptides with reduced helical propensity, p5 (coil) and p5 (Pro3) , exhibited significantly reduced binding to AA amyloid-laden organs. In contrast, peptide p5 (D) was retained by non-amyloid-related ligands in the liver and kidneys of both WT and AA mice, but it also bound AA amyloid in the spleen. The p5 (sheet) peptide specifically bound AA amyloid in vivo and was not retained by healthy tissues in WT animals. Modification of amyloid-targeting peptides using D-amino acids should be performed cautiously due to the introduction of unexpected secondary pharmacologic effects. Peptides that adopt a helical structure, to align charged amino acid side chains along one face, exhibit specific reactivity with amyloid; however, polybasic peptides with a propensity for β-sheet conformation are also amyloid-reactive and may yield a novel class of amyloid-targeting agents for imaging and therapy.

Revealing the membrane-bound structure of neurokinin A using neutron diffraction

NASA Astrophysics Data System (ADS)

Darkes, Malcolm J. M.; Hauss, Thomas; Dante, Silvia; Bradshaw, Jeremy P.

2000-03-01

Neurokinin A (or substance K) belongs to the tachykinin family, a group of small amphipathic peptides that bind to specific membrane-embedded, G-protein coupled receptors. The agonist/receptor complex is quaternary in nature because the receptor binding sites are thought to be located within the lipid bilayer and because the role of water cannot be ignored. The cell membrane acts as a solvent to accumulate peptide and an inducer of peptide secondary structure. The three-dimensional shape that the peptide assumes when associated to the cell membrane will be an important parameter with regards to the receptor selectivity and affinity. Neutron diffraction measurements were carried out in order to define the location of the N-terminus of the peptide in synthetic phospholipid multi-bilayer stacks.

Structural basis of molecular recognition of helical histone H3 tail by PHD finger domains.

PubMed

Bortoluzzi, Alessio; Amato, Anastasia; Lucas, Xavier; Blank, Manuel; Ciulli, Alessio

2017-05-04

The plant homeodomain (PHD) fingers are among the largest family of epigenetic domains, first characterized as readers of methylated H3K4. Readout of histone post-translational modifications by PHDs has been the subject of intense investigation; however, less is known about the recognition of secondary structure features within the histone tail itself. We solved the crystal structure of the PHD finger of the bromodomain adjacent to zinc finger 2A [BAZ2A, also known as TIP5 (TTF-I/interacting protein 5)] in complex with unmodified N-terminal histone H3 tail. The peptide is bound in a helical folded-back conformation after K4, induced by an acidic patch on the protein surface that prevents peptide binding in an extended conformation. Structural bioinformatics analyses identify a conserved Asp/Glu residue that we name 'acidic wall', found to be mutually exclusive with the conserved Trp for K4Me recognition. Neutralization or inversion of the charges at the acidic wall patch in BAZ2A, and homologous BAZ2B, weakened H3 binding. We identify simple mutations on H3 that strikingly enhance or reduce binding, as a result of their stabilization or destabilization of H3 helicity. Our work unravels the structural basis for binding of the helical H3 tail by PHD fingers and suggests that molecular recognition of secondary structure motifs within histone tails could represent an additional layer of regulation in epigenetic processes. © 2017 The Author(s).

Indexing and retrieving point and region objects

NASA Astrophysics Data System (ADS)

Ibrahim, Azzam T.; Fotouhi, Farshad A.

1996-03-01

R-tree and its variants are examples of spatial data structures for paged-secondary memory. To process a query, these structures require multiple path traversals. In this paper, we present a new image access method, SB+-tree which requires a single path traversal to process a query. Also, SB+-tree will allow commercial databases an access method for spatial objects without a major change, since most commercial databases already support B+-tree as an access method for text data. The SB+-tree can be used for zero and non-zero size data objects. Non-zero size objects are approximated by their minimum bounding rectangles (MBRs). The number of SB+-trees generated is dependent upon the number of dimensions of the approximation of the object. The structure supports efficient spatial operations such as regions-overlap, distance and direction. In this paper, we experimentally and analytically demonstrate the superiority of SB+-tree over R-tree.

Disorder and function: a review of the dehydrin protein family

PubMed Central

Graether, Steffen P.; Boddington, Kelly F.

2014-01-01

Dehydration proteins (dehydrins) are group 2 members of the late embryogenesis abundant (LEA) protein family. The protein architecture of dehydrins can be described by the presence of three types of conserved sequence motifs that have been named the K-, Y-, and S-segments. By definition, a dehydrin must contain at least one copy of the lysine-rich K-segment. Abiotic stresses such as drought, cold, and salinity cause the upregulation of dehydrin mRNA and protein levels. Despite the large body of genetic and protein evidence of the importance of these proteins in stress response, the in vivo protective mechanism is not fully known. In vitro experimental evidence from biochemical assays and localization experiments suggests multiple roles for dehydrins, including membrane protection, cryoprotection of enzymes, and protection from reactive oxygen species. Membrane binding by dehydrins is likely to be as a peripheral membrane protein, since the protein sequences are highly hydrophilic and contain many charged amino acids. Because of this, dehydrins in solution are intrinsically disordered proteins, that is, they have no well-defined secondary or tertiary structure. Despite their disorder, dehydrins have been shown to gain structure when bound to ligands such as membranes, and to possibly change their oligomeric state when bound to ions. We review what is currently known about dehydrin sequences and their structures, and examine the various ligands that have been shown to bind to this family of proteins. PMID:25400646

34 CFR 645.6 - What definitions apply to the Upward Bound Program?

Code of Federal Regulations, 2012 CFR

2012-07-01

..., chemistry, and physics. (iv) Three years of social studies. (v) One year of a language other than English... by the individual's State. Rigorous secondary school program of study means a program of study that... recognized as a rigorous secondary school program of study by the Secretary through the process described in...

34 CFR 645.6 - What definitions apply to the Upward Bound Program?

Code of Federal Regulations, 2014 CFR

2014-07-01

..., chemistry, and physics. (iv) Three years of social studies. (v) One year of a language other than English... by the individual's State. Rigorous secondary school program of study means a program of study that... recognized as a rigorous secondary school program of study by the Secretary through the process described in...

34 CFR 645.6 - What definitions apply to the Upward Bound Program?

Code of Federal Regulations, 2013 CFR

2013-07-01

..., chemistry, and physics. (iv) Three years of social studies. (v) One year of a language other than English... by the individual's State. Rigorous secondary school program of study means a program of study that... recognized as a rigorous secondary school program of study by the Secretary through the process described in...

34 CFR 645.6 - What definitions apply to the Upward Bound Program?

Code of Federal Regulations, 2011 CFR

2011-07-01

..., chemistry, and physics. (iv) Three years of social studies. (v) One year of a language other than English... by the individual's State. Rigorous secondary school program of study means a program of study that... recognized as a rigorous secondary school program of study by the Secretary through the process described in...

Redesigning Qatar's Post-Secondary Scholarship System. Research Brief

ERIC Educational Resources Information Center

Augustine, Catherine H.; Krop, Cathy

2008-01-01

At the request of Qatar's Supreme Education Council (SEC), RAND researchers devised a new set of scholarship programs for college-bound Qatari students and proposed the formation of an institute to manage the programs and oversee other post-secondary functions. The suggested system balanced support for local institutions of higher learning with…

Levels of a terpenoid glycoside (blumenin) and cell wall-bound phenolics in some cereal mycorrhizas.

PubMed Central

Maier, W; Peipp, H; Schmidt, J; Wray, V; Strack, D

1995-01-01

Four cereals, Hordeum vulgare (barley), Triticum aestivum (wheat), Secale cereal (rye), and Avena sativa (oat), were grown in a defined nutritional medium with and without the arbuscular mycorrhizal fungus Glomus intraradices. Levels of soluble and cell wall-bound secondary metabolites in the roots of mycorrhizal and nonmycorrhizal plants were determined by high-performance liquid chromatography during the first 6 to 8 weeks of plant development. Whereas there was no difference in the levels of the cell wall-bound hydroxycinnamic acids, 4-coumaric and ferulic acids, there was a fungus-induced change of the soluble secondary root metabolites. The most obvious effect observed in all four cereals was the induced accumulation of a terpenoid glycoside. This compound was isolated and identified by spectroscopic methods (nuclear magnetic resonance, mass spectrometry) to be a cyclohexenone derivative, i.e. blumenol C 9-O-(2'-O-beta-glucuronosyl)-beta-glucoside. The level of this compound was found to be directly correlated with the degree of root colonization. PMID:7480342

Phosphorylation regulates the secondary structure and function of dentin phosphoprotein peptides

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

Villarreal-Ramirez, Eduardo; Eliezer, David; Garduño-Juarez, Ramon

Dentin phosphoprotein (DPP) is the most acidic protein in vertebrates and structurally is classified as an intrinsically disordered protein. Functionally, DPP is related to dentin and bone formation, however the specifics of such association remain unknown. Here, we used atomistic molecular dynamics simulations to screen selected binding domains of DPP onto hydroxyapatite (HA), which is one of its important interacting partners. From these results, we selected a functionally relevant peptide, Ace-SSDSSDSSDSSDSSD-NH2 (named P5) and its phosphorylated form (named P5P), for experimental characterization. SAXS experiments indicated that in solution P5 was disordered, possibly in an extended conformation while P5P displayed moremore » compact globular conformations. Circular dichroism and FTIR confirmed that, either in the presence or absence of Ca2 +/HA, P5 adopts a random coil structure, whereas its phosphorylated counterpart, P5P, has a more compact arrangement associated with conformations that display β-sheet and α-helix motifs when bound to HA. In solution, P5 inhibited HA crystal growth, whereas at similar concentrations, P5P stimulated it. These findings suggest that phosphorylation controls the transient formation of secondary and tertiary structure of DPP peptides, and, most likely of DPP itself, which in turn controls HA growth in solution and possibly HA growth in mineralized tissues.« less

Reconstituted High-Density Lipoprotein Containing Human Growth Hormone-1 Shows Potent Tissue Regeneration Activity with Enhancement of Anti-Oxidant and Anti-Atherosclerotic Activities.

PubMed

Lee, Eun-Young; Kim, So-Hee; Cho, Kyung-Hyun

2015-06-01

Human growth hormone-1 (GH-1), somatotropin, is a peptide hormone that stimulates cell division in tissues such as bone and cartilage. To compare physiological activities in lipid-free and lipid-bound states, we expressed and incorporated GH-1 in reconstituted high-density lipoprotein (rHDL). GH-1 was expressed and purified using the pET30(a) vector and an Escherichia coli expression system. Purified GH-1 (at least 98% purity, 23 kD) was characterized and synthesized with apolipoproteinA-I (apoA-I), 1-palmitoyl-2-oleoyl-phosphatidylcholine (POPC), and cholesterol. Secondary structure analysis of GH-1 revealed 54% α-helical content in a lipid-free state and 65% α-helical content in a lipid-bound state along with blue-shifted tryptophan movement (around 2 nm). GH-1 caused less uptake of oxidized low-density lipoprotein (oxLDL) into macrophages and inhibited senescence of dermal cells in a dose-dependent manner. GH-1 in a lipid-bound state exerted stronger inhibitory activity than in a lipid-free state, indicating improved anti-atherosclerotic activity due to the lipid formulation. In a fin regeneration experiment using zebrafish (17 weeks old, n=9), GH-1 showed its highest regeneration speed without any side effects. GH-1-rHDL containing apoA-I showed 2.3-fold and 1.6-fold higher regeneration speeds than lipid-free GH-1 (in native state) and lipid-bound GH-1, respectively. Incorporation of GH-1 and apoA-I in HDL enhanced tissue regeneration activity of amputated tail fin, indicating a synergetic effect between GH-1 and apoA-I in a lipid-bound state.

The structure of a turbulent flow in a channel of complex shape

USGS Publications Warehouse

Tracy, Hubert Jerome

1976-01-01

Measurements of the Reynolds stresses and the mean motion pattern were made in a uniform turbulent motion in a conduit consisting of a large, nearly square section joined by a smaller rectangular section. The results indicate that the boundary shearing stress is nearly constant over large segments of the boundaries. The magnitudes of the lateral and the vertical components of turbulence are not the same near a boundary and the component normal to the boundary is smaller than the component parallel to the boundary. The difference in the two components in the corner regions of the channel produces secondary mean motions in the plane of the channel section. The strength of the motion depends upon the angle subtended by the corner. A principal function of the secondary motions is to transfer momentum into the corner regions and, elsewhere, to compensate for the excess force due to the shear gradients. In the absence of the secondary motions, the fluid must stagnate and separate from the boundaries in certain regions and be greatly accelerated in others. The secondary motions are conventionally described in terms of symmetrical rotations in cells bounded by the corner bisectors. The measured motion pattern is at variance with this view, unless the symmetry is confined to a very local region. (Woodard-USGS)

Analysis of the tertiary structure of the ribonuclease P ribozyme-substrate complex by site-specific photoaffinity crosslinking.

PubMed Central

Harris, M E; Kazantsev, A V; Chen, J L; Pace, N R

1997-01-01

Bacterial ribonuclease P (RNase P), an endonuclease involved in tRNA maturation, is a ribonucleoprotein containing a catalytic RNA. The secondary structure of this ribozyme is well-established, and a low-resolution model of the three-dimensional structure of the ribozyme-substrate complex has been proposed based on site-specific crosslinking and phylogenetic comparative data [Harris ME et al., 1994 EMBO J 13:3953-3963]. However, several substructures of that model were poorly constrained by the available data. In the present analysis, additional constraints between elements within the Escherichia coli RNase P RNA-pre-tRNA complex were determined by intra- and intermolecular crosslinking experiments. Circularly permuted RNase P RNAs were used to position an azidophenacyl photoactive crosslinking agent specifically at strategic sites within the ribozyme-substrate complex. Crosslink sites were mapped by primer extension and confirmed by analysis of the mobility of the crosslinked RNA lariats on denaturing acrylamide gels relative to circular and linear RNA standards. Crosslinked species generally retained significant catalytic activity, indicating that the results reflect the native ribozyme structure. The crosslinking results support the general configuration of the structure model and predicate new positions and orientations for helices that were previously poorly constrained by the data set. The expanded library of crosslinking constraints was used, together with secondary and tertiary structure identified by phylogenetic sequence comparisons, to refine significantly the model of RNase P RNA with bound substrate pre-tRNA. The crosslinking results and data from chemical-modification and mutational studies are discussed in the context of the current structural perspective on this ribozyme. PMID:9174092

Allosteric effects of gold nanoparticles on human serum albumin.

PubMed

Shao, Qing; Hall, Carol K

2017-01-07

The ability of nanoparticles to alter protein structure and dynamics plays an important role in their medical and biological applications. We investigate allosteric effects of gold nanoparticles on human serum albumin protein using molecular simulations. The extent to which bound nanoparticles influence the structure and dynamics of residues distant from the binding site is analyzed. The root mean square deviation, root mean square fluctuation and variation in the secondary structure of individual residues on a human serum albumin protein are calculated for four protein-gold nanoparticle binding complexes. The complexes are identified in a brute-force search process using an implicit-solvent coarse-grained model for proteins and nanoparticles. They are then converted to atomic resolution and their structural and dynamic properties are investigated using explicit-solvent atomistic molecular dynamics simulations. The results show that even though the albumin protein remains in a folded structure, the presence of a gold nanoparticle can cause more than 50% of the residues to decrease their flexibility significantly, and approximately 10% of the residues to change their secondary structure. These affected residues are distributed on the whole protein, even on regions that are distant from the nanoparticle. We analyze the changes in structure and flexibility of amino acid residues on a variety of binding sites on albumin and confirm that nanoparticles could allosterically affect the ability of albumin to bind fatty acids, thyroxin and metals. Our simulations suggest that allosteric effects must be considered when designing and deploying nanoparticles in medical and biological applications that depend on protein-nanoparticle interactions.

Structure and function of small heat shock/alpha-crystallin proteins: established concepts and emerging ideas.

PubMed

MacRae, T H

2000-06-01

Small heat shock/alpha-crystallin proteins are defined by conserved sequence of approximately 90 amino acid residues, termed the alpha-crystallin domain, which is bounded by variable amino- and carboxy-terminal extensions. These proteins form oligomers, most of uncertain quaternary structure, and oligomerization is prerequisite to their function as molecular chaperones. Sequence modelling and physical analyses show that the secondary structure of small heat shock/alpha-crystallin proteins is predominately beta-pleated sheet. Crystallography, site-directed spin-labelling and yeast two-hybrid selection demonstrate regions of secondary structure within the alpha-crystallin domain that interact during oligomer assembly, a process also dependent on the amino terminus. Oligomers are dynamic, exhibiting subunit exchange and organizational plasticity, perhaps leading to functional diversity. Exposure of hydrophobic residues by structural modification facilitates chaperoning where denaturing proteins in the molten globule state associate with oligomers. The flexible carboxy-terminal extension contributes to chaperone activity by enhancing the solubility of small heat shock/alpha-crystallin proteins. Site-directed mutagenesis has yielded proteins where the effect of the change on structure and function depends upon the residue modified, the organism under study and the analytical techniques used. Most revealing, substitution of a conserved arginine residue within the alpha-crystallin domain has a major impact on quaternary structure and chaperone action probably through realignment of beta-sheets. These mutations are linked to inherited diseases. Oligomer size is regulated by a stress-responsive cascade including MAPKAP kinase 2/3 and p38. Phosphorylation of small heat shock/alpha-crystallin proteins has important consequences within stressed cells, especially for microfilaments.

Nature and morphology of fumed oxides and features of interfacial phenomena

NASA Astrophysics Data System (ADS)

Gun'ko, V. M.; Zarko, V. I.; Goncharuk, O. V.; Matkovsky, A. K.; Remez, O. S.; Skubiszewska-Zięba, J.; Wojcik, G.; Walusiak, B.; Blitz, J. P.

2016-03-01

Individual and complex fumed nanooxides were studied using high-resolution transmission electron microscopy, X-ray diffraction, ultraviolet-visible (UV-vis) spectroscopy, differential scanning calorimetry, nuclear magnetic resonance spectroscopy, adsorption, desorption (evaporation), and quantum chemical methods. For mixed nanooxides in contrast to simple and small nanoparticles of individual silica or titania, complex core-shell nanoparticles (50-200 nm in size) with titania or alumina cores and silica or alumina shells can be destroyed under high-pressure cryogelation (HPCG), mechnochemical activation (MCA) that also affect the structure of aggregates of nanoparticles and agglomerates of aggregates becoming more compacted. This is accompanied by changes in color from white to beige of different tints and changes in the UV-vis spectra in the 300-600 nm range, as well as changes in crystalline structure of alumina. Any treatment of 'soft' nanooxides affects the interfacial behavior of polar and nonpolar adsorbates. For some of them, the hysteresis loops become strongly open. Rearrangement of secondary particles affects the freezing-melting point depression. Clusterization of adsorbates bound in pores causes diminution of heat effects during phase transition (freezing, fusion). Freezing point depression and increasing melting point cause significant hysteresis freezing-melting effects for adsorbates bound to oxide nanoparticles. The study shows that complex nanooxides can be more sensitive to external actions than simple nanooxides such as silica.

A Different Approach to Teaching Social Studies: Folk Songs History

ERIC Educational Resources Information Center

Tangülü, Zafer

2014-01-01

The purpose of this study is to determine the effect of teaching and learning the subjects of Social Studies with folk songs in secondary school students. This study is made in 2012-2013 Academic Year Spring Term with seventh grade students studying in secondary school bounded Mugla Provincial Directorate for National Education. 67 students have…

The structure of lactoferrin-binding protein B from Neisseria meningitidis suggests roles in iron acquisition and neutralization of host defences

PubMed Central

Brooks, Cory L.; Arutyunova, Elena; Lemieux, M. Joanne

2014-01-01

Pathogens have evolved a range of mechanisms to acquire iron from the host during infection. Several Gram-negative pathogens including members of the genera Neisseria and Moraxella have evolved two-component systems that can extract iron from the host glycoproteins lactoferrin and transferrin. The homologous iron-transport systems consist of a membrane-bound transporter and an accessory lipoprotein. While the mechanism behind iron acquisition from transferrin is well understood, relatively little is known regarding how iron is extracted from lactoferrin. Here, the crystal structure of the N-terminal domain (N-lobe) of the accessory lipoprotein lactoferrin-binding protein B (LbpB) from the pathogen Neisseria meningitidis is reported. The structure is highly homologous to the previously determined structures of the accessory lipoprotein transferrin-binding protein B (TbpB) and LbpB from the bovine pathogen Moraxella bovis. Docking the LbpB structure with lactoferrin reveals extensive binding interactions with the N1 subdomain of lactoferrin. The nature of the interaction precludes apolactoferrin from binding LbpB, ensuring the specificity of iron-loaded lactoferrin. The specificity of LbpB safeguards proper delivery of iron-bound lactoferrin to the transporter lactoferrin-binding protein A (LbpA). The structure also reveals a possible secondary role for LbpB in protecting the bacteria from host defences. Following proteolytic digestion of lactoferrin, a cationic peptide derived from the N-terminus is released. This peptide, called lactoferricin, exhibits potent antimicrobial effects. The docked model of LbpB with lactoferrin reveals that LbpB interacts extensively with the N-terminal lactoferricin region. This may provide a venue for preventing the production of the peptide by proteolysis, or directly sequestering the peptide, protecting the bacteria from the toxic effects of lactoferricin. PMID:25286931

Electron emission from surfaces resulting from low energy positron bombardment

NASA Astrophysics Data System (ADS)

Mukherjee, Saurabh

Measurements of the secondary electron energy spectra resulting from very low energy positron bombardment of a polycrystalline Au and Cu (100) surfaces are presented that provide evidence for a single step transition from an unbound scattering state to an image potential bound state. The primary positron energy threshold for secondary electron emission and energy cutoff of the positron induced secondary electron energy peak are consistent with an Auger like process in which an incident positron make a transition from a scattering state to a surface-image potential bound while transferring all of the energy difference to an outgoing secondary electron. We term this process: the Auger mediated quantum sticking effect (AQSE). The intensities of the positron induced secondary electron peak are used to estimate the probability of this process as a function of incident positron energy. Positron annihilation induced Auger spectra (PAES) of Cu and Au are presented that are free of all primary beam induced secondary electron background. This background was eliminated by setting the positron beam energy below AQSE threshold. The background free PAES spectra obtained include the first measurements of the low energy tail of CVV Auger transitions all the way down to zero kinetic energy. The integrated intensity of this tail is several times larger than Auger peak itself which provides strong evidence for multi-electron Auger processes.

Folding dynamics of linear emulsion polymers into 3D architectures

NASA Astrophysics Data System (ADS)

McMullen, Angus; Bargteil, Dylan; Brujic, Jasna

Colloidal polymers have been limited to inflexible, solid colloids. Here we show that the fluidity of emulsion droplets allows for the self-assembly of flexible droplet chains, which can subsequently be folded into 3D structures via secondary interactions. We achieve this using DNA-guided interactions, to initially form the chain, and then program its folding pathways. When two emulsion droplets labeled with complementary DNA meet, the balance of hybridization energy and droplet deformation yields an equilibrium patch size. Therefore, the concentration of DNA on the surface determines the number of droplet-droplet bonds in the assembly. We find that 96 % of bound droplets successfully self-assemble into chains. Droplet binding is a stochastic process, following a Poisson distribution of lengths. Since the fluid droplets can rearrange, we compare the dynamics of emulsion chains to that of polymers. We also trigger secondary interactions along the chain, causing the formation of specific loops or compact clusters. This approach will allow us to fold our emulsion polymers into a wide array of soft structures, giving us a powerful biomimetic colloidal system to investigate protein folding on the mesoscopic scale. This work was supported by the NSF MRSEC Program (DMR-0820341).

Porous medium convection at large Rayleigh number: Studies of coherent structure, transport, and reduced dynamics

NASA Astrophysics Data System (ADS)

Wen, Baole

Buoyancy-driven convection in fluid-saturated porous media is a key environmental and technological process, with applications ranging from carbon dioxide storage in terrestrial aquifers to the design of compact heat exchangers. Porous medium convection is also a paradigm for forced-dissipative infinite-dimensional dynamical systems, exhibiting spatiotemporally chaotic dynamics if not "true" turbulence. The objective of this dissertation research is to quantitatively characterize the dynamics and heat transport in two-dimensional horizontal and inclined porous medium convection between isothermal plane parallel boundaries at asymptotically large values of the Rayleigh number Ra by investigating the emergent, quasi-coherent flow. This investigation employs a complement of direct numerical simulations (DNS), secondary stability and dynamical systems theory, and variational analysis. The DNS confirm the remarkable tendency for the interior flow to self-organize into closely-spaced columnar plumes at sufficiently large Ra (up to Ra ≃ 105), with more complex spatiotemporal features being confined to boundary layers near the heated and cooled walls. The relatively simple form of the interior flow motivates investigation of unstable steady and time-periodic convective states at large Ra as a function of the domain aspect ratio L. To gain insight into the development of spatiotemporally chaotic convection, the (secondary) stability of these fully nonlinear states to small-amplitude disturbances is investigated using a spatial Floquet analysis. The results indicate that there exist two distinct modes of instability at large Ra: a bulk instability mode and a wall instability mode. The former usually is excited by long-wavelength disturbances and is generally much weaker than the latter. DNS, strategically initialized to investigate the fully nonlinear evolution of the most dangerous secondary instability modes, suggest that the (long time) mean inter-plume spacing in statistically-steady porous medium convection results from an interplay between the competing effects of these two types of instability. Upper bound analysis is then employed to investigate the dependence of the heat transport enhancement factor, i.e. the Nusselt number Nu, on Ra and L. To solve the optimization problems arising from the "background field" upper-bound variational analysis, a novel two-step algorithm in which time is introduced into the formulation is developed. The new algorithm obviates the need for numerical continuation, thereby enabling the best available bounds to be computed up to Ra ≈ 2.65 x 104. A mathematical proof is given to demonstrate that the only steady state to which this numerical algorithm can converge is the required global optimal of the variational problem. Using this algorithm, the dependence of the bounds on L( Ra) is explored, and a "minimal flow unit" is identified. Finally, the upper bound variational methodology is also shown to yield quantitatively useful predictions of Nu and to furnish a functional basis that is naturally adapted to the boundary layer dynamics at large Ra..

Locating Bound Structures in the Accelerating Universe

NASA Astrophysics Data System (ADS)

Pearson, David; Batuski, D. J.

2013-01-01

Given the overwhelming evidence of the universe’s accelerating expansion, the question of what structures are gravitationally bound becomes one of utmost interest. Dunner et al. 2006 (D06) and Busha et al. 2003 (B03) set out to answer this question analytically, and they arrived at fairly different answers owing to the differences in their assumptions of velocities at the present epoch. Applying their criteria to different superclusters, it’s possible to make predictions about what structures may be bound. We apply the criteria of D06 and B03 to the Aquarius, Microscopium, Corona Borealis, and Shapley superclusters to make predictions about what structures might be bound and compare with the results of simple N-body simulations to determine which method is a better predictor and to determine the likelihood that parts or all of the superclusters listed above are bound. We find that D06 tend to predict more structure to be bound than B03, and the results of the N-body simulations usually lie somewhere in between the two sets of predictions. Observational evidence, and simulation data suggests that pairs of clusters in Aquarius and Microscopium are gravitationally bound, and that Shapley contains a large complex of clusters that are bound, along with some additional bound pairs. The likelihood that any of the clusters in Corona Borealis are bound to one another is very small, contrary to the claims of Small et al. 1998, who claimed that the entire supercluster is likely gravitationally bound. Busha M. T., Adams F. C., Wechsler R. H., Evrard A. E., 2003, ApJ, 596, 713 Dunner R., Araya P. A., Meza A., Reisenegger A., 2006, MNRAS, 306, 803 Small T. A., Ma C., Sargent W. L. W., Hamilton D., 1998, ApJ, 492, 45

Delipidation of Cytochrome c Oxidase from Rhodobacter sphaeroides Destabilizes its Quaternary Structure

PubMed Central

Musatov, Andrej; Varhač, Rastislav; Hosler, Jonathan P.; Sedlák, Erik

2016-01-01

Delipidation of detergent-solubilized cytochrome c oxidase isolated from Rhodobacter sphaeroides (Rbs-CcO) has no apparent structural and/or functional effect on the protein, however affects its resistance against thermal or chemical denaturation. Phospholipase A2 (PLA2) hydrolysis of phospholipids that are co-purified with the enzyme removes all but two tightly bound phosphatidylethanolamines. Replacement of the removed phospholipids with nonionic detergent decreases both thermal stability of the enzyme and its resilience against the effect of chemical denaturants such as urea. In contrast to nondelipidated Rbs-CcO, the enzymatic activity of PLA2-treated Rbs-CcO is substantially diminished after exposure to high (>4M) urea concentration at room temperature without an alteration of its secondary structure. Absorbance spectroscopy and sedimentation velocity experiments revealed a strong correlation between intact tertiary structure of heme regions and quaternary structure, respectively, and the enzymatic activity of the protein. We concluded that phospholipid environment of Rbs-CcO has the protective role for stability of its tertiary and quaternary structures. PMID:26923069

Immunogenicity of Membrane-bound HIV-1 gp41 Membrane-proximal External Region (MPER) Segments Is Dominated by Residue Accessibility and Modulated by Stereochemistry*

PubMed Central

Kim, Mikyung; Song, Likai; Moon, James; Sun, Zhen-Yu J.; Bershteyn, Anna; Hanson, Melissa; Cain, Derek; Goka, Selasie; Kelsoe, Garnett; Wagner, Gerhard; Irvine, Darrell; Reinherz, Ellis L.

2013-01-01

Structural characterization of epitope-paratope pairs has contributed to the understanding of antigenicity. By contrast, few structural studies relate to immunogenicity, the process of antigen-induced immune responses in vivo. Using a lipid-arrayed membrane-proximal external region (MPER) of HIV-1 glycoprotein 41 as a model antigen, we investigated the influence of physicochemical properties on immunogenicity in relation to structural modifications of MPER/liposome vaccines. Anchoring the MPER to the membrane via an alkyl tail or transmembrane domain retained the MPER on liposomes in vivo, while preserving MPER secondary structure. However, structural modifications that affected MPER membrane orientation and antigenic residue accessibility strongly impacted induced antibody responses. The solvent-exposed MPER tryptophan residue (Trp-680) was immunodominant, focusing immune responses, despite sequence variability elsewhere. Nonetheless, immunogenicity could be readily manipulated using site-directed mutagenesis or structural constraints to modulate amino acid surface display. These studies provide fundamental insights for immunogen design aimed at targeting B cell antibody responses. PMID:24047898

Disneyization: A framework for understanding illicit drug use in bounded play spaces.

PubMed

Turner, Tim

2018-05-17

This paper combines evidence from an ethnographic study of illicit drug use amongst tourists in Ibiza with Bryman's (2004) theoretical model of Disneyization. The principal aim was to construct a new conceptual framework that may help scholars, practitioners and policy makers make sense of dynamic patterns of illegal drug use across bounded play spaces such as tourist resorts, music festivals and nightclubs. Ethnographic fieldwork employing a grounded theory design was undertaken over three summers in tourist locations on the Balearic island of Ibiza, including nightclubs, bars, cafes, beaches, airports and hotels. Field notes from participant observation were supplemented with data from semi-structured interviews (n = 56) and secondary sources gathered from tourist marketing. The framework of Disneyization has been discussed in terms of 5 constructs: theming, hybrid consumption, branding, performative labour and atmospheres; each having a specific role in relation to understanding illicit drug use in bounded play spaces. Thus: Theming sets the stage, by physically and symbolically demarcating space with indelible themes of hedonism that open up the possibility of illicit drug use. Hybrid-consumption blurs the distinction between legal and illegal forms of intoxication, making the trading and consumption of illegal drugs appear like a natural feature of the consumer space. Branding demonstrates how participants construct intricate hierarchies of taste and credibility related to drug of choice. Performative labour re-enforces hybrid consumption, with participants working in the bounded play spaces of Ibiza immersed within the illicit drug market. Atmospheres represents the alchemic synergy of bounded play space and is important to understanding illicit drug use as a sensorial, deeply immersive but transitory experience. This research offers Disneyization as a new conceptual framework for making sense of deeply complex spatial, socio-cultural, psychological and economic processes that underpin dynamic patterns of drug use in bounded play spaces. Copyright © 2018 Elsevier B.V. All rights reserved.

Influence of selected physicochemical parameters on microbiological activity of mucks.

NASA Astrophysics Data System (ADS)

Całka, A.; Sokołowska, Z.; Warchulska, P.; Dąbek-Szreniawska, M.

2009-04-01

One of the basic factor decided about soil fertility are microorganisms that together with flora, determine trend and character of biochemical processes as well totality of fundamental transformations connected with biogeochemistry and physicochemical properties of soil. Determination of general bacteria number, quantity of selected groups of microorganisms and investigation of respiration intensity let estimate microbiological activity of soil. Intensity of microbiological processes is directly connected with physicochemical soil parameters. In that case, such structural parameters as bulk density, porosity, surface or carbon content play significant role. Microbiological activity also changes within the bounds of mucks with different stage of humification and secondary transformation. Knowledge of relations between structural properties, microorganism activity and degree of transformation and humification can lead to better understanding microbiological processes as well enable to estimate microbiological activity at given physicochemical conditions and at progressing process of soil transformation. The study was carried out on two peaty-moorsh (muck) soils at different state of secondary transformation and humification degree. Soil samples were collected from Polesie Lubelskie (layer depth: 5 - 25 cm). Investigated mucks originated from soils formed from low peatbogs. Soil sample marked as I belonged to muck group weakly secondary transformed. Second sample (II) represented soil group with middle stage of secondary transformation. The main purpose of the research was to examine the relations between some physicochemical and surface properties and their biological activity. Total number and respiration activity of microorganisms were determined. The effectiveness of utilizing the carbon substances from the soil by the bacteria increased simultaneously with the transformation state of the peat-muck soils. Quantity of organic carbon decreased distinctly in the soil at the higher stage of secondary transformation and it influenced quantity and activity of soil microorganisms. Bulk density and surface increased with increasing secondary transformation degree. On the other hand, porosity decreased with increasing secondary transformation index. Process of secondary transformation influenced the soil environment for the microbes by changing the physicochemical properties. This way it influenced the number of microorganisms and caused changes of biological activity in the soils.

Structural modeling of the N-terminal signal–receiving domain of IκBα

PubMed Central

Yazdi, Samira; Durdagi, Serdar; Naumann, Michael; Stein, Matthias

2015-01-01

The transcription factor nuclear factor-κB (NF-κB) exerts essential roles in many biological processes including cell growth, apoptosis and innate and adaptive immunity. The NF-κB inhibitor (IκBα) retains NF-κB in the cytoplasm and thus inhibits nuclear localization of NF-κB and its association with DNA. Recent protein crystal structures of the C-terminal part of IκBα in complex with NF-κB provided insights into the protein-protein interactions but could not reveal structural details about the N-terminal signal receiving domain (SRD). The SRD of IκBα contains a degron, formed following phosphorylation by IκB kinases (IKK). In current protein X-ray structures, however, the SRD is not resolved and assumed to be disordered. Here, we combined secondary structure annotation and domain threading followed by long molecular dynamics (MD) simulations and showed that the SRD possesses well-defined secondary structure elements. We show that the SRD contains 3 additional stable α-helices supplementing the six ARDs present in crystallized IκBα. The IκBα/NF-κB protein-protein complex remained intact and stable during the entire simulations. Also in solution, free IκBα retains its structural integrity. Differences in structural topology and dynamics were observed by comparing the structures of NF-κB free and NF-κB bound IκBα-complex. This study paves the way for investigating the signaling properties of the SRD in the IκBα degron. A detailed atomic scale understanding of molecular mechanism of NF-κB activation, regulation and the protein-protein interactions may assist to design and develop novel chronic inflammation modulators. PMID:26157801

Backbone conformational preferences of an intrinsically disordered protein in solution.

PubMed

Espinoza-Fonseca, L Michel; Ilizaliturri-Flores, Ian; Correa-Basurto, José

2012-06-01

We have performed a 4-μs molecular dynamics simulation to investigate the native conformational preferences of the intrinsically disordered kinase-inducible domain (KID) of the transcription factor CREB in solution. There is solid experimental evidence showing that KID does not possess a bound-like structure in solution; however, it has been proposed that coil-to-helix transitions upon binding to its binding partner (CBP) are template-driven. While these studies indicate that IDPs possess a bias towards the bound structure, they do not provide direct evidence on the time-dependent conformational preferences of IDPs in atomic detail. Our simulation captured intrinsic conformational characteristics of KID that are in good agreement with experimental data such as a very small percentage of helical structure in its segment α(B) and structural disorder in solution. We used dihedral principal component analysis dPCA to map the conformations of KID in the microsecond timescale. By using principal components as reaction coordinates, we further constructed dPCA-based free energy landscapes of KID. Analysis of the free energy landscapes showed that KID is best characterized as a conformational ensemble of rapidly interconverting conformations. Interestingly, we found that despite the conformational heterogeneity of the backbone and the absence of substantial secondary structure, KID does not randomly sample the conformational space in solution: analysis of the (Φ, Ψ) dihedral angles showed that several individual residues of KID possess a strong bias toward the helical region of the Ramachandran plot. We suggest that the intrinsic conformational preferences of KID provide a bias toward the folded state without having to populate bound-like conformations before binding. Furthermore, we argue that these conformational preferences do not represent actual structural constraints which drive binding through a single pathway, which allows for specific interactions with multiple binding partners. Based on this evidence, we propose that the backbone conformational preferences of KID provide a thermodynamic advantage for folding and binding without negatively affecting the kinetics of binding. We further discuss the relation of our results to previous studies to rationalize the functional implications of the conformational preferences of IDPs, such as the optimization of structural disorder in protein-protein interactions. This study illustrates the importance in obtaining atomistic information of intrinsically disordered proteins in real time to reveal functional features arising from their complex conformational space.

Crystal Structures of the Luciferase and Green Fluorescent Protein from Renilla reniformis

PubMed Central

Loening, Andreas Markus; Fenn, Timothy David; Gambhir, Sanjiv Sam

2009-01-01

Due to its ability to emit light, the luciferase from Renilla reniformis (RLuc) is widely employed in molecular biology as a reporter gene in both cell culture experiments and small animal imaging. To accomplish this bioluminesce, the 37 KDa enzyme catalyzes the degradation of its substrate coelenterazine in the presence of molecular oxygen, resulting in the product coelenteramide, carbon dioxide, and the desired photon of light. We successfully crystallized a stabilized variant of this important protein (RLuc8), and present the first structures for any coelenterazine-using luciferase. These structures are based on high resolution data measured to 1.4 Å and demonstrate a classic α/β-hydrolase fold. We also present data of a coe-lenteramide bound-luciferase, and reason that this structure represents a secondary conformational form following shift of the product out of the primary active site. During the course of this work, the structure of the luciferase’s accessory green fluorescent protein (RrGFP) was determined as well and shown to be highly similar to that of Aequorea GFP. PMID:17980388

Crystal structures of the luciferase and green fluorescent protein from Renilla reniformis.

PubMed

Loening, Andreas Markus; Fenn, Timothy David; Gambhir, Sanjiv Sam

2007-12-07

Due to its ability to emit light, the luciferase from Renilla reniformis (RLuc) is widely employed in molecular biology as a reporter gene in cell culture experiments and small animal imaging. To accomplish this bioluminescence, the 37-kDa enzyme catalyzes the degradation of its substrate coelenterazine in the presence of molecular oxygen, resulting in the product coelenteramide, carbon dioxide, and the desired photon of light. We successfully crystallized a stabilized variant of this important protein (RLuc8) and herein present the first structures for any coelenterazine-using luciferase. These structures are based on high-resolution data measured to 1.4 A and demonstrate a classic alpha/beta-hydrolase fold. We also present data of a coelenteramide-bound luciferase and reason that this structure represents a secondary conformational form following shift of the product out of the primary active site. During the course of this work, the structure of the luciferase's accessory green fluorescent protein (RrGFP) was also determined and shown to be highly similar to that of Aequorea victoria GFP.

Structure of Urtica dioica agglutinin isolectin I: dimer formation mediated by two zinc ions bound at the sugar-binding site.

PubMed

Harata, K; Schubert, W D; Muraki, M

2001-11-01

Ultica dioica agglutinin, a plant lectin from the stinging nettle, consists of a total of seven individual isolectins. One of these structures, isolectin I, was determined at 1.9 A resolution by the X-ray method. The crystals belong to the space group P2(1) and the asymmetric unit contains two molecules related by local twofold symmetry. The molecule consists of two hevein-like chitin-binding domains lacking distinct secondary structure, but four disulfide bonds in each domain maintain the tertiary structure. The backbone structure of the two independent molecules is essentially identical and this is similarly true of the sugar-binding sites. In the crystal, the C-terminal domains bind Zn(2+) ions at the sugar-binding site. Owing to their location near a pseudo-twofold axis, the two zinc ions link the two independent molecules in a tail-to-tail arrangement: thus, His47 of molecule 1 and His67 of molecule 2 coordinate the first zinc ion, while the second zinc ion links Asp75 of molecule 1 and His47 of molecule 2.

Human γ-Glutamyl Transpeptidase 1: STRUCTURES OF THE FREE ENZYME, INHIBITOR-BOUND TETRAHEDRAL TRANSITION STATES, AND GLUTAMATE-BOUND ENZYME REVEAL NOVEL MOVEMENT WITHIN THE ACTIVE SITE DURING CATALYSIS.

PubMed

Terzyan, Simon S; Burgett, Anthony W G; Heroux, Annie; Smith, Clyde A; Mooers, Blaine H M; Hanigan, Marie H

2015-07-10

γ-Glutamyl transpeptidase 1 (GGT1) is a cell surface, N-terminal nucleophile hydrolase that cleaves glutathione and other γ-glutamyl compounds. GGT1 expression is essential in cysteine homeostasis, and its induction has been implicated in the pathology of asthma, reperfusion injury, and cancer. In this study, we report four new crystal structures of human GGT1 (hGGT1) that show conformational changes within the active site as the enzyme progresses from the free enzyme to inhibitor-bound tetrahedral transition states and finally to the glutamate-bound structure prior to the release of this final product of the reaction. The structure of the apoenzyme shows flexibility within the active site. The serine-borate-bound hGGT1 crystal structure demonstrates that serine-borate occupies the active site of the enzyme, resulting in an enzyme-inhibitor complex that replicates the enzyme's tetrahedral intermediate/transition state. The structure of GGsTop-bound hGGT1 reveals its interactions with the enzyme and why neutral phosphonate diesters are more potent inhibitors than monoanionic phosphonates. These structures are the first structures for any eukaryotic GGT that include a molecule in the active site covalently bound to the catalytic Thr-381. The glutamate-bound structure shows the conformation of the enzyme prior to release of the final product and reveals novel information regarding the displacement of the main chain atoms that form the oxyanion hole and movement of the lid loop region when the active site is occupied. These data provide new insights into the mechanism of hGGT1-catalyzed reactions and will be invaluable in the development of new classes of hGGT1 inhibitors for therapeutic use. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Human γ-glutamyl transpeptidase 1: Structures of the free enzyme, inhibitor-bound tetrahedral transition states, and glutamate-bound enzyme reveal novel movement within the active site during catalysis [Human gamma-glutamyl transpeptidase: Inhibitor binding and movement within the active site

DOE PAGES

Terzyan, Simon S.; Burgett, Anthony W. G.; Heroux, Annie; ...

2015-05-26

γ-Glutamyl transpeptidase 1 (GGT1) is a cell surface, N-terminal nucleophile hydrolase that cleaves glutathione and other γ-glutamyl compounds. GGT1 expression is essential in cysteine homeostasis, and its induction has been implicated in the pathology of asthma, reperfusion injury, and cancer. In this study, we report four new crystal structures of human GGT1 (hGGT1) that show conformational changes within the active site as the enzyme progresses from the free enzyme to inhibitor-bound tetrahedral transition states and finally to the glutamate-bound structure prior to the release of this final product of the reaction. The structure of the apoenzyme shows flexibility within themore » active site. The serine-borate-bound hGGT1 crystal structure demonstrates that serine-borate occupies the active site of the enzyme, resulting in an enzyme-inhibitor complex that replicates the enzyme's tetrahedral intermediate/transition state. The structure of GGsTop-bound hGGT1 reveals its interactions with the enzyme and why neutral phosphonate diesters are more potent inhibitors than monoanionic phosphonates. These structures are the first structures for any eukaryotic GGT that include a molecule in the active site covalently bound to the catalytic Thr-381. The glutamate-bound structure shows the conformation of the enzyme prior to release of the final product and reveals novel information regarding the displacement of the main chain atoms that form the oxyanion hole and movement of the lid loop region when the active site is occupied. Lastly,tThese data provide new insights into the mechanism of hGGT1-catalyzed reactions and will be invaluable in the development of new classes of hGGT1 inhibitors for therapeutic use.« less

Human γ-glutamyl transpeptidase 1: Structures of the free enzyme, inhibitor-bound tetrahedral transition states, and glutamate-bound enzyme reveal novel movement within the active site during catalysis [Human gamma-glutamyl transpeptidase: Inhibitor binding and movement within the active site

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

Terzyan, Simon S.; Burgett, Anthony W. G.; Heroux, Annie

γ-Glutamyl transpeptidase 1 (GGT1) is a cell surface, N-terminal nucleophile hydrolase that cleaves glutathione and other γ-glutamyl compounds. GGT1 expression is essential in cysteine homeostasis, and its induction has been implicated in the pathology of asthma, reperfusion injury, and cancer. In this study, we report four new crystal structures of human GGT1 (hGGT1) that show conformational changes within the active site as the enzyme progresses from the free enzyme to inhibitor-bound tetrahedral transition states and finally to the glutamate-bound structure prior to the release of this final product of the reaction. The structure of the apoenzyme shows flexibility within themore » active site. The serine-borate-bound hGGT1 crystal structure demonstrates that serine-borate occupies the active site of the enzyme, resulting in an enzyme-inhibitor complex that replicates the enzyme's tetrahedral intermediate/transition state. The structure of GGsTop-bound hGGT1 reveals its interactions with the enzyme and why neutral phosphonate diesters are more potent inhibitors than monoanionic phosphonates. These structures are the first structures for any eukaryotic GGT that include a molecule in the active site covalently bound to the catalytic Thr-381. The glutamate-bound structure shows the conformation of the enzyme prior to release of the final product and reveals novel information regarding the displacement of the main chain atoms that form the oxyanion hole and movement of the lid loop region when the active site is occupied. Lastly,tThese data provide new insights into the mechanism of hGGT1-catalyzed reactions and will be invaluable in the development of new classes of hGGT1 inhibitors for therapeutic use.« less

Unliganded HIV-1 gp120 core structures assume the CD4-bound conformation with regulation by quaternary interactions and variable loops

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

Kwon, Young Do; Finzi, Andrés; Wu, Xueling

2013-03-04

The HIV-1 envelope (Env) spike (gp120{sub 3}/gp41{sub 3}) undergoes considerable structural rearrangements to mediate virus entry into cells and to evade the host immune response. Engagement of CD4, the primary human receptor, fixes a particular conformation and primes Env for entry. The CD4-bound state, however, is prone to spontaneous inactivation and susceptible to antibody neutralization. How does unliganded HIV-1 maintain CD4-binding capacity and regulate transitions to the CD4-bound state? To define this mechanistically, we determined crystal structures of unliganded core gp120 from HIV-1 clades B, C, and E. Notably, all of these unliganded HIV-1 structures resembled the CD4-bound state. Conformationalmore » fixation with ligand selection and thermodynamic analysis of full-length and core gp120 interactions revealed that the tendency of HIV-1 gp120 to adopt the CD4-bound conformation was restrained by the V1/V2- and V3-variable loops. In parallel, we determined the structure of core gp120 in complex with the small molecule, NBD-556, which specifically recognizes the CD4-bound conformation of gp120. Neutralization by NBD-556 indicated that Env spikes on primary isolates rarely assume the CD4-bound conformation spontaneously, although they could do so when quaternary restraints were loosened. Together, the results suggest that the CD4-bound conformation represents a 'ground state' for the gp120 core, with variable loop and quaternary interactions restraining unliganded gp120 from 'snapping' into this conformation. A mechanism of control involving deformations in unliganded structure from a functionally critical state (e.g., the CD4-bound state) provides advantages in terms of HIV-1 Env structural diversity and resistance to antibodies and inhibitors, while maintaining elements essential for entry.« less

Investigating plausible mechanisms for the photo-induced partial unfolding of a globular protein

NASA Astrophysics Data System (ADS)

Parker, James E.

Two hypotheses are proposed to explain the photo-induced unfolding of β-lactoglobulin (BLG) that occurs when non-covalently bound to a dye molecule, meso-tetrakis (p-sulfonatophenyl) porphyrin (TSPP), and illuminated by a laser in the post-Tanford transition configuration. The first involves a photo-induced electron transfer from the porphyrin to the protein. The second involves the production of kynurenine by singlet oxygen that is generated during photo-excitation of the porphyrin. To evaluate these hypotheses, a series of computational and experimental results have been combined to establish the physical state of the BLG-TSPP complex and to estimate the likelihood of a post-irradiation event to initiate the partial unfolding. Determining the binding site location is crucial to establish the position of the photo-induced events and the likely end-product. A study of the vibronic state of the BLG-TSPP complex using resonant Raman and absorption spectroscopy coupled with density functional theory (DFT) and docking simulations is used to estimate the location of the binding site. Once the binding site is found, molecular dynamics simulations of the post-irradiation event relaxations in the protein are used to estimate the resulting secondary structure. This structure is compared to experimental estimates of the secondary structure of the unfolded protein to determine which hypothesis is the most likely mechanism to explain the unfolding.

Unusual sugar specificity of banana lectin from Musa paradisiaca and its probable evolutionary origin. Crystallographic and modelling studies.

PubMed

Singh, D D; Saikrishnan, K; Kumar, Prashant; Surolia, A; Sekar, K; Vijayan, M

2005-10-01

The crystal structure of a complex of methyl-alpha-D-mannoside with banana lectin from Musa paradisiaca reveals two primary binding sites in the lectin, unlike in other lectins with beta-prism I fold which essentially consists of three Greek key motifs. It has been suggested that the fold evolved through successive gene duplication and fusion of an ancestral Greek key motif. In other lectins, all from dicots, the primary binding site exists on one of the three motifs in the three-fold symmetric molecule. Banana is a monocot, and the three motifs have not diverged enough to obliterate sequence similarity among them. Two Greek key motifs in it carry one primary binding site each. A common secondary binding site exists on the third Greek key. Modelling shows that both the primary sites can support 1-2, 1-3, and 1-6 linked mannosides with the second residue interacting in each case primarily with the secondary binding site. Modelling also readily leads to a bound branched mannopentose with the nonreducing ends of the two branches anchored at the two primary binding sites, providing a structural explanation for the lectin's specificity for branched alpha-mannans. A comparison of the dimeric banana lectin with other beta-prism I fold lectins, provides interesting insights into the variability in their quaternary structure.

Fast concurrent array-based stacks, queues and deques using fetch-and-increment-bounded, fetch-and-decrement-bounded and store-on-twin synchronization primitives

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

Chen, Dong; Gara, Alana; Heidelberger, Philip

Implementation primitives for concurrent array-based stacks, queues, double-ended queues (deques) and wrapped deques are provided. In one aspect, each element of the stack, queue, deque or wrapped deque data structure has its own ticket lock, allowing multiple threads to concurrently use multiple elements of the data structure and thus achieving high performance. In another aspect, new synchronization primitives FetchAndIncrementBounded (Counter, Bound) and FetchAndDecrementBounded (Counter, Bound) are implemented. These primitives can be implemented in hardware and thus promise a very fast throughput for queues, stacks and double-ended queues.

Thick electrodes including nanoparticles having electroactive materials and methods of making same

DOEpatents

Xiao, Jie; Lu, Dongping; Liu, Jun; Zhang, Jiguang; Graff, Gordon L.

2017-02-21

Electrodes having nanostructure and/or utilizing nanoparticles of active materials and having high mass loadings of the active materials can be made to be physically robust and free of cracks and pinholes. The electrodes include nanoparticles having electroactive material, which nanoparticles are aggregated with carbon into larger secondary particles. The secondary particles can be bound with a binder to form the electrode.

Nuclear export receptor CRM1 recognizes diverse conformations in nuclear export signals.

PubMed

Fung, Ho Yee Joyce; Fu, Szu-Chin; Chook, Yuh Min

2017-03-10

Nuclear export receptor CRM1 binds highly variable nuclear export signals (NESs) in hundreds of different cargoes. Previously we have shown that CRM1 binds NESs in both polypeptide orientations (Fung et al., 2015). Here, we show crystal structures of CRM1 bound to eight additional NESs which reveal diverse conformations that range from loop-like to all-helix, which occupy different extents of the invariant NES-binding groove. Analysis of all NES structures show 5-6 distinct backbone conformations where the only conserved secondary structural element is one turn of helix that binds the central portion of the CRM1 groove. All NESs also participate in main chain hydrogen bonding with human CRM1 Lys568 side chain, which acts as a specificity filter that prevents binding of non-NES peptides. The large conformational range of NES backbones explains the lack of a fixed pattern for its 3-5 hydrophobic anchor residues, which in turn explains the large array of peptide sequences that can function as NESs.

Deciphering the toxicity of bisphenol a to Candida rugosa lipase through spectrophotometric methods.

PubMed

Zhang, Rui; Zhao, Lining; Liu, Rutao

2016-10-01

Bisphenol A is widely used in the manufacture of food packaging and beverage containers and can invade our food and cause contamination. Candida rugose lipase has been a versatile enzyme for biocatalysis and biotransformations to produce useful materials for food, pharmaceutical and flavor. The interactions between bisphenol A and Candida rugosa lipase in vitro were studied by UV-vis, steady-state fluorescence, circular dichroism, synchronous fluorescence, light scattering spectra, molecular docking and enzyme activity assay to better understand the toxicity and toxic mechanisms of bisphenol A. The intrinsic fluorescence of the tryptophan amino acid residue and the secondary structure of the globular protein candida rugose lipase were made use of to thoroughly investigate the structural changes caused by bisphenol A. The results of the fluorescence indicated that bisphenol A interacted with candida rugose lipase and made tryptophan be exposed to a hydrophobic environment. Multi-spectroscopic measurements showed that the addition of bisphenol A increased the intrinsic fluorescence of Candida rugosa lipase, loosened its skeleton structure and changed its secondary structure. Also, the increased activity of Candida rugosa lipase revealed that the position or the structure of the catalytic triad of Candida rugosa lipase may be changed. The molecular docking results showed that bisphenol A bound with the residue Serine 209 which could be another reason for the increased activity of Candida rugosa lipase. Moreover, as can be seen from the results of resonance light scattering and dynamic light scattering, the volume of the Candida rugosa lipase was decreased and the lid may be stripped. Copyright © 2016 Elsevier B.V. All rights reserved.

TIME STRUCTURE OF GAMMA-RAY SIGNALS GENERATED IN LINE-OF-SIGHT INTERACTIONS OF COSMIC RAYS FROM DISTANT BLAZARS

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

Prosekin, Anton; Aharonian, Felix; Essey, Warren

2012-10-01

Blazars are expected to produce both gamma rays and cosmic rays. Therefore, observed high-energy gamma rays from distant blazars may contain a significant contribution from secondary gamma rays produced along the line of sight by the interactions of cosmic-ray protons with background photons. Unlike the standard models of blazars that consider only the primary photons emitted at the source, models that include the cosmic-ray contribution predict that even {approx}10 TeV photons should be detectable from distant objects with redshifts as high as z {>=} 0.1. Secondary photons contribute to signals of point sources only if the intergalactic magnetic fields aremore » very small, B {approx}< 10{sup -14} G, and their detection can be used to set upper bounds on magnetic fields along the line of sight. Secondary gamma rays have distinct spectral and temporal features. We explore the temporal properties of such signals using a semi-analytical formalism and detailed numerical simulations, which account for all the relevant processes, including magnetic deflections. In particular, we elucidate the interplay of time delays coming from the proton deflections and from the electromagnetic cascade, and we find that, at multi-TeV energies, secondary gamma rays can show variability on timescales of years for B {approx} 10{sup -15} G.« less

Comprehensive studies on the nature of interaction between carboxylated multi-walled carbon nanotubes and bovine serum albumin.

PubMed

Lou, Kai; Zhu, Zhaohua; Zhang, Hongmei; Wang, Yanqing; Wang, Xiaojiong; Cao, Jian

2016-01-05

Herein, the interaction between carboxylated multi-walled carbon nanotubes (MWCNTs-COOH) and bovine serum albumin has been investigated by using circular dichroism, UV-vis, and fluorescence spectroscopic methods and molecular modeling in order to better understand the basic behavior of carbon nanotubes in biological systems. The spectral results showed that MWCNTs-COOH bound to BSA and induced the relatively large changes in secondary structure of protein by mainly hydrophobic forces and π-π stacking interactions. Thermal denaturation of BSA in the presence of MWCNTs-COOH indicated that carbon nanotubes acted as a structure destabilizer for BSA. In addition, the putative binding site of MWCNTs-COOH on BSA was near to domain II. With regard to human health, the present study could provide a better understanding of the biological properties, cytotocicity of surface modified carbon nanotubes. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Mathematical model of susceptibility, resistance, and resilience in the within-host dynamics between a Plasmodium parasite and the immune system.

PubMed

Yan, Yi; Adam, Brian; Galinski, Mary; C Kissinger, Jessica; Moreno, Alberto; Gutierrez, Juan B

2015-12-01

We developed a coupled age-structured partial differential equation model to capture the disease dynamics during blood-stage malaria. The addition of age structure for the parasite population, with respect to previous models, allows us to better characterize the interaction between the malaria parasite and red blood cells during infection. Here we prove that the system we propose is well-posed and there exist at least two global states. We further demonstrate that the numerical simulation of the system coincides with clinically observed outcomes of primary and secondary malaria infection. The well-posedness of this system guarantees that the behavior of the model remains smooth, bounded, and continuously dependent on initial conditions; calibration with clinical data will constrain domains of parameters and variables to physiological ranges. Copyright © 2015 Elsevier Inc. All rights reserved.

An investigation of the effects of spanwise wall oscillation on the structure of a turbulent boundary layer

NASA Astrophysics Data System (ADS)

Trujillo, Steven Mathew

Transition of a fluid boundary layer from a laminar to a turbulent regime is accompanied by a large increase in skin friction drag. The ability to manipulate the flow or its bounding geometry to reduce this drag effectively has been a long-sought goal in contemporary fluid mechanics. Recently, workers have demonstrated that continuous lateral oscillation of the flow's bounding surface is one means to this goal, producing significant drag reduction. The present study was performed to understand better the mechanism by which such a flow achieves drag reduction. An oscillating wall section was installed in a water channel facility, and the resulting flow was studied using laser Doppler velocimetry, hot-film anemometry, and visualization techniques. Traditional mean and fluctuating statistics were examined, as well as statistics computed from conditionally-sampled turbulent events. The dependence of these quantities on the phase of the oscillating surface's motion was also studied. Visualization-based studies were employed to provide insight into the structural changes brought on by the wall oscillation. The most dramatic changes effected by the wall motion were seen as reductions in frequency of bursts and sweeps, events which concentrate large production of Reynolds stress and which ultimately augment wall skin friction. These Reynolds-stress reductions were reflected in reductions in mean and fluctuating quantifies in the lower regions of the boundary layer. Other velocity measurements confirmed earlier workers' speculations that the secondary flow induced by the oscillating wall is comparable to Stokes' solution for an oscillating plate in a quiescent fluid. Other than this secondary flow, however, the boundary layer displayed essentially no dependence on the phase of the wall motion. A simple cost analysis showed that, in general, the energy cost required to implement this technique is greater than the savings it produces. The visualizations of the flow revealed a more uniform flow in the near-wall region resulting from wall oscillation. Quantitative analyses of the visualizations supported the velocity-based Reynolds-stress reductions; the same data also revealed that the quasi-streamwise vortical structures above the wall did not appear to be altered significantly by the wall motion.

Fluoride-Mediated Capture of a Noncovalent Bound State of a Reversible Covalent Enzyme Inhibitor: X-ray Crystallographic Analysis of an Exceptionally Potent [alpha]-Ketoheterocycle Inhibitor of Fatty Acid Amide Hydrolase

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

Mileni, Mauro; Garfunkle, Joie; Ezzili, Cyrine

2011-11-02

Two cocrystal X-ray structures of the exceptionally potent {alpha}-ketoheterocycle inhibitor 1 (K{sub i} = 290 pM) bound to a humanized variant of rat fatty acid amide hydrolase (FAAH) are disclosed, representing noncovalently and covalently bound states of the same inhibitor with the enzyme. Key to securing the structure of the noncovalently bound state of the inhibitor was the inclusion of fluoride ion in the crystallization conditions that is proposed to bind the oxyanion hole precluding inhibitor covalent adduct formation with stabilization of the tetrahedral hemiketal. This permitted the opportunity to detect important noncovalent interactions stabilizing the binding of the inhibitormore » within the FAAH active site independent of the covalent reaction. Remarkably, noncovalently bound 1 in the presence of fluoride appears to capture the active site in the same 'in action' state with the three catalytic residues Ser241-Ser217-Lys142 occupying essentially identical positions observed in the covalently bound structure of 1, suggesting that this technique of introducing fluoride may have important applications in structural studies beyond inhibiting substrate or inhibitor oxyanion hole binding. Key insights to emerge from the studies include the observations that noncovalently bound 1 binds in its ketone (not gem diol) form, that the terminal phenyl group in the acyl side chain of the inhibitor serves as the key anchoring interaction overriding the intricate polar interactions in the cytosolic port, and that the role of the central activating heterocycle is dominated by its intrinsic electron-withdrawing properties. These two structures are also briefly compared with five X-ray structures of {alpha}-ketoheterocycle-based inhibitors bound to FAAH recently disclosed.« less

Conformational Plasticity of the Influenza A M2 Transmembrane Helix in Lipid Bilayers Under Varying pH, Drug Binding and Membrane Thickness

PubMed Central

Hu, Fanghao; Luo, Wenbin; Cady, Sarah D.; Hong, Mei

2010-01-01

Membrane proteins change their conformations to respond to environmental cues, thus conformational plasticity is important for function. The influenza A M2 protein forms an acid-activated proton channel important for the virus lifecycle. Here we have used solid-state NMR spectroscopy to examine the conformational plasticity of membrane-bound transmembrane domain of M2 (M2TM). 13C and 15N chemical shifts indicate coupled conformational changes of several pore-facing residues due to changes in bilayer thickness, drug binding and pH. The structural changes are attributed to the formation of a well-defined helical kink at G34 in the drug-bound state and in thick lipid bilayers, non-ideal backbone conformation of the secondary-gate residue V27 in the presence of drug, and non-ideal conformation of the proton-sensing residue H37 at high pH. The chemical shifts constrained the (ϕ, ψ) torsion angles for three basis states, the equilibrium among which explains the multiple resonances per site in the NMR spectra under different combinations of bilayer thickness, drug binding and pH conditions. Thus, conformational plasticity is important for the proton conduction and inhibition of M2TM. The study illustrates the utility of NMR chemical shifts for probing the structural plasticity and folding of membrane proteins. PMID:20883664

Pd-Catalyzed N-Arylation of Secondary Acyclic Amides: Catalyst Development, Scope, and Computational Study

PubMed Central

Hicks, Jacqueline D.; Hyde, Alan M.; Cuezva, Alberto Martinez; Buchwald, Stephen L.

2009-01-01

We report the efficient N-arylation of acyclic secondary amides and related nucleophiles with aryl nonaflates, triflates, and chlorides. This method allows for easy variation of the aromatic component in tertiary aryl amides. A new biaryl phosphine with P-bound 3,5-(bis)trifluoromethylphenyl groups was found to be uniquely effective for this amidation. The critical aspects of the ligand were explored through synthetic, mechanistic, and computational studies. Systematic variation of the ligand revealed the importance of (1) a methoxy group on the aromatic carbon of the “top ring” ortho to the phosphorus and (2) two highly electron-withdrawing P-bound 3,5-(bis)trifluoromethylphenyl groups. Computational studies suggest the electron-deficient nature of the ligand is important in facilitating amide binding to the LPd(II)(Ph)(X) intermediate. PMID:19886610

Helix A Stabilization Precedes Amino-terminal Lobe Activation upon Calcium Binding to Calmodulin

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

Chen, Baowei; Lowry, David; Mayer, M. Uljana

2008-08-09

The structural coupling between opposing domains of CaM was investigated using the conformationally sensitive biarsenical probe 4,5-bis(1,3,2-dithioarsolan-2-yl)-resorufin (ReAsH), which upon binding to an engineered tetracysteine binding motif near the end of helix A (Thr-5 to Phe-19) becomes highly fluorescent. Changes in conformation and dynamics are reflective of the native CaM structure, as there is no change in the 1H- 15N HSQC NMR spectrum in comparison to wild-type CaM. We find evidence of a conformational intermediate associated with CaM activation, where calcium occupancy of sites in the amino-terminal and carboxyl-terminal lobes of CaM differentially affect the fluorescence intensity of bound ReAsH.more » Insight into the structure of the conformational intermediate is possible from a consideration of calcium-dependent changes in rates of ReAsH binding and helix A mobility, which respectively distinguish secondary structural changes associated with helix A stabilization from the tertiary structural reorganization of the amino-terminal lobe of CaM necessary for high-affinity binding to target proteins. Helix A stabilization is associated with calcium occupancy of sites in the carboxyl-terminal lobe (Kd = 0.36 ± 0.04 μM), which results in a reduction in the rate of ReAsH binding from 4900 M -1 sec -1 to 370 M -1 sec -1. In comparison, tertiary structural changes involving helix A and other structural elements in the amino-terminal lobe requires calcium-occupancy of amino-terminal sites (Kd = 18 ± 3 μM). Observed secondary and tertiary structural changes involving helix A in response to the sequential calcium occupancy of carboxyl- and amino-terminal lobe calcium binding sites suggest an important involvement of helix A in mediating the structural coupling between the opposing domains of CaM. These results are discussed in terms of a model in which carboxyl-terminal lobe calcium activation induces secondary structural changes within the interdomain linker that release helix A, thereby facilitating the formation of calcium binding sites in the amino-terminal lobe and linked tertiary structural rearrangements to form a high-affinity binding cleft that can associate with target proteins.« less

Thermal crystallization mechanism of silk fibroin protein

NASA Astrophysics Data System (ADS)

Hu, Xiao

In this thesis, the thermal crystallization mechanism of silk fibroin protein from Bombyx mori silkworm, was treated as a model for the general study of protein based materials, combining theories from both biophysics and polymer physics fields. A systematic and scientific path way to model the dynamic beta-sheet crystallization process of silk fibroin protein was presented in the following sequence: (1) The crystallinity, fractions of secondary structures, and phase compositions in silk fibroin proteins at any transition stage were determined. Two experimental methods, Fourier transform infrared spectroscopy (FTIR) with Fourier self-deconvolution, and specific reversing heat capacity, were used together for the first time for modeling the static structures and phases in the silk fibroin proteins. The protein secondary structure fractions during the crystallization were quantitatively determined. The possibility of existence of a "rigid amorphous phase" in silk protein was also discussed. (2) The function of bound water during the crystallization process of silk fibroin was studied using heat capacity, and used to build a silk-water dynamic crystallization model. The fundamental concepts and thermal properties of silk fibroin with/without bound water were discussed. Results show that intermolecular bound water molecules, acting as a plasticizer, will cause silk to display a water-induced glass transition around 80°C. During heating, water is lost, and the change of the microenvironment in the silk fibroin chains induces a mesophase prior to thermal crystallization. Real time FTIR during heating and isothermal holding above Tg show the tyrosine side chain changes only during the former process, while beta sheet crystallization occurs only during the latter process. Analogy is made between the crystallization of synthetic polymers according to the four-state scheme of Strobl, and the crystallization process of silk fibroin, which includes an intermediate precursor stage before crystallization. (3) The beta-sheet crystallization kinetics in silk fibroin protein were measured using X-ray, FTIR and heat flow, and the structure reveals the formation mechanism of the silk crystal network. Avrami kinetics theories, which were established for studies of synthetic polymer crystal growth, were for the first time extended to investigate protein self-assembly in multiblock silk fibroin samples. The Avrami exponent, n, was close to two for all methods, indicating formation of beta sheet crystals in silk proteins is different from the 3-D spherulitic crystal growth found in most synthetic homopolymers. A microphase separation pattern after chymotrypsin enzyme biodegradation was shown in the protein structures using scanning electron microscopy. A model was then used to explain the crystallization of silk fibroin protein by analogy to block copolymers. (4) The effects of metal ions during the crystallization of silk fibroin was investigated using thermal analysis. Advanced thermal analysis methods were used to analyze the thermal protein-metallic ion interactions in silk fibroin proteins. Results show that K+ and Ca2+ metallic salts play different roles in silk fibroin proteins, which either reduce (K+) or increase (Ca2+ ) the glass transition (Tg) of pure silk protein and affect the thermal stability of this structure.

Isolation and characterization of the genes for two small RNAs of herpesvirus papio and their comparison with Epstein-Barr virus-encoded EBER RNAs.

PubMed

Howe, J G; Shu, M D

1988-08-01

Genes for the Epstein-Barr virus-encoded RNAs (EBERs), two low-molecular-weight RNAs encoded by the human gammaherpesvirus Epstein-Barr virus (EBV), hybridize to two small RNAs in a baboon cell line that contains a similar virus, herpesvirus papio (HVP). The genes for the HVP RNAs (HVP-1 and HVP-2) are located together in the small unique region at the left end of the viral genome and are transcribed by RNA polymerase III in a rightward direction, similar to the EBERs. There is significant similarity between EBER1 and HVP-1 RNA, except for an insert of 22 nucleotides which increases the length of HVP-1 RNA to 190 nucleotides. There is less similarity between the sequences of EBER2 and HVP-2 RNA, but both have a length of about 170 nucleotides. The predicted secondary structure of each HVP RNA is remarkably similar to that of the respective EBER, implying that the secondary structures are important for function. Upstream from the initiation sites of all four RNA genes are several highly conserved sequences which may function in the regulation of transcription. The HVP RNAs, together with the EBERs, are highly abundant in transformed cells and are efficiently bound by the cellular La protein.

Accelerating the Conformational Sampling of Intrinsically Disordered Proteins.

PubMed

Do, Trang Nhu; Choy, Wing-Yiu; Karttunen, Mikko

2014-11-11

Intrinsically disordered proteins (IDPs) are a class of proteins lacking a well-defined secondary structure. Instead, they are able to attain multiple conformations, bind to multiple targets, and respond to changes in their surroundings. Functionally, IDPs have been associated with molecular recognition, cell regulation, and signal transduction. The dynamic conformational ensemble of IDPs is highly environmental and binding partner dependent, rendering the characterization of IDPs extremely challenging. Here, we compare the sampling efficiencies of conventional molecular dynamics (MD), well-tempered metadynamics (WT-META), and bias-exchange metadynamics (BE-META). The total simulation time was over 10 μs, and a 20-mer peptide derived from the Neh2 domain of the Nuclear factor erythroid 2-related factor 2 (Nrf2) protein was simulated. BE-META, with a neutral replica and seven biased replicas employing a set of seven relevant collective variables (CVs), provided the most reliable and efficient sampling. Finally, we propose a free-energy reconstruction method based on the probability distribution of the secondary structure contents. This postprocessing analysis confirms the presence of not only the β-hairpin conformation of the free Neh2 peptide but also its rare bound-state-like conformation, both of that have been experimentally observed. In addition, our simulations also predict other possible conformations to be verified with future experiments.

Crystalline and amorphous cellulose in the secondary walls of Arabidopsis.

PubMed

Ruel, Katia; Nishiyama, Yoshiharu; Joseleau, Jean-Paul

2012-09-01

In the cell walls of higher plants, cellulose chains are present in crystalline microfibril, with an amorphous part at the surface, or present as amorphous material. To assess the distribution and relative occurrence of the two forms of cellulose in the inflorescence stem of Arabidopsis, we used two carbohydrate-binding modules, CBM3a and CBM28, specific for crystalline and amorphous cellulose, respectively, with immunogold detection in TEM. The binding of the two CBMs displayed specific patterns suggesting that the synthesis of cellulose leads to variable nanodomains of cellulose structures according to cell type. In developing cell walls, only CBM3a bound significantly to the incipient primary walls, indicating that at the onset of its deposition cellulose is in a crystalline structure. As the secondary wall develops, the labeling with both CBMs becomes more intense. The variation of the labeling pattern by CBM3a between transverse and longitudinal sections appeared related to microfibril orientation and differed between fibers and vessels. Although the two CBMs do not allow the description of the complete status of cellulose microstructures, they revealed the dynamics of the deposition of crystalline and amorphous forms of cellulose during wall formation and between cell types adapting cellulose microstructures to the cell function. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

Equilibrium structures and flows of polar and nonpolar liquids in different carbon nanotubes

NASA Astrophysics Data System (ADS)

Abramyan, Andrey K.; Bessonov, Nick M.; Mirantsev, Leonid V.; Chevrychkina, Anastasiia A.

2018-03-01

Molecular dynamics (MD) simulations of equilibrium structures and flows of polar water and nonpolar methane confined by single-walled carbon nanotubes (SWCNTs) with circular and square cross sections and bounding walls with regular graphene structure and random (amorphous) distribution of carbon atoms have been performed. The results of these simulations show that equilibrium structures of both confined liquids depend strongly on the shape of the cross section of SWCNTs, whereas the structure of their bounding walls has a minor influence on these structures. On contrary, the external pressure driven water and methane flows through above mentioned SWCNTs depend significantly on both the shape of their cross sections and the structure of their bounding walls.

Rotation of Guanine Amino Groups in G-Quadruplexes: A Probe for Local Structure and Ligand Binding.

PubMed

Adrian, Michael; Winnerdy, Fernaldo Richtia; Heddi, Brahim; Phan, Anh Tuân

2017-08-22

Nucleic acids are dynamic molecules whose functions may depend on their conformational fluctuations and local motions. In particular, amino groups are dynamic components of nucleic acids that participate in the formation of various secondary structures such as G-quadruplexes. Here, we present a cost-efficient NMR method to quantify the rotational dynamics of guanine amino groups in G-quadruplex nucleic acids. An isolated spectrum of amino protons from a specific tetrad-bound guanine can be extracted from the nuclear Overhauser effect spectroscopy spectrum based on the close proximity between the intra-residue imino and amino protons. We apply the method in different structural contexts of G-quadruplexes and their complexes. Our results highlight the role of stacking and hydrogen-bond interactions in restraining amino-group rotation. The measurement of the rotation rate of individual amino groups could give insight into the dynamic processes occurring at specific locations within G-quadruplex nucleic acids, providing valuable probes for local structure, dynamics, and ligand binding. Copyright © 2017 Biophysical Society. Published by Elsevier Inc. All rights reserved.

The Ancient Greeks Speak to Us: A New Humanistic Approach to Classical Greek and Greek Culture for Secondary Schools. Teacher's Guide-Level Alpha.

ERIC Educational Resources Information Center

Masciantonio, Rudolph; And Others

A humanistic approach to the study of classical Greek and Greek culture at the secondary school level is detailed in this guide. References to the student programed text and other multisensory instructional materials used in the system focus on instructional objectives geared to students who are not necessarily college-bound. The standard Attic…

Upper bound dose values for meson radiation in heavy-ion therapy.

PubMed

Rabin, C; Gonçalves, M; Duarte, S B; González-Sprinberg, G A

2018-06-01

Radiation treatment of cancer has evolved to include massive particle beams, instead of traditional irradiation procedures. Thus, patient doses and worker radiological protection have become issues of constant concern in the use of these new technologies, especially for proton- and heavy-ion-therapy. In the beam energies of interest of heavy-ion-therapy, secondary particle radiation comes from proton, neutron, and neutral and charged pions produced in the nuclear collisions of the beam with human tissue atoms. This work, for the first time, offers the upper bound of meson radiation dose in organic tissues due to secondary meson radiation in heavy-ion therapy. A model based on intranuclear collision has been used to follow in time the nuclear reaction and to determine the secondary radiation due to the meson yield produced in the beam interaction with nuclei in the tissue-equivalent media and water. The multiplicity, energy spectrum, and angular distribution of these pions, as well as their decay products, have been calculated in different scenarios for the nuclear reaction mechanism. The results of the produced secondary meson particles has been used to estimate the energy deposited in tissue using a cylindrical phantom by a transport Monte Carlo simulation and we have concluded that these mesons contribute at most 0.1% of the total prescribed dose.

Structural Changes Due to Antagonist Binding in Ligand Binding Pocket of Androgen Receptor Elucidated Through Molecular Dynamics Simulations.

PubMed

Sakkiah, Sugunadevi; Kusko, Rebecca; Pan, Bohu; Guo, Wenjing; Ge, Weigong; Tong, Weida; Hong, Huixiao

2018-01-01

When a small molecule binds to the androgen receptor (AR), a conformational change can occur which impacts subsequent binding of co-regulator proteins and DNA. In order to accurately study this mechanism, the scientific community needs a crystal structure of the Wild type AR (WT-AR) ligand binding domain, bound with antagonist. To address this open need, we leveraged molecular docking and molecular dynamics (MD) simulations to construct a structure of the WT-AR ligand binding domain bound with antagonist bicalutamide. The structure of mutant AR (Mut-AR) bound with this same antagonist informed this study. After molecular docking analysis pinpointed the suitable binding orientation of a ligand in AR, the model was further optimized through 1 μs of MD simulations. Using this approach, three molecular systems were studied: (1) WT-AR bound with agonist R1881, (2) WT-AR bound with antagonist bicalutamide, and (3) Mut-AR bound with bicalutamide. Our structures were very similar to the experimentally determined structures of both WT-AR with R1881 and Mut-AR with bicalutamide, demonstrating the trustworthiness of this approach. In our model, when WT-AR is bound with bicalutamide, Val716/Lys720/Gln733, or Met734/Gln738/Glu897 move and thus disturb the positive and negative charge clumps of the AF2 site. This disruption of the AF2 site is key for understanding the impact of antagonist binding on subsequent co-regulator binding. In conclusion, the antagonist induced structural changes in WT-AR detailed in this study will enable further AR research and will facilitate AR targeting drug discovery.

Structure of a double-domain phosphagen kinase reveals an asymmetric arrangement of the tandem domains.

PubMed

Wang, Zhiming; Qiao, Zhu; Ye, Sheng; Zhang, Rongguang

2015-04-01

Tandem duplications and fusions of single genes have led to magnificent expansions in the divergence of protein structures and functions over evolutionary timescales. One of the possible results is polydomain enzymes with interdomain cooperativities, few examples of which have been structurally characterized at the full-length level to explore their innate synergistic mechanisms. This work reports the crystal structures of a double-domain phosphagen kinase in both apo and ligand-bound states, revealing a novel asymmetric L-shaped arrangement of the two domains. Unexpectedly, the interdomain connections are not based on a flexible hinge linker but on a rigid secondary-structure element: a long α-helix that tethers the tandem domains in relatively fixed positions. Besides the connective helix, the two domains also contact each other directly and form an interdomain interface in which hydrogen bonds and hydrophobic interactions further stabilize the L-shaped domain arrangement. Molecular-dynamics simulations show that the interface is generally stable, suggesting that the asymmetric domain arrangement crystallographically observed in the present study is not a conformational state simply restrained by crystal-packing forces. It is possible that the asymmetrically arranged tandem domains could provide a structural basis for further studies of the interdomain synergy.

The crystal structure of the calcium-bound con-G[Q6A] peptide reveals a novel metal-dependent helical trimer

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

Cnudde, Sara E.; Prorok, Mary; Jia, Xaofei

2012-02-15

The ability to form and control both secondary structure and oligomerization in short peptides has proven to be challenging owing to the structural instability of such peptides. The conantokin peptides are a family of {gamma}-carboxyglutamic acid containing peptides produced in the venoms of predatory sea snails of the Conus family. They are examples of short peptides that form stable helical structures, especially in the presence of divalent cations. Both monomeric and dimeric conantokin peptides have been identified and represent a new mechanism of helix association, 'the metallozipper motif' that is devoid of a hydrophobic interface between monomers. In the presentmore » study, a parallel/antiparallel three-helix bundle was identified and its crystal structure determined at high resolution. The three helices are almost perfectly parallel and represent a novel helix-helix association. The trimer interface is dominated by metal chelation between the three helices, and contains no interfacial hydrophobic interactions. It is now possible to produce stable monomeric, dimeric, or trimeric metallozippers depending on the peptide sequence and metal ion. Such structures have important applications in protein design.« less

Molecular evidence for biodegradation of geomacromolecules

NASA Astrophysics Data System (ADS)

Jenisch-Anton, A.; Adam, P.; Michaelis, W.; Connan, J.; Herrmann, D.; Rohmer, M.; Albrecht, P.

2000-10-01

The biodegradability of macromolecular organic structures of geological origin was investigated by performing in vitro studies. Cultures of the common Nocardioides simplex were grown, first, on a high molecular weight, asymmetric thioether (1-(phytanylsulfanyl)-octadecane 1) and then on macromolecular fractions isolated from a sulfur-rich oil. Gross data indicate that bacteria convert macromolecular substances to material of higher polarity by oxidizing the abundant thioethers to sulfones and sulfoxides and by introducing new functionalities, such as carboxylic acid, keto or hydroxyl groups. Furthermore, bacteria remineralize the macromolecular structures. Bacterially induced alterations were also studied on a molecular level after chemical desulfurization of the macromolecular structure. Thus, it could be established that the amounts of linear hydrocarbons in the macromolecular structure are decreased relative to branched and cyclic structures due to a preferential bacterial attack of the linear moieties bound to the macromolecules. This is further supported by the detection of S-bound fatty acids resulting from the bacterial oxidation of S-bound n-alkanes. Moreover, N. simplex also degraded sulfur-bound steranes by oxidation of the steroid side-chain leading to S-bound steroid acids.

The nucleotide-free state of heterotrimeric G proteins α-subunit adopts a highly stable conformation.

PubMed

Andhirka, Sai Krishna; Vignesh, Ravichandran; Aradhyam, Gopala Krishna

2017-08-01

Deciphering the mechanism of activation of heterotrimeric G proteins by their cognate receptors continues to be an intriguing area of research. The recently solved crystal structure of the ternary complex captured the receptor-bound α-subunit in an open conformation, without bound nucleotide has improved our understanding of the activation process. Despite these advancements, the mechanism by which the receptor causes GDP release from the α-subunit remains elusive. To elucidate the mechanism of activation, we studied guanine nucleotide-induced structural stability of the α-subunit (in response to thermal/chaotrope-mediated stress). Inherent stabilities of the inactive (GDP-bound) and active (GTP-bound) forms contribute antagonistically to the difference in conformational stability whereas the GDP-bound protein is able to switch to a stable intermediate state, GTP-bound protein loses this ability. Partial perturbation of the protein fold reveals the underlying influence of the bound nucleotide providing an insight into the mechanism of activation. An extra stable, pretransition intermediate, 'empty pocket' state (conformationally active-state like) in the unfolding pathway of GDP-bound protein mimics a gating system - the activation process having to overcome this stable intermediate state. We demonstrate that a relatively more complex conformational fold of the GDP-bound protein is at the core of the gating system. We report capturing this threshold, 'metastable empty pocket' conformation (the gate) of α-subunit of G protein and hypothesize that the receptor activates the G protein by enabling it to achieve this structure through mild structural perturbation. © 2017 Federation of European Biochemical Societies.

Structures of the Substrate-free and Product-bound Forms of HmuO, a Heme Oxygenase from Corynebacterium diphtheriae

PubMed Central

Unno, Masaki; Ardèvol, Albert; Rovira, Carme; Ikeda-Saito, Masao

2013-01-01

Heme oxygenase catalyzes the degradation of heme to biliverdin, iron, and carbon monoxide. Here, we present crystal structures of the substrate-free, Fe3+-biliverdin-bound, and biliverdin-bound forms of HmuO, a heme oxygenase from Corynebacterium diphtheriae, refined to 1.80, 1.90, and 1.85 Å resolution, respectively. In the substrate-free structure, the proximal and distal helices, which tightly bracket the substrate heme in the substrate-bound heme complex, move apart, and the proximal helix is partially unwound. These features are supported by the molecular dynamic simulations. The structure implies that the heme binding fixes the enzyme active site structure, including the water hydrogen bond network critical for heme degradation. The biliverdin groups assume the helical conformation and are located in the heme pocket in the crystal structures of the Fe3+-biliverdin-bound and the biliverdin-bound HmuO, prepared by in situ heme oxygenase reaction from the heme complex crystals. The proximal His serves as the Fe3+-biliverdin axial ligand in the former complex and forms a hydrogen bond through a bridging water molecule with the biliverdin pyrrole nitrogen atoms in the latter complex. In both structures, salt bridges between one of the biliverdin propionate groups and the Arg and Lys residues further stabilize biliverdin at the HmuO heme pocket. Additionally, the crystal structure of a mixture of two intermediates between the Fe3+-biliverdin and biliverdin complexes has been determined at 1.70 Å resolution, implying a possible route for iron exit. PMID:24106279

Rosetta FlexPepDock ab-initio: simultaneous folding, docking and refinement of peptides onto their receptors.

PubMed

Raveh, Barak; London, Nir; Zimmerman, Lior; Schueler-Furman, Ora

2011-04-29

Flexible peptides that fold upon binding to another protein molecule mediate a large number of regulatory interactions in the living cell and may provide highly specific recognition modules. We present Rosetta FlexPepDock ab-initio, a protocol for simultaneous docking and de-novo folding of peptides, starting from an approximate specification of the peptide binding site. Using the Rosetta fragments library and a coarse-grained structural representation of the peptide and the receptor, FlexPepDock ab-initio samples efficiently and simultaneously the space of possible peptide backbone conformations and rigid-body orientations over the receptor surface of a given binding site. The subsequent all-atom refinement of the coarse-grained models includes full side-chain modeling of both the receptor and the peptide, resulting in high-resolution models in which key side-chain interactions are recapitulated. The protocol was applied to a benchmark in which peptides were modeled over receptors in either their bound backbone conformations or in their free, unbound form. Near-native peptide conformations were identified in 18/26 of the bound cases and 7/14 of the unbound cases. The protocol performs well on peptides from various classes of secondary structures, including coiled peptides with unusual turns and kinks. The results presented here significantly extend the scope of state-of-the-art methods for high-resolution peptide modeling, which can now be applied to a wide variety of peptide-protein interactions where no prior information about the peptide backbone conformation is available, enabling detailed structure-based studies and manipulation of those interactions. © 2011 Raveh et al.

Rosetta FlexPepDock ab-initio: Simultaneous Folding, Docking and Refinement of Peptides onto Their Receptors

PubMed Central

Raveh, Barak; London, Nir; Zimmerman, Lior; Schueler-Furman, Ora

2011-01-01

Flexible peptides that fold upon binding to another protein molecule mediate a large number of regulatory interactions in the living cell and may provide highly specific recognition modules. We present Rosetta FlexPepDock ab-initio, a protocol for simultaneous docking and de-novo folding of peptides, starting from an approximate specification of the peptide binding site. Using the Rosetta fragments library and a coarse-grained structural representation of the peptide and the receptor, FlexPepDock ab-initio samples efficiently and simultaneously the space of possible peptide backbone conformations and rigid-body orientations over the receptor surface of a given binding site. The subsequent all-atom refinement of the coarse-grained models includes full side-chain modeling of both the receptor and the peptide, resulting in high-resolution models in which key side-chain interactions are recapitulated. The protocol was applied to a benchmark in which peptides were modeled over receptors in either their bound backbone conformations or in their free, unbound form. Near-native peptide conformations were identified in 18/26 of the bound cases and 7/14 of the unbound cases. The protocol performs well on peptides from various classes of secondary structures, including coiled peptides with unusual turns and kinks. The results presented here significantly extend the scope of state-of-the-art methods for high-resolution peptide modeling, which can now be applied to a wide variety of peptide-protein interactions where no prior information about the peptide backbone conformation is available, enabling detailed structure-based studies and manipulation of those interactions. PMID:21572516

A detached eddy simulation model for the study of lateral separation zones along a large canyon-bound river

NASA Astrophysics Data System (ADS)

Alvarez, Laura V.; Schmeeckle, Mark W.; Grams, Paul E.

2017-01-01

Lateral flow separation occurs in rivers where banks exhibit strong curvature. In canyon-bound rivers, lateral recirculation zones are the principal storage of fine-sediment deposits. A parallelized, three-dimensional, turbulence-resolving model was developed to study the flow structures along lateral separation zones located in two pools along the Colorado River in Marble Canyon. The model employs the detached eddy simulation (DES) technique, which resolves turbulence structures larger than the grid spacing in the interior of the flow. The DES-3D model is validated using Acoustic Doppler Current Profiler flow measurements taken during the 2008 controlled flood release from Glen Canyon Dam. A point-to-point validation using a number of skill metrics, often employed in hydrological research, is proposed here for fluvial modeling. The validation results show predictive capabilities of the DES model. The model reproduces the pattern and magnitude of the velocity in the lateral recirculation zone, including the size and position of the primary and secondary eddy cells, and return current. The lateral recirculation zone is open, having continuous import of fluid upstream of the point of reattachment and export by the recirculation return current downstream of the point of separation. Differences in magnitude and direction of near-bed and near-surface velocity vectors are found, resulting in an inward vertical spiral. Interaction between the recirculation return current and the main flow is dynamic, with large temporal changes in flow direction and magnitude. Turbulence structures with a predominately vertical axis of vorticity are observed in the shear layer becoming three-dimensional without preferred orientation downstream.

Ranking Enzyme Structures in the PDB by Bound Ligand Similarity to Biological Substrates.

PubMed

Tyzack, Jonathan D; Fernando, Laurent; Ribeiro, Antonio J M; Borkakoti, Neera; Thornton, Janet M

2018-04-03

There are numerous applications that use the structures of protein-ligand complexes from the PDB, such as 3D pharmacophore identification, virtual screening, and fragment-based drug design. The structures underlying these applications are potentially much more informative if they contain biologically relevant bound ligands, with high similarity to the cognate ligands. We present a study of ligand-enzyme complexes that compares the similarity of bound and cognate ligands, enabling the best matches to be identified. We calculate the molecular similarity scores using a method called PARITY (proportion of atoms residing in identical topology), which can conveniently be combined to give a similarity score for all cognate reactants or products in the reaction. Thus, we generate a rank-ordered list of related PDB structures, according to the biological similarity of the ligands bound in the structures. Copyright © 2018 The Authors. Published by Elsevier Ltd.. All rights reserved.

Structural mechanism of JH delivery in hemolymph by JHBP of silkworm, Bombyx mori

PubMed Central

Suzuki, Rintaro; Fujimoto, Zui; Shiotsuki, Takahiro; Tsuchiya, Wataru; Momma, Mitsuru; Tase, Akira; Miyazawa, Mitsuhiro; Yamazaki, Toshimasa

2011-01-01

Juvenile hormone (JH) plays crucial roles in many aspects of the insect life. All the JH actions are initiated by transport of JH in the hemolymph as a complex with JH-binding protein (JHBP) to target tissues. Here, we report structural mechanism of JH delivery by JHBP based upon the crystal and solution structures of apo and JH-bound JHBP. In solution, apo-JHBP exists in equilibrium of multiple conformations with different orientations of the gate helix for the hormone-binding pocket ranging from closed to open forms. JH-binding to the gate-open form results in the fully closed JHBP-JH complex structure where the bound JH is completely buried inside the protein. JH-bound JHBP opens the gate helix to release the bound hormone likely by sensing the less polar environment at the membrane surface of target cells. This is the first report that provides structural insight into JH signaling. PMID:22355650

Rice yellow stunt rhabdovirus protein 6 suppresses systemic RNA silencing by blocking RDR6-mediated secondary siRNA synthesis.

PubMed

Guo, Hongyan; Song, Xiaoguang; Xie, Chuanmiao; Huo, Yan; Zhang, Fujie; Chen, Xiaoying; Geng, Yunfeng; Fang, Rongxiang

2013-08-01

The P6 protein of Rice yellow stunt rhabdovirus (RYSV) is a virion structural protein that can be phosphorylated in vitro. However its exact function remains elusive. We found that P6 enhanced the virulence of Potato virus X (PVX) in Nicotiana benthamiana and N. tabacum plants, suggesting that it might function as a suppressor of RNA silencing. We examined the mechanism of P6-mediated silencing suppression by transiently expressing P6 in both N. benthamiana leaves and rice protoplasts. Our results showed that P6 could repress the production of secondary siRNAs and inhibit systemic green fluorescent protein RNA silencing but did not interfere with local RNA silencing in N. benthamiana plants or in rice protoplasts. Intriguingly, P6 and RDR6 had overlapping subcellular localization and P6 bound both rice and Arabidopsis RDR6 in vivo. Furthermore, transgenic rice plants expressing P6 showed enhanced susceptibility to infection by Rice stripe virus. Hence, we propose that P6 is part of the RYSV's counter-defense machinery against the plant RNA silencing system and plays a role mainly in affecting RDR6-mediated secondary siRNA synthesis. Our work provides a new perspective on how a plant-infecting nucleorhabdovirus may counteract host RNA silencing-mediated antiviral defense.

Revisiting and re-engineering the classical zinc finger peptide: consensus peptide-1 (CP-1).

PubMed

Besold, Angelique N; Widger, Leland R; Namuswe, Frances; Michalek, Jamie L; Michel, Sarah L J; Goldberg, David P

2016-04-01

Zinc plays key structural and catalytic roles in biology. Structural zinc sites are often referred to as zinc finger (ZF) sites, and the classical ZF contains a Cys2His2 motif that is involved in coordinating Zn(II). An optimized Cys2His2 ZF, named consensus peptide 1 (CP-1), was identified more than 20 years ago using a limited set of sequenced proteins. We have reexamined the CP-1 sequence, using our current, much larger database of sequenced proteins that have been identified from high-throughput sequencing methods, and found the sequence to be largely unchanged. The CCHH ligand set of CP-1 was then altered to a CAHH motif to impart hydrolytic activity. This ligand set mimics the His2Cys ligand set of peptide deformylase (PDF), a hydrolytically active M(II)-centered (M = Zn or Fe) protein. The resultant peptide [CP-1(CAHH)] was evaluated for its ability to coordinate Zn(II) and Co(II) ions, adopt secondary structure, and promote hydrolysis. CP-1(CAHH) was found to coordinate Co(II) and Zn(II) and a pentacoordinate geometry for Co(II)-CP-1(CAHH) was implicated from UV-vis data. This suggests a His2Cys(H2O)2 environment at the metal center. The Zn(II)-bound CP-1(CAHH) was shown to adopt partial secondary structure by 1-D (1)H NMR spectroscopy. Both Zn(II)-CP-1(CAHH) and Co(II)-CP-1(CAHH) show good hydrolytic activity toward the test substrate 4-nitrophenyl acetate, exhibiting faster rates than most active synthetic Zn(II) complexes.

Homogenization-based interval analysis for structural-acoustic problem involving periodical composites and multi-scale uncertain-but-bounded parameters.

PubMed

Chen, Ning; Yu, Dejie; Xia, Baizhan; Liu, Jian; Ma, Zhengdong

2017-04-01

This paper presents a homogenization-based interval analysis method for the prediction of coupled structural-acoustic systems involving periodical composites and multi-scale uncertain-but-bounded parameters. In the structural-acoustic system, the macro plate structure is assumed to be composed of a periodically uniform microstructure. The equivalent macro material properties of the microstructure are computed using the homogenization method. By integrating the first-order Taylor expansion interval analysis method with the homogenization-based finite element method, a homogenization-based interval finite element method (HIFEM) is developed to solve a periodical composite structural-acoustic system with multi-scale uncertain-but-bounded parameters. The corresponding formulations of the HIFEM are deduced. A subinterval technique is also introduced into the HIFEM for higher accuracy. Numerical examples of a hexahedral box and an automobile passenger compartment are given to demonstrate the efficiency of the presented method for a periodical composite structural-acoustic system with multi-scale uncertain-but-bounded parameters.

The 15-K neutron structure of saccharide-free concanavalin A.

PubMed

Blakeley, M P; Kalb, A J; Helliwell, J R; Myles, D A A

2004-11-23

The positions of the ordered hydrogen isotopes of a protein and its bound solvent can be determined by using neutron crystallography. Furthermore, by collecting neutron data at cryo temperatures, the dynamic disorder within a protein crystal is reduced, which may lead to improved definition of the nuclear density. It has proved possible to cryo-cool very large Con A protein crystals (>1.5 mm3) suitable for high-resolution neutron and x-ray structure analysis. We can thereby report the neutron crystal structure of the saccharide-free form of Con A and its bound water, including 167 intact D2O molecules and 60 oxygen atoms at 15 K to 2.5-A resolution, along with the 1.65-A x-ray structure of an identical crystal at 100 K. Comparison with the 293-K neutron structure shows that the bound water molecules are better ordered and have lower average B factors than those at room temperature. Overall, twice as many bound waters (as D2O) are identified at 15 K than at 293 K. We note that alteration of bound water orientations occurs between 293 and 15 K; such changes, as illustrated here with this example, could be important more generally in protein crystal structure analysis and ligand design. Methodologically, this successful neutron cryo protein structure refinement opens up categories of neutron protein crystallography, including freeze-trapped structures and cryo to room temperature comparisons.

Combining Sequential Extractions and X-ray Absorption Spectroscopy for Quantitative and Qualitative Zinc Speciation in Soil

NASA Astrophysics Data System (ADS)

Bauer, Tatiana; Minkina, Tatiana; Batukaev, Abdulmalik; Nevidomskaya, Dina; Burachevskaya, Marina; Tsitsuashvili, Viktoriya; Urazgildieva, Kamilya

2017-04-01

The combined use of X-ray absorption spectrometry and extractive fractionation is an effective approach for studying the interaction of metal ions with soil compounds and identifying the phases-carriers of metals in soil and their stable fixation. These studies were carried out using the technique of X-ray absorption spectroscopy and chemical extractive fractionation. In a model experiment the samples taken in Calcic Chernozem were artificially contaminated with higher portion of Zn(NO3)2 (2000 mg/kg). The metal were incubated in soil samples for 2 year. The samples of soil mineral and organic phases (calcite, kaolinite, bentonite, humic acids) were saturated with Zn2+ from a solution of nitrate salts of metal. The total content of Zn in soil and soil various phases was determined using the X-ray fluorescence method. Extended X-ray absorption fine structure (EXAFS) Zn was measured at the Structural Materials Science beamline of the Kurchatov Center for Synchrotron Radiation. Sequential fractionation of Zn in soil conducted by Tessier method (Tessier et al., 1979) which determining 5 fractions of metals in soil: exchangeable, bound to Fe-Mn oxide, bound to carbonate, bound to the organic matter, and bound to silicate (residual). This methodology has so far more than 4000 citations (Web of Science), which demonstrates the popularity of this approach. Much Zn compounds are contained in uncontaminated soils in stable primary and secondary silicates inherited from the parental rocks (67% of the total concentrations in all fractions), which is a regional trait of soils in the fore-Caucasian plain. Extracted fractionation of metal compounds in soil samples, artificially contaminated with Zn salts, indicates the priority holding of Zn2+ ions by silicates, carbonates and Fe-Mn oxides. The Zn content significantly increases in the exchangeable fraction. Atomic structure study of the soil various phases saturated with Zn2+ ion by using (XANES) X-ray absorption spectroscopy allowed the determination of mechanism of metal ions interaction with soil phases and the resulting types of chemical bonds. Interaction with soil components modifies the electron structure of the metal ions themselves. The soil contamination with Zn is accompanied by decreasing the stable connection between metal and soil components. Interacting with humic acids in chernozem, the Zn2+ ion is coordinated by functional groups and ligands and forms unstable outer-sphere complexes. Zinc included into octahedral structures of layered minerals and hydro(oxides) can be inner-and outer-sphere adsorbed. The Zn2+ ions enable to replace Ca2+ ions in octahedral positions being coordinated with carbonate ions as ligands, thus forming absorbed complexes at the surface of mineral calcite. This work was supported by grant of the Russian Scientific Foundation № 16-14-10217.

SP.ACE: taking secondary school students' hearts and minds "up, up and away"

NASA Astrophysics Data System (ADS)

de Schrijver, Erik

2005-08-01

Secondary school students were given the opportunity to build and fly "pongsats" (small experiments weighing under 100 grams each, and packed inside a ping pong ball) on high-altitude balloons bound for 100000 feet, or 30 km: the edge of space. The need to acquire the knowledge and know-how to build successful experiments gave birth to the SP.ACE project. Over their last 3 years of secondary education, students are now learning about flight vehicles, the physical conditions in space, microcontrollers, sensors, programming, data logging, flight path analysis, and much more.

Structural redesign of lipase B from Candida antarctica by circular permutation and incremental truncation.

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

Qian, Zhen; Horton, John R.; Cheng, Xiadong

2009-11-02

Circular permutation of Candida antarctica lipase B yields several enzyme variants with substantially increased catalytic activity. To better understand the structural and functional consequences of protein termini reorganization, we have applied protein engineering and x-ray crystallography to cp283, one of the most active hydrolase variants. Our initial investigation has focused on the role of an extended surface loop, created by linking the native N- and C-termini, on protein integrity. Incremental truncation of the loop partially compensates for observed losses in secondary structure and the permutants temperature of unfolding. Unexpectedly, the improvements are accompanied by quaternary-structure changes from monomer to dimer.more » The crystal structures of one truncated variant (cp283{Delta}7) in the apo-form determined at 1.49 {angstrom} resolution and with a bound phosphonate inhibitor at 1.69 {angstrom} resolution confirmed the formation of a homodimer by swapping of the enzyme's 35-residue N-terminal region. Separately, the new protein termini at amino acid positions 282/283 convert the narrow access tunnel to the catalytic triad into a broad crevice for accelerated substrate entry and product exit while preserving the native active-site topology for optimal catalytic turnover.« less

Structures of Substrate-And Inhibitor-Bound Adenosine Deaminase From a Human Malaria Parasite Show a Dramatic Conformational Change And Shed Light on Drug Selectivity

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

Larson, E.T.; Deng, W.; Krumm, B.E.

Plasmodium and other apicomplexan parasites are deficient in purine biosynthesis, relying instead on the salvage of purines from their host environment. Therefore, interference with the purine salvage pathway is an attractive therapeutic target. The plasmodial enzyme adenosine deaminase (ADA) plays a central role in purine salvage and, unlike mammalian ADA homologs, has a further secondary role in methylthiopurine recycling. For this reason, plasmodial ADA accepts a wider range of substrates, as it is responsible for deamination of both adenosine and 5{prime}-methylthioadenosine. The latter substrate is not accepted by mammalian ADA homologs. The structural basis for this natural difference in specificitymore » between plasmodial and mammalian ADA has not been well understood. We now report crystal structures of Plasmodium vivax ADA in complex with adenosine, guanosine, and the picomolar inhibitor 2{prime}-deoxycoformycin. These structures highlight a drastic conformational change in plasmodial ADA upon substrate binding that has not been observed for mammalian ADA enzymes. Further, these complexes illuminate the structural basis for the differential substrate specificity and potential drug selectivity between mammalian and parasite enzymes.« less

Absence of residual structure in the intrinsically disordered regulatory protein CP12 in its reduced state

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

Launay, Hélène; Barré, Patrick; Puppo, Carine

2016-08-12

The redox switch protein CP12 is a key player of the regulation of the Benson–Calvin cycle. Its oxidation state is controlled by the formation/dissociation of two intramolecular disulphide bridges during the day/night cycle. CP12 was known to be globally intrinsically disordered on a large scale in its reduced state, while being partly ordered in the oxidised state. By combining Nuclear Magnetic Resonance and Small Angle X-ray Scattering experiments, we showed that, contrary to secondary structure or disorder predictions, reduced CP12 is fully disordered, with no transient or local residual structure likely to be precursor of the structures identified in themore » oxidised active state and/or in the bound state with GAPDH or PRK. These results highlight the diversity of the mechanisms of regulation of conditionally disordered redox switches, and question the stability of oxidised CP12 scaffold. - Highlights: • CP12 is predicted to form two helices in its N-terminal sequence. • Reduced CP12 is disordered as a random coil according to SAXS. • Limited or no transient structures are observed in reduced CP12 by NMR.« less

Contribution of proline to the pre-structuring tendency of transient helical secondary structure elements in intrinsically disordered proteins.

PubMed

Lee, Chewook; Kalmar, Lajos; Xue, Bin; Tompa, Peter; Daughdrill, Gary W; Uversky, Vladimir N; Han, Kyou-Hoon

2014-03-01

IDPs function without relying on three-dimensional structures. No clear rationale for such a behavior is available yet. PreSMos are transient secondary structures observed in the target-free IDPs and serve as the target-binding "active" motifs in IDPs. Prolines are frequently found in the flanking regions of PreSMos. Contribution of prolines to the conformational stability of the helical PreSMos in IDPs is investigated. MD simulations are performed for several IDP segments containing a helical PreSMo and the flanking prolines. To measure the influence of flanking-prolines on the structural content of a helical PreSMo calculations were done for wild type as well as for mutant segments with Pro→Asp, His, Lys, or Ala. The change in the helicity due to removal of a proline was measured both for the PreSMo region and for the flanking regions. The α-helical content in ~70% of the helical PreSMos at the early stage of simulation decreases due to replacement of an N-terminal flanking proline by other residues whereas the helix content in nearly all PreSMos increases when the same replacements occur at the C-terminal flanking region. The helix destabilizing/terminating role of the C-terminal flanking prolines is more pronounced than the helix promoting effect of the N-terminal flanking prolines. This work represents a novel example demonstrating that a proline is encoded in an IDP with a defined purpose. The helical PreSMos presage their target-bound conformations. As they most likely mediate IDP-target binding via conformational selection their helical content can be an important feature for IDP function. Copyright © 2013 Elsevier B.V. All rights reserved.

Effects of Water on the Single-Chain Elasticity of Poly(U) RNA.

PubMed

Luo, Zhonglong; Cheng, Bo; Cui, Shuxun

2015-06-09

Water, the dominant component under the physiological condition, is a complicated solvent which greatly affects the properties of solute molecules. Here, we utilize atomic force microscope-based single-molecule force spectroscopy to study the influence of water on the single-molecule elasticity of an unstructured single-stranded RNA (poly(U)). In nonpolar solvents, RNA presents its inherent elasticity, which is consistent with the theoretical single-chain elasticity calculated by quantum mechanics calculations. In aqueous buffers, however, an additional energy of 1.88 kJ/mol·base is needed for the stretching of the ssRNA chain. This energy is consumed by the bound water rearrangement (Ew) during chain elongation. Further experimental results indicate that the Ew value is uncorrelated to the salt concentrations and stretching velocity. The results obtained in an 8 M guanidine·HCl solution provide more evidence that the bound water molecules around RNA give rise to the observed deviation between aqueous and nonaqueous environments. Compared to synthetic water-soluble polymers, the value of Ew of RNA is much lower. The weak interference of water is supposed to be the precondition for the RNA secondary structure to exist in aqueous solution.

Spectrum sharing between a surveillance radar and secondary Wi-Fi networks

NASA Astrophysics Data System (ADS)

Hessar, Farzad; Roy, Sumit

2016-06-01

Co-existence between unlicensed networks that share spectrum spatio-temporally with terrestrial (e.g. Air Traffic Control) and shipborne radars in 3-GHz band is attracting significant interest. Similar to every primary-secondary coexistence scenario, interference from unlicensed devices to a primary receiver must be within acceptable bounds. In this work, we formulate the spectrum sharing problem between a pulsed, search radar (primary) and 802.11 WLAN as the secondary. We compute the protection region for such a search radar for a) a single secondary user (initially) as well as b) a random spatial distribution of multiple secondary users. Furthermore, we also analyze the interference to the WiFi devices from the radar's transmissions to estimate the impact on achievable WLAN throughput as a function of distance to the primary radar.

Structure of the Tetrameric p53 Tumor Suppressor Bound to DNA

DTIC Science & Technology

2002-05-01

been unable to prepare suitable p53/DNA cocrystals for structure determination. Nonetheless, we have successfully determined the medium resolution (2.7A... cocrystallization with longer DNA targets or DNA targets assembled into nucleosome core particles. The structure of tetrameric p53 bound to DNA will provide

Structure Functions of Bound Neutrons

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

Sebastian Kuhn

2005-04-01

We describe an experiment measuring electron scattering on a neutron bound in deuterium with coincident detection of a fast, backward-going spectator proton. Our data map out the relative importance of the pure PWIA spectator mechanism and final state interactions in various kinematic regions, and give a first glimpse of the modification of the structure function of a bound neutron as a function of its off-shell mass. We also discuss a new experimental program to study the structure of a free neutron by extending the same technique to much lower spectator momenta.

Influence of the Relative Enamel Abrasivity (REA) of Toothpastes on the Uptake of KOH-soluble and Structurally Bound Fluoride.

PubMed

Elmazi, Valbona; Sener, Beatrice; Attin, Thomas; Imfeld, Thomas; Wegehaupt, Florian J

2015-01-01

To determine the influence of the relative enamel abrasivity (REA) of fluoridated toothpaste on the uptake of KOH-soluble and structurally bound fluoride into enamel. Bovine enamel samples were randomly allocated to 6 groups (n=36 per group). Groups A to C were treated with sodium fluoride (NaF) toothpastes and groups D to F with amine fluoride (AmF) toothpastes (1500 ppm F each). The REA in groups A and D was 2, in groups B and E it was 6 and in groups C and F it was 9. Twice a day, 18 samples of each group were immersed for 2 min in a slurry (toothpaste:artificial saliva=1:3), while the remaining samples were brushed with the respective slurry (2.5 N force; 60 strokes/min; 2 min). All samples were stored at 37°C and 100% humidity. After five days, the amount of KOH-soluble and structurally bound fluoride was determined and statistically compared by Scheffe's post-hoc tests. REA value and mode of application (immersion or brushing) had no significant influence on the amount of either kind of fluoride from NaF toothpastes. Only for the NaF toothpaste with REA 6 was the amount of KOH-soluble fluoride significantly higher after brushing. With AmF toothpastes, KOH-soluble and structurally bound fluoride concentrations were significantly higher when the samples were brushed. Furthermore, in the REA-2 group, the amounts of KOH-soluble fluoride (brushed or immersed) and structurally bound fluoride (brushed) were significantly higher than in the other groups. The REA dependency of KOH-soluble and structurally bound fluoride was found only for the AmF toothpastes. Using AmF toothpaste, the mode of application influenced the uptake of KOH-soluble and structurally bound fluoride into enamel.

Identification of New Serum Biomarkers for Early Breast Cancer Diagnosis and Prognosis Using Lipid Microarrays

DTIC Science & Technology

2008-09-01

specific for asialo-GM1 bound specifically to GM1, but not to the closely related gangliosides GM1 or GM2 (Fig. 2). Monoclonal antibodies raised...against GD3 specifically bound GD3, but not to asialo-GM1, GM1 and GM2 (Fig. 2). The secondary antibodies did not show reactivity against lipids (data not...fluorescent intensity on different membranes. Asialo GM1 GM1 GM2 GD3 Fig 2. Lipids on the PVDF membrane can be detected by specific

Identification of New Serum Biomarkers for Early Breast Cancer Diagnosis and Prognosis Using Lipid Microarrays

DTIC Science & Technology

2007-09-01

specific for asialo-GM1 bound specifically to GM1, but not to the closely related gangliosides GM1 or GM2 (Fig. 2). Monoclonal antibodies raised against...GD3 specifically bound GD3, but not to asialo-GM1, GM1 and GM2 (Fig. 2). The secondary antibodies did not show reactivity against lipids (data not...fluorescent intensity on different membranes. Asialo GM1 GM1 GM2 GD3 Fig 2. Lipids on the PVDF membrane can be detected by specific antibodies

A de novo redesign of the WW domain

PubMed Central

Kraemer-Pecore, Christina M.; Lecomte, Juliette T.J.; Desjarlais, John R.

2003-01-01

We have used a sequence prediction algorithm and a novel sampling method to design protein sequences for the WW domain, a small β-sheet motif. The procedure, referred to as SPANS, designs sequences to be compatible with an ensemble of closely related polypeptide backbones, mimicking the inherent flexibility of proteins. Two designed sequences (termed SPANS-WW1 and SPANS-WW2), using only naturally occurring l-amino acids, were selected for study and the corresponding polypeptides were prepared in Escherichia coli. Circular dichroism data suggested that both purified polypeptides adopted secondary structure features related to those of the target without the aid of disulfide bridges or bound cofactors. The structure exhibited by SPANS-WW2 melted cooperatively by raising the temperature of the solution. Further analysis of this polypeptide by proton nuclear magnetic resonance spectroscopy demonstrated that at 5°C, it folds into a structure closely resembling a natural WW domain. This achievement constitutes one of a small number of successful de novo protein designs through fully automated computational methods and highlights the feasibility of including backbone flexibility in the design strategy. PMID:14500877

A de novo redesign of the WW domain.

PubMed

Kraemer-Pecore, Christina M; Lecomte, Juliette T J; Desjarlais, John R

2003-10-01

We have used a sequence prediction algorithm and a novel sampling method to design protein sequences for the WW domain, a small beta-sheet motif. The procedure, referred to as SPANS, designs sequences to be compatible with an ensemble of closely related polypeptide backbones, mimicking the inherent flexibility of proteins. Two designed sequences (termed SPANS-WW1 and SPANS-WW2), using only naturally occurring L-amino acids, were selected for study and the corresponding polypeptides were prepared in Escherichia coli. Circular dichroism data suggested that both purified polypeptides adopted secondary structure features related to those of the target without the aid of disulfide bridges or bound cofactors. The structure exhibited by SPANS-WW2 melted cooperatively by raising the temperature of the solution. Further analysis of this polypeptide by proton nuclear magnetic resonance spectroscopy demonstrated that at 5 degrees C, it folds into a structure closely resembling a natural WW domain. This achievement constitutes one of a small number of successful de novo protein designs through fully automated computational methods and highlights the feasibility of including backbone flexibility in the design strategy.

Low-velocity impact tests on fibrous composite sandwich structures

NASA Technical Reports Server (NTRS)

Sharma, A. V.

1981-01-01

The effect of low-velocity projectile impact on the load-carrying ability of the composite sandwich structural components is investigated experimentally, the impact simulating the damage caused by runway debris and the accidental dropping of hand tools during servicing on secondary aircraft structures made with composites. The sandwich-type beam specimens were fabricated with graphite/epoxy face sheets, aluminum honeycomb core, and a steel (back) plate. A four-point beam-loading apparatus was used, and the ultimate strength, ultimate strain, and residual strength of the composites were determined. A faired curve is presented indicating the lower bound of the failure threshold for each of the laminate configurations tested in compression and tension as a function of the projectile impact energy. It is shown that strength degradation due to impact is dependent on the laminate configuration and the fiber/matrix combination. The laminates having more angle plies near the impact surface and unidirectional plies elsewhere seem to show extensive interply and intraply fiber delaminations at failure relative to the laminates with a cross-ply on the impact surface.

VC-dimension of univariate decision trees.

PubMed

Yildiz, Olcay Taner

2015-02-01

In this paper, we give and prove the lower bounds of the Vapnik-Chervonenkis (VC)-dimension of the univariate decision tree hypothesis class. The VC-dimension of the univariate decision tree depends on the VC-dimension values of its subtrees and the number of inputs. Via a search algorithm that calculates the VC-dimension of univariate decision trees exhaustively, we show that our VC-dimension bounds are tight for simple trees. To verify that the VC-dimension bounds are useful, we also use them to get VC-generalization bounds for complexity control using structural risk minimization in decision trees, i.e., pruning. Our simulation results show that structural risk minimization pruning using the VC-dimension bounds finds trees that are more accurate as those pruned using cross validation.

Characterization of the binding of 2-mercaptobenzimidazole to bovine serum albumin.

PubMed

Teng, Yue; Zou, Luyi; Huang, Ming; Zong, Wansong

2015-04-01

2-Mercaptobenzimidazole (MBI) is widely utilized as a corrosion inhibitor, copper-plating brightener and rubber accelerator. The residue of MBI in the environment is potentially harmful to human health. In this article, the interaction of MBI with bovine serum albumin (BSA) was explored using spectroscopic and molecular docking methods under physiological conditions. The positively charged MBI can spontaneously bind with the negatively charged BSA through electrostatic forces with one binding site. The site marker competition experiments and the molecular docking study revealed that MBI bound into site II (subdomain IIIA) of BSA, which further led to some secondary structure and microenvironmental changes of BSA. This work provides useful information on understanding the toxicological actions of MBI at the molecular level. Copyright © 2015 John Wiley & Sons, Ltd.

Biochemical identification of Argonaute 2 as the sole protein required for RNA-induced silencing complex activity

PubMed Central

Rand, Tim A.; Ginalski, Krzysztof; Grishin, Nick V.; Wang, Xiaodong

2004-01-01

RNA interference is carried out by the small double-stranded RNA-induced silencing complex (RISC). The RISC-bound small RNA guides the RISC complex to identify and cleave mRNAs with complementary sequences. The proteins that make up the RISC complex and cleave mRNA have not been unequivocally defined. Here, we report the biochemical purification of RISC activity to homogeneity from Drosophila Schnieder 2 cell extracts. Argonaute 2 (Ago-2) is the sole protein component present in the purified, functional RISC. By using a bioinformatics method that combines sequence-profile analysis with predicted protein secondary structure, we found homology between the PIWI domain of Ago-2 and endonuclease V and identified potential active-site amino acid residues within the PIWI domain of Ago-2. PMID:15452342

Biochemical identification of Argonaute 2 as the sole protein required for RNA-induced silencing complex activity.

PubMed

Rand, Tim A; Ginalski, Krzysztof; Grishin, Nick V; Wang, Xiaodong

2004-10-05

RNA interference is carried out by the small double-stranded RNA-induced silencing complex (RISC). The RISC-bound small RNA guides the RISC complex to identify and cleave mRNAs with complementary sequences. The proteins that make up the RISC complex and cleave mRNA have not been unequivocally defined. Here, we report the biochemical purification of RISC activity to homogeneity from Drosophila Schnieder 2 cell extracts. Argonaute 2 (Ago-2) is the sole protein component present in the purified, functional RISC. By using a bioinformatics method that combines sequence-profile analysis with predicted protein secondary structure, we found homology between the PIWI domain of Ago-2 and endonuclease V and identified potential active-site amino acid residues within the PIWI domain of Ago-2.

Molecular simulations and Markov state modeling reveal the structural diversity and dynamics of a theophylline-binding RNA aptamer in its unbound state

PubMed Central

Warfield, Becka M.

2017-01-01

RNA aptamers are oligonucleotides that bind with high specificity and affinity to target ligands. In the absence of bound ligand, secondary structures of RNA aptamers are generally stable, but single-stranded and loop regions, including ligand binding sites, lack defined structures and exist as ensembles of conformations. For example, the well-characterized theophylline-binding aptamer forms a highly stable binding site when bound to theophylline, but the binding site is unstable and disordered when theophylline is absent. Experimental methods have not revealed at atomic resolution the conformations that the theophylline aptamer explores in its unbound state. Consequently, in the present study we applied 21 microseconds of molecular dynamics simulations to structurally characterize the ensemble of conformations that the aptamer adopts in the absence of theophylline. Moreover, we apply Markov state modeling to predict the kinetics of transitions between unbound conformational states. Our simulation results agree with experimental observations that the theophylline binding site is found in many distinct binding-incompetent states and show that these states lack a binding pocket that can accommodate theophylline. The binding-incompetent states interconvert with binding-competent states through structural rearrangement of the binding site on the nanosecond to microsecond timescale. Moreover, we have simulated the complete theophylline binding pathway. Our binding simulations supplement prior experimental observations of slow theophylline binding kinetics by showing that the binding site must undergo a large conformational rearrangement after the aptamer and theophylline form an initial complex, most notably, a major rearrangement of the C27 base from a buried to solvent-exposed orientation. Theophylline appears to bind by a combination of conformational selection and induced fit mechanisms. Finally, our modeling indicates that when Mg2+ ions are present the population of binding-competent aptamer states increases more than twofold. This population change, rather than direct interactions between Mg2+ and theophylline, accounts for altered theophylline binding kinetics. PMID:28437473

Isolation and characterization of the genes for two small RNAs of herpesvirus papio and their comparison with Epstein-Barr virus-encoded EBER RNAs.

PubMed Central

Howe, J G; Shu, M D

1988-01-01

Genes for the Epstein-Barr virus-encoded RNAs (EBERs), two low-molecular-weight RNAs encoded by the human gammaherpesvirus Epstein-Barr virus (EBV), hybridize to two small RNAs in a baboon cell line that contains a similar virus, herpesvirus papio (HVP). The genes for the HVP RNAs (HVP-1 and HVP-2) are located together in the small unique region at the left end of the viral genome and are transcribed by RNA polymerase III in a rightward direction, similar to the EBERs. There is significant similarity between EBER1 and HVP-1 RNA, except for an insert of 22 nucleotides which increases the length of HVP-1 RNA to 190 nucleotides. There is less similarity between the sequences of EBER2 and HVP-2 RNA, but both have a length of about 170 nucleotides. The predicted secondary structure of each HVP RNA is remarkably similar to that of the respective EBER, implying that the secondary structures are important for function. Upstream from the initiation sites of all four RNA genes are several highly conserved sequences which may function in the regulation of transcription. The HVP RNAs, together with the EBERs, are highly abundant in transformed cells and are efficiently bound by the cellular La protein. Images PMID:2839701

Social network influences on smoking, drinking and drug use in secondary school: centrifugal and centripetal forces.

PubMed

Fletcher, Adam; Bonell, Chris

2013-06-01

We explore how school experiences and social networks structure young people's substance use in different institutional contexts. The concepts of 'selection' and 'influence' are situated within the context of bounded agency, counter-school cultures and Bourdieusian notions of capital. We employed individual and group interviews, network-mapping, and observations at two contrasting English secondary schools. Both schools were characterised by extended social network structures that appeared to influence patterns of substance use, although the mechanisms via which this occurred varied according to school context. At Grange House school (suburban context) a minority of students from disadvantaged families were alienated by the attainment-focused regime, marginalised by a strong peer-led centrifugal force pushing them outwards, and substance use was an alternative source of bonding and identity for these students. In contrast, at North Street a centripetal force operated whereby the majority of students were pulled towards highly-visible, normative markers of 'safe', 'road culture', such as cannabis use and gang-involvement, as they attempted to fit in and survive in an inner-city school environment. We conclude that health inequalities may be reproduced through these distinctive centrifugal and centripetal forces in different institutional contexts, and this should be the focus of quantitative examination in the UK and elsewhere. © 2013 The Authors. Sociology of Health & Illness © 2013 Foundation for the Sociology of Health & Illness/John Wiley & Sons Ltd.

Mapping the Free Energy Landscape of PKA Inhibition and Activation: A Double-Conformational Selection Model for the Tandem cAMP-Binding Domains of PKA RIα

PubMed Central

Akimoto, Madoka; McNicholl, Eric Tyler; Ramkissoon, Avinash; Moleschi, Kody; Taylor, Susan S.; Melacini, Giuseppe

2015-01-01

Protein Kinase A (PKA) is the major receptor for the cyclic adenosine monophosphate (cAMP) secondary messenger in eukaryotes. cAMP binds to two tandem cAMP-binding domains (CBD-A and -B) within the regulatory subunit of PKA (R), unleashing the activity of the catalytic subunit (C). While CBD-A in RIα is required for PKA inhibition and activation, CBD-B functions as a “gatekeeper” domain that modulates the control exerted by CBD-A. Preliminary evidence suggests that CBD-B dynamics are critical for its gatekeeper function. To test this hypothesis, here we investigate by Nuclear Magnetic Resonance (NMR) the two-domain construct RIα (91–379) in its apo, cAMP2, and C-bound forms. Our comparative NMR analyses lead to a double conformational selection model in which each apo CBD dynamically samples both active and inactive states independently of the adjacent CBD within a nearly degenerate free energy landscape. Such degeneracy is critical to explain the sensitivity of CBD-B to weak interactions with C and its high affinity for cAMP. Binding of cAMP eliminates this degeneracy, as it selectively stabilizes the active conformation within each CBD and inter-CBD contacts, which require both cAMP and W260. The latter is contributed by CBD-B and mediates capping of the cAMP bound to CBD-A. The inter-CBD interface is dispensable for intra-CBD conformational selection, but is indispensable for full activation of PKA as it occludes C-subunit recognition sites within CBD-A. In addition, the two structurally homologous cAMP-bound CBDs exhibit marked differences in their residual dynamics profiles, supporting the notion that conservation of structure does not necessarily imply conservation of dynamics. PMID:26618408

A three-dimensional construction of the active site (region 507-749) of human neutral endopeptidase (EC.3.4.24.11).

PubMed

Tiraboschi, G; Jullian, N; Thery, V; Antonczak, S; Fournie-Zaluski, M C; Roques, B P

1999-02-01

A three-dimensional model of the 507-749 region of neutral endopeptidase-24.11 (NEP; E.C.3.4.24.11) was constructed integrating the results of secondary structure predictions and sequence homologies with the bacterial endopeptidase thermolysin. Additional data were extracted from the structure of two other metalloproteases, astacin and stromelysin. The resulting model accounts for the main biological properties of NEP and has been used to describe the environment close to the zinc atom defining the catalytic site. The analysis of several thiol inhibitors, complexed in the model active site, revealed the presence of a large hydrophobic pocket at the S1' subsite level. This is supported by the nature of the constitutive amino acids. The computed energies of bound inhibitors correspond with the relative affinities of the stereoisomers of benzofused macrocycle derivatives of thiorphan. The model could be used to facilitate the design of new NEP inhibitors, as illustrated in the paper.

Immunoinformatic Analysis of Crimean Congo Hemorrhagic Fever Virus Glycoproteins and Epitope Prediction for Synthetic Peptide Vaccine.

PubMed

Tipu, Hamid Nawaz

2016-02-01

To determine the Crimean Congo Hemorrhagic Fever (CCHF) virus M segement glycoprotein's immunoinformatic parameters, and identify Human Leukocyte Antigen (HLA) class I binders as candidates for synthetic peptide vaccines. Cross-sectional study. Combined Military Hospital, Khuzdar Cantt, in May 2015. Data acquisition, antigenicity prediction, secondary and tertiary structure prediction, residue analysis were done using immunoinformatics tools. HLAclass I binders in glycoprotein's sequence were identified at nanomer length using NetMHC 3.4 and mapped onto tertiary structure. Docking was done for strongest binder against its corresponding allele with CABS-dock. HLAA*0101, 0201, 0301, 2402, 2601 and B*0702, 0801, 2705, 3901, 4001, 5801, 1501 were analyzed against two glycoprotein components of the virus. Atotal of 35 nanomers from GP1, and 3 from GP2 were identified. HLAB*0702 bound maximum number of peptides (6), while HLAB*4001 showed strongest binding affinity. HLAspecific glycoproteins epitope prediction can help identify synthetic peptide vaccine candidates.

Binding of (-)-epigallocatechin-3-gallate with thermally-induced bovine serum albumin/ι-carrageenan particles.

PubMed

Li, Jinbing; Wang, Xiaoyong

2015-02-01

Novel thermally-induced BSA/ι-carrageenan particles are used as a protective carrier for (-)-epigallocatechin-3-gallate (EGCG). The addition of EGCG to BSA/ι-carrageenan particles can highly quench the intrinsic fluorescence of BSA, which is explained in terms of the binding of EGCG to the hydrophobic pockets of BSA mainly through the hydrophobic force. According to the double logarithm equation, the binding constant is determined as 1.1×10(8)M(-1) for the binding of EGCG with BSA/ι-carrageenan particles. The high binding affinity is ascribed to both the molecular structure of EGCG and the partial unfolding state of BSA in BSA/ι-carrageenan particles. The circular dichroism spectra and calculated α-helix of BSA suggest that the bound EGCG leads to a more random secondary structure of BSA. Furthermore, BSA/ι-carrageenan particles are found to be superior to native BSA and pure BSA particles for improving the stability and radical scavenging activity of EGCG. Copyright © 2014 Elsevier Ltd. All rights reserved.

Molecular dynamics simulations of lysozyme-lipid systems: probing the early steps of protein aggregation.

PubMed

Trusova, Valeriya M; Gorbenko, Galyna P

2017-07-10

Using the molecular dynamics simulation, the role of lipids in the lysozyme transition into the aggregation-competent conformation has been clarified. Analysis of the changes of lysozyme secondary structure upon its interactions with the model bilayer membranes composed of phosphatidylcholine and its mixtures with phosphatidylglycerol (10, 40, and 80 mol%) within the time interval of 100 ns showed that lipid-bound protein is characterized by the increased content of β-structures. Along with this, the formation of protein-lipid complexes was accompanied by the increase in the gyration radius and the decrease in RMSD of polypeptide chain. The results obtained were interpreted in terms of the partial unfolding of lysozyme molecule on the lipid matrix, with the magnitude of this effect being increased with increasing the fraction of anionic lipids. Based on the results of molecular dynamics simulation, a hypothetical model of the nucleation of lysozyme amyloid fibrils in a membrane environment was suggested.

Crystal structures of the methyltransferase and helicase from the ZIKA 1947 MR766 Uganda strain

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

Bukrejewska, Malgorzata; Derewenda, Urszula; Radwanska, Malwina

2017-08-15

Two nonstructural proteins encoded byZika virusstrain MR766 RNA, a methyltransferase and a helicase, were crystallized and their structures were solved and refined at 2.10 and 2.01 Å resolution, respectively. The NS5 methyltransferase contains a boundS-adenosyl-L-methionine (SAM) co-substrate. The NS3 helicase is in the apo form. Comparison with published crystal structures of the helicase in the apo, nucleotide-bound and single-stranded RNA (ssRNA)-bound states suggests that binding of ssRNA to the helicase may occur through conformational selection rather than induced fit.

New insights into flavivirus biology: the influence of pH over interactions between prM and E proteins

NASA Astrophysics Data System (ADS)

Oliveira, Edson R. A.; de Alencastro, Ricardo B.; Horta, Bruno A. C.

2017-11-01

Diseases caused by flaviviruses, such as dengue and zika, are globally recognized as major threats. During infection, a critical point in their replicative cycle is the maturation step, which occurs throughout the cellular exocytic pathway. This step is a pH-dependent process that involves the modification of the viral envelope by converting prM (pre-membrane) into M (membrane) proteins with the release of a "pr peptide". After this reaction, the pr peptides remain bound to the viral envelope while the virions cross the acidic trans-Golgi network, and are released only at neutral pH after secretion of the virus particles. Despite this current knowledge, the molecular basis of the flavivirus maturation step is largely unknown. Here, based on the crystal structure of the dengue pr-E complex ("pr peptide" bound to virus envelope protein) and using molecular dynamics simulations, we found that the pH shift from acidic to neutral yields considerable structural changes in the system. Dynamic cross correlation maps and root mean square deviation analyses revealed that the pr-E junction is clearly unstable under neutral pH. Secondary structure analysis also revealed that the fusion loop region, present in the E protein, is sensitive to pH and tends to unstructure at a neutral environment. Moreover, we found that five residues present in the E protein, Gly102, His244, Thr70, Thr68 and Asn67 are critical to confer stability to the pr-E complex while inside the Golgi apparatus. This work brings details about the dynamical behavior of the pr-E system, helps to better understand the flavivirus biology and may also be of use in the development of novel antiviral strategies.

Structural Studies on Intact Clostridium botulinum Neurotoxins Complexed with Inhibitors Leading to Drug Design

DTIC Science & Technology

2007-02-01

699. 5. Hanson, M. A., and Stevens, R. C. (2000) Cocrystal structure of synaptobrevin-ll bound to botulinum neurotoxin type B at 2.0 A resolution...R. C. (2000). Cocrystal structure of synaptobrevin-II bound to botulinum neurotoxin type B at 2.0 A resolution. Nature Struct. Biol. 7, 687–692. 23

Leveraging structure determination with fragment screening for infectious disease drug targets: MECP synthase from Burkholderia pseudomallei

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

Begley, Darren W.; Hartley, Robert C.; Davies, Douglas R.

As part of the Seattle Structural Genomics Center for Infectious Disease, we seek to enhance structural genomics with ligand-bound structure data which can serve as a blueprint for structure-based drug design. We have adapted fragment-based screening methods to our structural genomics pipeline to generate multiple ligand-bound structures of high priority drug targets from pathogenic organisms. In this study, we report fragment screening methods and structure determination results for 2C-methyl-D-erythritol-2,4-cyclo-diphosphate (MECP) synthase from Burkholderia pseudomallei, the gram-negative bacterium which causes melioidosis. Screening by nuclear magnetic resonance spectroscopy as well as crystal soaking followed by X-ray diffraction led to the identification ofmore » several small molecules which bind this enzyme in a critical metabolic pathway. A series of complex structures obtained with screening hits reveal distinct binding pockets and a range of small molecules which form complexes with the target. Additional soaks with these compounds further demonstrate a subset of fragments to only bind the protein when present in specific combinations. This ensemble of fragment-bound complexes illuminates several characteristics of MECP synthase, including a previously unknown binding surface external to the catalytic active site. These ligand-bound structures now serve to guide medicinal chemists and structural biologists in rational design of novel inhibitors for this enzyme.« less

A semi-supervised learning approach for RNA secondary structure prediction.

PubMed

Yonemoto, Haruka; Asai, Kiyoshi; Hamada, Michiaki

2015-08-01

RNA secondary structure prediction is a key technology in RNA bioinformatics. Most algorithms for RNA secondary structure prediction use probabilistic models, in which the model parameters are trained with reliable RNA secondary structures. Because of the difficulty of determining RNA secondary structures by experimental procedures, such as NMR or X-ray crystal structural analyses, there are still many RNA sequences that could be useful for training whose secondary structures have not been experimentally determined. In this paper, we introduce a novel semi-supervised learning approach for training parameters in a probabilistic model of RNA secondary structures in which we employ not only RNA sequences with annotated secondary structures but also ones with unknown secondary structures. Our model is based on a hybrid of generative (stochastic context-free grammars) and discriminative models (conditional random fields) that has been successfully applied to natural language processing. Computational experiments indicate that the accuracy of secondary structure prediction is improved by incorporating RNA sequences with unknown secondary structures into training. To our knowledge, this is the first study of a semi-supervised learning approach for RNA secondary structure prediction. This technique will be useful when the number of reliable structures is limited. Copyright © 2015 Elsevier Ltd. All rights reserved.

Cell-bound lipases from Burkholderia sp. ZYB002: gene sequence analysis, expression, enzymatic characterization, and 3D structural model.

PubMed

Shu, Zhengyu; Lin, Hong; Shi, Shaolei; Mu, Xiangduo; Liu, Yanru; Huang, Jianzhong

2016-05-03

The whole-cell lipase from Burkholderia cepacia has been used as a biocatalyst in organic synthesis. However, there is no report in the literature on the component or the gene sequence of the cell-bound lipase from this species. Qualitative analysis of the cell-bound lipase would help to illuminate the regulation mechanism of gene expression and further improve the yield of the cell-bound lipase by gene engineering. Three predictive cell-bound lipases, lipA, lipC21 and lipC24, from Burkholderia sp. ZYB002 were cloned and expressed in E. coli. Both LipA and LipC24 displayed the lipase activity. LipC24 was a novel mesophilic enzyme and displayed preference for medium-chain-length acyl groups (C10-C14). The 3D structural model of LipC24 revealed the open Y-type active site. LipA displayed 96 % amino acid sequence identity with the known extracellular lipase. lipA-inactivation and lipC24-inactivation decreased the total cell-bound lipase activity of Burkholderia sp. ZYB002 by 42 % and 14 %, respectively. The cell-bound lipase activity from Burkholderia sp. ZYB002 originated from a multi-enzyme mixture with LipA as the main component. LipC24 was a novel lipase and displayed different enzymatic characteristics and structural model with LipA. Besides LipA and LipC24, other type of the cell-bound lipases (or esterases) should exist.

Apo And Calcium-Bound Crystal Structures of Alpha-11 Giardin, An Unusual Annexin From 'Giardia Lamblia'

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

Pathuri, P.; Nguyen, E.T.; Svard, S.G.

2007-07-12

Alpha-11 giardin is a member of the multi-gene alpha-giardin family in the intestinal protozoan, Giardia lamblia. This gene family shares an ancestry with the annexin super family, whose common characteristic is calcium-dependent binding to membranes that contain acidic phospholipids. Several alpha giardins are highly expressed during parasite-induced diarrhea in humans. Despite being a member of a large family of proteins, little is known about the function and cellular localization of alpha-11 giardin, although giardins are often associated with the cytoskeleton. It has been shown that Giardia exhibits high levels of alpha-11 giardin mRNA transcript throughout its life cycle; however, constitutivemore » over-expression of this protein is lethal to the parasite. Determining the three-dimensional structure of an alpha-giardin is essential to identifying functional domains shared in the alpha-giardin family. Here we report the crystal structures of the apo and Ca{sup 2+}-bound forms of alpha-11 giardin, the first alpha giardin to be characterized structurally. Crystals of apo and Ca{sup 2+}-bound alpha-11 giardin diffracted to 1.1 angstroms and 2.93 angstroms, respectively. The crystal structure of selenium-substituted apo alpha-11 giardin reveals a planar array of four tandem repeats of predominantly {alpha}-helical domains, reminiscent of previously determined annexin structures, making this the highest-resolution structure of an annexin to date. The apo alpha-11 giardin structure also reveals a hydrophobic core formed between repeats I/IV and II/III, a region typically hydrophilic in other annexins. Surprisingly, the Ca{sup 2+}-bound structure contains only a single calcium ion, located in the DE loop of repeat I and coordinated differently from the two types of calcium sites observed in previous annexin structures. The apo and Ca{sup 2+}-bound alpha-11 giardin structures assume overall similar conformations; however, Ca2+-bound alpha-11 giardin crystallized in a lower-symmetry space group with four molecules in the asymmetric unit. Vesicle-binding studies suggest that alpha-11 giardin, unlike most other annexins, does not bind to vesicles composed of acidic phospholipids in a calcium-dependent manner.« less

Diphytanyl glycerol ether distributions in sediments of the Orca Basin

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

Pease, T.K.; VanVleet, E.S.; Barre, J.S.

1992-09-01

Archaebacterially produced diphytanyl glycerol ether (DPGE) was examined in core sediments from the Orca Basin, an anoxic hypersaline basin in the northwestern Gulf of Mexico, to observe its spatial variability and potential origin. A differential extraction protocol was employed to quantify the isopranyl glycerol ethers associated with unbound, intermediate-bound, and kerogen-bound lipid fractions. Archaebacterial lipids were evident at all depths for the unbound and intermediate-bound fractions. Concentrations of DPGE ranged from 0.51 to 2.91 [mu]g/g dry sediment at the surface and showed secondary maxima deeper in basin sediments. Intermediate-bound DPGE concentrations exhibited an inverse relationship to unbound DPGE concentrations. Kerogen-boundmore » DPGE concentrations were normally below detection limits. Earlier studies describing the general homogeneity of lipid components within the overlying brine and at the brine/seawater interface suggest that the large-scale sedimentary DPGE variations observed in this study result from spatial and temporal variations in in-situ production by methanogenic or extremely halophilic archaebacteria.« less

Reversibly bound chloride in the atrial natriuretic peptide receptor hormone-binding domain: possible allosteric regulation and a conserved structural motif for the chloride-binding site.

PubMed

Ogawa, Haruo; Qiu, Yue; Philo, John S; Arakawa, Tsutomu; Ogata, Craig M; Misono, Kunio S

2010-03-01

The binding of atrial natriuretic peptide (ANP) to its receptor requires chloride, and it is chloride concentration dependent. The extracellular domain (ECD) of the ANP receptor (ANPR) contains a chloride near the ANP-binding site, suggesting a possible regulatory role. The bound chloride, however, is completely buried in the polypeptide fold, and its functional role has remained unclear. Here, we have confirmed that chloride is necessary for ANP binding to the recombinant ECD or the full-length ANPR expressed in CHO cells. ECD without chloride (ECD(-)) did not bind ANP. Its binding activity was fully restored by bromide or chloride addition. A new X-ray structure of the bromide-bound ECD is essentially identical to that of the chloride-bound ECD. Furthermore, bromide atoms are localized at the same positions as chloride atoms both in the apo and in the ANP-bound structures, indicating exchangeable and reversible halide binding. Far-UV CD and thermal unfolding data show that ECD(-) largely retains the native structure. Sedimentation equilibrium in the absence of chloride shows that ECD(-) forms a strongly associated dimer, possibly preventing the structural rearrangement of the two monomers that is necessary for ANP binding. The primary and tertiary structures of the chloride-binding site in ANPR are highly conserved among receptor-guanylate cyclases and metabotropic glutamate receptors. The chloride-dependent ANP binding, reversible chloride binding, and the highly conserved chloride-binding site motif suggest a regulatory role for the receptor bound chloride. Chloride-dependent regulation of ANPR may operate in the kidney, modulating ANP-induced natriuresis.

Reversibly Bound Chloride in the Atrial Natriuretic Peptide Receptor Hormone Binding Domain: Possible Allosteric Regulation and a Conserved Structural Motif for the Chloride-binding Site

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

Ogawa, H.; Qiu, Y; Philo, J

2010-01-01

The binding of atrial natriuretic peptide (ANP) to its receptor requires chloride, and it is chloride concentration dependent. The extracellular domain (ECD) of the ANP receptor (ANPR) contains a chloride near the ANP-binding site, suggesting a possible regulatory role. The bound chloride, however, is completely buried in the polypeptide fold, and its functional role has remained unclear. Here, we have confirmed that chloride is necessary for ANP binding to the recombinant ECD or the full-length ANPR expressed in CHO cells. ECD without chloride (ECD(-)) did not bind ANP. Its binding activity was fully restored by bromide or chloride addition. Amore » new X-ray structure of the bromide-bound ECD is essentially identical to that of the chloride-bound ECD. Furthermore, bromide atoms are localized at the same positions as chloride atoms both in the apo and in the ANP-bound structures, indicating exchangeable and reversible halide binding. Far-UV CD and thermal unfolding data show that ECD(-) largely retains the native structure. Sedimentation equilibrium in the absence of chloride shows that ECD(-) forms a strongly associated dimer, possibly preventing the structural rearrangement of the two monomers that is necessary for ANP binding. The primary and tertiary structures of the chloride-binding site in ANPR are highly conserved among receptor-guanylate cyclases and metabotropic glutamate receptors. The chloride-dependent ANP binding, reversible chloride binding, and the highly conserved chloride-binding site motif suggest a regulatory role for the receptor bound chloride. Chloride-dependent regulation of ANPR may operate in the kidney, modulating ANP-induced natriuresis.« less

Reversibly bound chloride in the atrial natriuretic peptide receptor hormone-binding domain: Possible allosteric regulation and a conserved structural motif for the chloride-binding site

PubMed Central

Ogawa, Haruo; Qiu, Yue; Philo, John S; Arakawa, Tsutomu; Ogata, Craig M; Misono, Kunio S

2010-01-01

The binding of atrial natriuretic peptide (ANP) to its receptor requires chloride, and it is chloride concentration dependent. The extracellular domain (ECD) of the ANP receptor (ANPR) contains a chloride near the ANP-binding site, suggesting a possible regulatory role. The bound chloride, however, is completely buried in the polypeptide fold, and its functional role has remained unclear. Here, we have confirmed that chloride is necessary for ANP binding to the recombinant ECD or the full-length ANPR expressed in CHO cells. ECD without chloride (ECD(−)) did not bind ANP. Its binding activity was fully restored by bromide or chloride addition. A new X-ray structure of the bromide-bound ECD is essentially identical to that of the chloride-bound ECD. Furthermore, bromide atoms are localized at the same positions as chloride atoms both in the apo and in the ANP-bound structures, indicating exchangeable and reversible halide binding. Far-UV CD and thermal unfolding data show that ECD(−) largely retains the native structure. Sedimentation equilibrium in the absence of chloride shows that ECD(−) forms a strongly associated dimer, possibly preventing the structural rearrangement of the two monomers that is necessary for ANP binding. The primary and tertiary structures of the chloride-binding site in ANPR are highly conserved among receptor-guanylate cyclases and metabotropic glutamate receptors. The chloride-dependent ANP binding, reversible chloride binding, and the highly conserved chloride-binding site motif suggest a regulatory role for the receptor bound chloride. Chloride-dependent regulation of ANPR may operate in the kidney, modulating ANP-induced natriuresis. PMID:20066666

"Experiential" Professional Development: Improving World Language Pedagogy inside Spanish Classrooms

ERIC Educational Resources Information Center

Burke, Brigid Moira

2012-01-01

"Experiential" professional development (EPD), influenced by Expeditionary Learning Outward Bound design, was integrated in the classrooms of secondary Spanish teachers to create opportunities for them to learn to use communicative language teaching (CLT) through experience. Teachers collaborated with colleagues, students, and a…

Structure and symmetry inform gating principles of ionotropic glutamate receptors.

PubMed

Zhu, Shujia; Gouaux, Eric

2017-01-01

Ionotropic glutamate receptors (iGluRs) transduce signals derived from release of the excitatory neurotransmitter glutamate from pre-synaptic neurons into excitation of post-synaptic neurons on a millisecond time-scale. In recent years, the elucidation of full-length iGluR structures of NMDA, AMPA and kainate receptors by X-ray crystallography and single particle cryo-electron microscopy has greatly enhanced our understanding of the interrelationships between receptor architecture and gating mechanism. Here we briefly review full-length iGluR structures and discuss the similarities and differences between NMDA receptors and non-NMDA iGluRs. We focus on distinct conformations, including ligand-free, agonist-bound active, agonist-bound desensitized and antagonist-bound conformations as well as modulator and auxiliary protein-bound states. These findings provide insights into structure-based mechanisms of iGluR gating and modulation which together shape the amplitude and time course of the excitatory postsynaptic potential. This article is part of the Special Issue entitled 'Ionotropic glutamate receptors'. Copyright © 2016 Elsevier Ltd. All rights reserved.

A fresh look at linear cosmological constraints on a decaying Dark Matter component

NASA Astrophysics Data System (ADS)

Poulin, Vivian; Serpico, Pasquale D.; Lesgourgues, Julien

2016-08-01

We consider a cosmological model in which a fraction fdcdm of the Dark Matter (DM) is allowed to decay in an invisible relativistic component, and compute the resulting constraints on both the decay width (or inverse lifetime) Γdcdm and fdcdm from purely gravitational arguments. We report a full derivation of the Boltzmann hierarchy, correcting a mistake in previous literature, and compute the impact of the decay—as a function of the lifetime—on the CMB and matter power spectra. From CMB only, we obtain that no more than 3.8% of the DM could have decayed in the time between recombination and today (all bounds quoted at 95% CL). We also comment on the important application of this bound to the case where primordial black holes constitute DM, a scenario notoriously difficult to constrain. For lifetimes longer than the age of the Universe, the bounds can be cast as fdcdmΓdcdm < 6.3×10-3 Gyr-1. For the first time, we also checked that degeneracies with massive neutrinos are broken when information from the large scale structure is used. Even secondary effects like CMB lensing suffice to this purpose. Decaying DM models have been invoked to solve a possible tension between low redshift astronomical measurements of σ8 and Ωm and the ones inferred by Planck. We reassess this claim finding that with the most recent BAO, HST and σ8 data extracted from the CFHT survey, the tension is only slightly reduced despite the two additional free parameters. Nonetheless, the existing tension explains why the bound on fdcdmΓdcdm loosens to fdcdmΓdcdm < 15.9×10-3 Gyr-1 when including such additional data. The bound however improves to fdcdmΓdcdm < 5.9 ×10-3 Gyr-1 if only data consistent with the CMB are included. This highlights the importance of establishing whether the tension is due to real physical effects or unaccounted systematics, for settling the reach of achievable constraints on decaying DM.

RNA-SSPT: RNA Secondary Structure Prediction Tools.

PubMed

Ahmad, Freed; Mahboob, Shahid; Gulzar, Tahsin; Din, Salah U; Hanif, Tanzeela; Ahmad, Hifza; Afzal, Muhammad

2013-01-01

The prediction of RNA structure is useful for understanding evolution for both in silico and in vitro studies. Physical methods like NMR studies to predict RNA secondary structure are expensive and difficult. Computational RNA secondary structure prediction is easier. Comparative sequence analysis provides the best solution. But secondary structure prediction of a single RNA sequence is challenging. RNA-SSPT is a tool that computationally predicts secondary structure of a single RNA sequence. Most of the RNA secondary structure prediction tools do not allow pseudoknots in the structure or are unable to locate them. Nussinov dynamic programming algorithm has been implemented in RNA-SSPT. The current studies shows only energetically most favorable secondary structure is required and the algorithm modification is also available that produces base pairs to lower the total free energy of the secondary structure. For visualization of RNA secondary structure, NAVIEW in C language is used and modified in C# for tool requirement. RNA-SSPT is built in C# using Dot Net 2.0 in Microsoft Visual Studio 2005 Professional edition. The accuracy of RNA-SSPT is tested in terms of Sensitivity and Positive Predicted Value. It is a tool which serves both secondary structure prediction and secondary structure visualization purposes.

RNA-SSPT: RNA Secondary Structure Prediction Tools

PubMed Central

Ahmad, Freed; Mahboob, Shahid; Gulzar, Tahsin; din, Salah U; Hanif, Tanzeela; Ahmad, Hifza; Afzal, Muhammad

2013-01-01

The prediction of RNA structure is useful for understanding evolution for both in silico and in vitro studies. Physical methods like NMR studies to predict RNA secondary structure are expensive and difficult. Computational RNA secondary structure prediction is easier. Comparative sequence analysis provides the best solution. But secondary structure prediction of a single RNA sequence is challenging. RNA-SSPT is a tool that computationally predicts secondary structure of a single RNA sequence. Most of the RNA secondary structure prediction tools do not allow pseudoknots in the structure or are unable to locate them. Nussinov dynamic programming algorithm has been implemented in RNA-SSPT. The current studies shows only energetically most favorable secondary structure is required and the algorithm modification is also available that produces base pairs to lower the total free energy of the secondary structure. For visualization of RNA secondary structure, NAVIEW in C language is used and modified in C# for tool requirement. RNA-SSPT is built in C# using Dot Net 2.0 in Microsoft Visual Studio 2005 Professional edition. The accuracy of RNA-SSPT is tested in terms of Sensitivity and Positive Predicted Value. It is a tool which serves both secondary structure prediction and secondary structure visualization purposes. PMID:24250115

Hidden Markov model approach for identifying the modular framework of the protein backbone.

PubMed

Camproux, A C; Tuffery, P; Chevrolat, J P; Boisvieux, J F; Hazout, S

1999-12-01

The hidden Markov model (HMM) was used to identify recurrent short 3D structural building blocks (SBBs) describing protein backbones, independently of any a priori knowledge. Polypeptide chains are decomposed into a series of short segments defined by their inter-alpha-carbon distances. Basically, the model takes into account the sequentiality of the observed segments and assumes that each one corresponds to one of several possible SBBs. Fitting the model to a database of non-redundant proteins allowed us to decode proteins in terms of 12 distinct SBBs with different roles in protein structure. Some SBBs correspond to classical regular secondary structures. Others correspond to a significant subdivision of their bounding regions previously considered to be a single pattern. The major contribution of the HMM is that this model implicitly takes into account the sequential connections between SBBs and thus describes the most probable pathways by which the blocks are connected to form the framework of the protein structures. Validation of the SBBs code was performed by extracting SBB series repeated in recoding proteins and examining their structural similarities. Preliminary results on the sequence specificity of SBBs suggest promising perspectives for the prediction of SBBs or series of SBBs from the protein sequences.

LUTE telescope structural design

NASA Technical Reports Server (NTRS)

Ruthven, Gregory

1993-01-01

The major objective of the Lunar Ultraviolet Transit Experiment (LUTE) Telescope Structural Design Study was to investigate the feasibility of designing an ultralightweight 1-m aperture system within optical performance requirements and mass budget constraints. This study uses the results from our previous studies on LUTE as a basis for further developing the LUTE structural architecture. After summarizing our results in Section 2, Section 3 begins with the overall logic we used to determine which telescope 'structural form' should be adopted for further analysis and weight estimates. Specific telescope component analysis showing calculated fundamental frequencies and how they compare with our derived requirements are included. 'First-order' component stress analyses to ensure telescope optical and structural component (i.e. mirrors & main bulkhead) weights are realistic are presented. Layouts of both the primary and tertiary mirrors showing dimensions that are consistent with both our weight and frequency calculations also form part of Section 3. Section 4 presents our calculated values for the predicted thermally induced primary-to-secondary mirror despace motion due to the large temperature range over which LUTE must operate. Two different telescope design approaches (one which utilizes fused quartz metering rods and one which assumes the entire telescope is fabricated from beryllium) are considered in this analysis. We bound the secondary mirror focus mechanism range (in despace) based on these two telescope configurations. In Section 5 we show our overall design of the UVTA (Ultraviolet Telescope Assembly) via an 'exploded view' of the sub-system. The 'exploded view' is annotated to help aid in the understanding of each sub-assembly. We also include a two view layout of the UVTA from which telescope and telescope component dimensions can be measured. We conclude our study with a set of recommendations not only with respect to the LUTE structural architecture but also on other topics related to the overall feasibility of the LUTE telescope sub-system.

Fragment-based discovery of a potent NAMPT inhibitor.

PubMed

Korepanova, Alla; Longenecker, Kenton L; Pratt, Steve D; Panchal, Sanjay C; Clark, Richard F; Lake, Marc; Gopalakrishnan, Sujatha M; Raich, Diana; Sun, Chaohong; Petros, Andrew M

2017-12-12

NAMPT expression is elevated in many cancers, making this protein a potential target for anticancer therapy. We have carried out both NMR based and TR-FRET based fragment screens against human NAMPT and identified six novel binders with a range of potencies. Co-crystal structures were obtained for two of the fragments bound to NAMPT while for the other four fragments force-field driven docking was employed to generate a bound pose. Based on structural insights arising from comparison of the bound fragment poses to that of bound FK866 we were able to synthetically elaborate one of the fragments into a potent NAMPT inhibitor. Copyright © 2017 Elsevier Ltd. All rights reserved.

The structures and properties of proton- and alkali-bound cysteine dimers.

PubMed

Ieritano, Christian; Carr, Patrick J J; Hasan, Moaraj; Burt, Michael; Marta, Rick A; Steinmetz, Vincent; Fillion, Eric; McMahon, Terrance B; Scott Hopkins, W

2016-02-14

The proton-, lithium-, and sodium-bound cysteine dimers have been investigated in a joint computational and experimental infrared multiple photon dissociation (IRMPD) study. IRMPD spectra in the 1000-2000 cm(-1) region show that protonation is localized on an amine group, and that intermolecular hydrogen bonding occurs between the protonated amine and the carbonyl oxygen of the neutral Cys moiety. Alkali-bound dimers adopt structures reminiscent of those observed for the monomeric Cys·Li(+) and Cys·Na(+) species. Calculations of the heavier Cys2·M(+) (M = K, Rb or Cs) species suggest that these are significantly less strongly bound than the lighter (M = H, Li, or Na) dimers.

Structure and dynamics of protein waters revealed by radiolysis and mass spectrometry

PubMed Central

Gupta, Sayan; D’Mello, Rhijuta; Chance, Mark R.

2012-01-01

Water is critical for the structure, stability, and functions of macromolecules. Diffraction and NMR studies have revealed structure and dynamics of bound waters at atomic resolution. However, localizing the sites and measuring the dynamics of bound waters, particularly on timescales relevant to catalysis and macromolecular assembly, is quite challenging. Here we demonstrate two techniques: first, temperature-dependent radiolytic hydroxyl radical labeling with a mass spectrometry (MS)-based readout to identify sites of bulk and bound water interactions with surface and internal residue side chains, and second, H218O radiolytic exchange coupled MS to measure the millisecond dynamics of bound water interactions with various internal residue side chains. Through an application of the methods to cytochrome c and ubiquitin, we identify sites of water binding and measure the millisecond dynamics of bound waters in protein crevices. As these MS-based techniques are very sensitive and not protein size limited, they promise to provide unique insights into protein–water interactions and water dynamics for both small and large proteins and their complexes. PMID:22927377

Eastward Bound

ERIC Educational Resources Information Center

Ly, Phuong

2012-01-01

In recent years, the economic boom in Asia has resulted in more families sending their children to independent secondary schools in North America, Europe, and Australia. Donations, however, haven't been as forthcoming. Schools often have to start from scratch in educating parents about the need for fundraising. Independent schools, more so than…

Distance-Based Configurational Entropy of Proteins from Molecular Dynamics Simulations

PubMed Central

Fogolari, Federico; Corazza, Alessandra; Fortuna, Sara; Soler, Miguel Angel; VanSchouwen, Bryan; Brancolini, Giorgia; Corni, Stefano; Melacini, Giuseppe; Esposito, Gennaro

2015-01-01

Estimation of configurational entropy from molecular dynamics trajectories is a difficult task which is often performed using quasi-harmonic or histogram analysis. An entirely different approach, proposed recently, estimates local density distribution around each conformational sample by measuring the distance from its nearest neighbors. In this work we show this theoretically well grounded the method can be easily applied to estimate the entropy from conformational sampling. We consider a set of systems that are representative of important biomolecular processes. In particular: reference entropies for amino acids in unfolded proteins are obtained from a database of residues not participating in secondary structure elements;the conformational entropy of folding of β2-microglobulin is computed from molecular dynamics simulations using reference entropies for the unfolded state;backbone conformational entropy is computed from molecular dynamics simulations of four different states of the EPAC protein and compared with order parameters (often used as a measure of entropy);the conformational and rototranslational entropy of binding is computed from simulations of 20 tripeptides bound to the peptide binding protein OppA and of β2-microglobulin bound to a citrate coated gold surface. This work shows the potential of the method in the most representative biological processes involving proteins, and provides a valuable alternative, principally in the shown cases, where other approaches are problematic. PMID:26177039

Distance-Based Configurational Entropy of Proteins from Molecular Dynamics Simulations.

PubMed

Fogolari, Federico; Corazza, Alessandra; Fortuna, Sara; Soler, Miguel Angel; VanSchouwen, Bryan; Brancolini, Giorgia; Corni, Stefano; Melacini, Giuseppe; Esposito, Gennaro

2015-01-01

Estimation of configurational entropy from molecular dynamics trajectories is a difficult task which is often performed using quasi-harmonic or histogram analysis. An entirely different approach, proposed recently, estimates local density distribution around each conformational sample by measuring the distance from its nearest neighbors. In this work we show this theoretically well grounded the method can be easily applied to estimate the entropy from conformational sampling. We consider a set of systems that are representative of important biomolecular processes. In particular: reference entropies for amino acids in unfolded proteins are obtained from a database of residues not participating in secondary structure elements;the conformational entropy of folding of β2-microglobulin is computed from molecular dynamics simulations using reference entropies for the unfolded state;backbone conformational entropy is computed from molecular dynamics simulations of four different states of the EPAC protein and compared with order parameters (often used as a measure of entropy);the conformational and rototranslational entropy of binding is computed from simulations of 20 tripeptides bound to the peptide binding protein OppA and of β2-microglobulin bound to a citrate coated gold surface. This work shows the potential of the method in the most representative biological processes involving proteins, and provides a valuable alternative, principally in the shown cases, where other approaches are problematic.

In silico studies of the properties of water hydrating a small protein

NASA Astrophysics Data System (ADS)

Sinha, Sudipta Kumar; Jana, Madhurima; Chakraborty, Kausik; Bandyopadhyay, Sanjoy

2014-12-01

Atomistic molecular dynamics simulation of an aqueous solution of the small protein HP-36 has been carried out with explicit solvent at room temperature. Efforts have been made to explore the influence of the protein on the relative packing and ordering of water molecules around its secondary structures, namely, three α-helices. The calculations reveal that the inhomogeneous water ordering and density distributions around the helices are correlated with their relative hydrophobicity. Importantly, we have identified the existence of a narrow relatively dehydrated region containing randomly organized "quasi-free" water molecules beyond the first layer of "bound" waters at the protein surface. These water molecules with relatively weaker binding energies form the transition state separating the "bound" and "free" water molecules at the interface. Further, increased contribution of solid-like caging motions of water molecules around the protein is found to be responsible for reduced fluidity of the hydration layer. Interestingly, we notice that the hydration layer of helix-3 is more fluidic with relatively higher entropy as compared to the hydration layers of the other two helical segments. Such characteristics of helix-3 hydration layer correlate well with the activity of HP-36, as helix-3 contains the active site of the protein.

Propagation of waves in a bounded random layer with laminar structure

NASA Technical Reports Server (NTRS)

Karam, M. A.; Fung, A. K.

1985-01-01

A closed form solution has been developed to obtain the intensity propagating in a bounded layer with laminar structure. Then, the brightness temperature due to an arbitrary temperature profile has been derived. Results are specialized to a half space to compare with those reported in the literature.

The Influence of Verb-Bound Syntactic Preferences on the Processing of Syntactic Structures

ERIC Educational Resources Information Center

Segaert, Katrien; Weber, Kirsten; Cladder-Micus, Mira; Hagoort, Peter

2014-01-01

Speakers sometimes repeat syntactic structures across sentences, a phenomenon called syntactic priming. We investigated the influence of verb-bound syntactic preferences on syntactic priming effects in response choices and response latencies for German ditransitive sentences. In the response choices we found "inverse preference effects":…

K-Partite RNA Secondary Structures

NASA Astrophysics Data System (ADS)

Jiang, Minghui; Tejada, Pedro J.; Lasisi, Ramoni O.; Cheng, Shanhong; Fechser, D. Scott

RNA secondary structure prediction is a fundamental problem in structural bioinformatics. The prediction problem is difficult because RNA secondary structures may contain pseudoknots formed by crossing base pairs. We introduce k-partite secondary structures as a simple classification of RNA secondary structures with pseudoknots. An RNA secondary structure is k-partite if it is the union of k pseudoknot-free sub-structures. Most known RNA secondary structures are either bipartite or tripartite. We show that there exists a constant number k such that any secondary structure can be modified into a k-partite secondary structure with approximately the same free energy. This offers a partial explanation of the prevalence of k-partite secondary structures with small k. We give a complete characterization of the computational complexities of recognizing k-partite secondary structures for all k ≥ 2, and show that this recognition problem is essentially the same as the k-colorability problem on circle graphs. We present two simple heuristics, iterated peeling and first-fit packing, for finding k-partite RNA secondary structures. For maximizing the number of base pair stackings, our iterated peeling heuristic achieves a constant approximation ratio of at most k for 2 ≤ k ≤ 5, and at most frac6{1-(1-6/k)^k} le frac6{1-e^{-6}} < 6.01491 for k ≥ 6. Experiment on sequences from PseudoBase shows that our first-fit packing heuristic outperforms the leading method HotKnots in predicting RNA secondary structures with pseudoknots. Source code, data set, and experimental results are available at http://www.cs.usu.edu/ mjiang/rna/kpartite/.

Molecular Mechanism of Scanning and Start Codon Selection in Eukaryotes

PubMed Central

Hinnebusch, Alan G.

2011-01-01

Summary: The correct translation of mRNA depends critically on the ability to initiate at the right AUG codon. For most mRNAs in eukaryotic cells, this is accomplished by the scanning mechanism, wherein the small (40S) ribosomal subunit attaches to the 5′ end of the mRNA and then inspects the leader base by base for an AUG in a suitable context, using complementarity with the anticodon of methionyl initiator tRNA (Met-tRNAiMet) as the key means of identifying AUG. Over the past decade, a combination of yeast genetics, biochemical analysis in reconstituted systems, and structural biology has enabled great progress in deciphering the mechanism of ribosomal scanning. A robust molecular model now exists, describing the roles of initiation factors, notably eukaryotic initiation factor 1 (eIF1) and eIF1A, in stabilizing an “open” conformation of the 40S subunit with Met-tRNAiMet bound in a low-affinity state conducive to scanning and in triggering rearrangement into a “closed” conformation incompatible with scanning, which features Met-tRNAiMet more tightly bound to the “P” site and base paired with AUG. It has also emerged that multiple DEAD-box RNA helicases participate in producing a single-stranded “landing pad” for the 40S subunit and in removing the secondary structure to enable the mRNA to traverse the 40S mRNA-binding channel in the single-stranded form for base-by-base inspection in the P site. PMID:21885680

Structural insight into GRIP1-PDZ6 in Alzheimer's disease: study from protein expression data to molecular dynamics simulations.

PubMed

Chatterjee, Paulami; Roy, Debjani

2017-08-01

Protein-protein interaction domain, PDZ, plays a critical role in efficient synaptic transmission in brain. Dysfunction of synaptic transmission is thought to be the underlying basis of many neuropsychiatric and neurodegenerative disorders including Alzheimer's disease (AD). In this study, Glutamate Receptor Interacting Protein1 (GRIP1) was identified as one of the most important differentially expressed, topologically significant proteins in the protein-protein interaction network. To date, very few studies have analyzed the detailed structural basis of PDZ-mediated protein interaction of GRIP1. In order to gain better understanding of structural and dynamic basis of these interactions, we employed molecular dynamics (MD) simulations of GRIP1-PDZ6 dimer bound with Liprin-alpha and GRIP1-PDZ6 dimer alone each with 100 ns simulations. The analyses of MD simulations of Liprin-alpha bound GRIP1-PDZ6 dimer show considerable conformational differences than that of peptide-free dimer in terms of SASA, hydrogen bonding patterns, and along principal component 1 (PC1). Our study also furnishes insight into the structural attunement of the PDZ6 domains of Liprin-alpha bound GRIP1 that is attributed by significant shift of the Liprin-alpha recognition helix in the simulated peptide-bound dimer compared to the crystal structure and simulated peptide-free dimer. It is evident that PDZ6 domains of peptide-bound dimer show differential movements along PC1 than that of peptide-free dimers. Thus, Liprin-alpha also serves an important role in conferring conformational changes along the dimeric interface of the peptide-bound dimer. Results reported here provide information that may lead to novel therapeutic approaches in AD.

Effects of atmospheric relative humidity on Stratum Corneum structure at the molecular level: ex vivo Raman spectroscopy analysis.

PubMed

Vyumvuhore, Raoul; Tfayli, Ali; Duplan, Hélène; Delalleau, Alexandre; Manfait, Michel; Baillet-Guffroy, Arlette

2013-07-21

Skin hydration plays an important role in the optimal physical properties and physiological functions of the skin. Despite the advancements in the last decade, dry skin remains the most common characteristic of human skin disorders. Thus, it is important to understand the effect of hydration on Stratum Corneum (SC) components. In this respect, our interest consists in correlating the variations of unbound and bound water content in the SC with structural and organizational changes in lipids and proteins using a non-invasive technique: Raman spectroscopy. Raman spectra were acquired on human SC at different relative humidity (RH) levels (4-75%). The content of different types of water, bound and free, was measured using the second derivative and curve fitting of the Raman bands in the range of 3100-3700 cm(-1). Changes in lipidic order were evaluated using νC-C and νC-H. To analyze the effect of RH on the protein structure, we examined in the Amide I region, the Fermi doublet of tyrosine, and the νasymCH3 vibration. The contributions of totally bound water were found not to vary with humidity, while partially bound water varied with three different rates. Unbound water increased greatly when all sites for bound water were saturated. Lipid organization as well as protein deployment was found to be optimal at intermediate RH values (around 60%), which correspond to the maximum of SC water binding capacity. This analysis highlights the relationship between bound water, the SC barrier state and the protein structure and elucidates the optimal conditions. Moreover, our results showed that increased content of unbound water in the SC induces disorder in the structures of lipids and proteins.

Subject- and Experience-Bound Differences in Teachers' Conceptual Understanding of Sustainable Development

ERIC Educational Resources Information Center

Borg, C.; Gericke, N.; Höglund, H.-O.; Bergman, E.

2014-01-01

This article describes the results of a nationwide questionnaire study of 3229 Swedish upper secondary school teachers' conceptual understanding of sustainable development in relation to their subject discipline and teaching experience. Previous research has shown that teachers have difficulties understanding the complex concept of sustainable…

Limits on estimating the width of thin tubular structures in 3D images.

PubMed

Wörz, Stefan; Rohr, Karl

2006-01-01

This work studies limits on estimating the width of thin tubular structures in 3D images. Based on nonlinear estimation theory we analyze the minimal stochastic error of estimating the width. Given a 3D analytic model of the image intensities of tubular structures, we derive a closed-form expression for the Cramér-Rao bound of the width estimate under image noise. We use the derived lower bound as a benchmark and compare it with three previously proposed accuracy limits for vessel width estimation. Moreover, by experimental investigations we demonstrate that the derived lower bound can be achieved by fitting a 3D parametric intensity model directly to the image data.

Structural and dynamical properties of recombining ultracold neutral plasma

NASA Astrophysics Data System (ADS)

Tiwari, Sanat Kumar; Shaffer, Nathaniel R.; Baalrud, Scott D.

2017-10-01

An ultracold plasma (UCP) is an evolving collection of free charges and bound charges (Rydberg atoms). Over time, bound species concentration increases due to recombination. We present the structural and dynamical properties of an evolving UCP using classical molecular dynamics simulation. Coulomb collapse is avoided using a repulsive core with the attractive Coulomb potential. The repulsive core size controls the concentration of bound states, as it determines the depth of the potential well between opposite charges. We vary the repulsive core size to emulate the quasi-static state of plasma at different time during the evolution. Binary, chain and ring-like bound states are observed in the simulation carried out at different coupling strengths and repulsive core size. The effect of bound states can be seen as molecular peaks in the radial distribution function (RDF). The thermodynamic properties associated with the free charges can be analyzed from RDF by separating free from bound states. These bound states also change the dynamical properties of the plasma. The electron velocity auto-correlation displays oscillations due to the orbital motion in bound states. These bound states act like a neutral species, damping electron plasmon modes and broadening the ion acoustic mode. This work is supported by AFOSR Grant Number FA9550-16-1-0221. It used computational resources by XSEDE, which is supported by NSF Grant Number ACI-1053575.

Protein-protein binding before and after photo-modification of albumin

NASA Astrophysics Data System (ADS)

Rozinek, Sarah C.; Glickman, Randolph D.; Thomas, Robert J.; Brancaleon, Lorenzo

2016-03-01

Bioeffects of directed-optical-energy encompass a wide range of applications. One aspect of photochemical interactions involves irradiating a photosensitizer with visible light in order to induce protein unfolding and consequent changes in function. In the past, irradiation of several dye-protein combinations has revealed effects on protein structure. Beta lactoglobulin, human serum albumin (HSA) and tubulin have all been photo-modified with meso-tetrakis(4- sulfonatophenyl)porphyrin (TSPP) bound, but only in the case of tubulin has binding caused a verified loss of biological function (loss of ability to form microtubules) as a result of this light-induced structural change. The current work questions if the photo-induced structural changes that occur to HSA, are sufficient to disable its biological function of binding to osteonectin. The albumin-binding protein, osteonectin, is about half the molecular weight of HSA, so the two proteins and their bound product can be separated and quantified by size exclusion high performance liquid chromatography. TSPP was first bound to HSA and irradiated, photo-modifying the structure of HSA. Then native HSA or photo-modified HSA (both with TSPP bound) were compared, to assess loss in HSA's innate binding ability as a result of light-induced structure modification.

Robust stability bounds for multi-delay networked control systems

NASA Astrophysics Data System (ADS)

Seitz, Timothy; Yedavalli, Rama K.; Behbahani, Alireza

2018-04-01

In this paper, the robust stability of a perturbed linear continuous-time system is examined when controlled using a sampled-data networked control system (NCS) framework. Three new robust stability bounds on the time-invariant perturbations to the original continuous-time plant matrix are presented guaranteeing stability for the corresponding discrete closed-loop augmented delay-free system (ADFS) with multiple time-varying sensor and actuator delays. The bounds are differentiated from previous work by accounting for the sampled-data nature of the NCS and for separate communication delays for each sensor and actuator, not a single delay. Therefore, this paper expands the knowledge base in multiple inputs multiple outputs (MIMO) sampled-data time delay systems. Bounds are presented for unstructured, semi-structured, and structured perturbations.

Crystal structure of cGMP-dependent protein kinase Iβ cyclic nucleotide-binding-B domain : Rp-cGMPS complex reveals an apo-like, inactive conformation

DOE PAGES

Campbell, James C.; VanSchouwen, Bryan; Lorenz, Robin; ...

2016-12-23

The R-diastereomer of phosphorothioate analogs of cGMP, Rp-cGMPS, is one of few known inhibitors of cGMP-dependent protein kinase I (PKG I); however, its mechanism of inhibition is currently not fully understood. We determined the crystal structure of the PKG Iβ cyclic nucleotide-binding domain (PKG Iβ CNB-B), considered a ‘gatekeeper’ for cGMP activation, bound to Rp-cGMPS at 1.3 Å. Our structural and NMR data show that PKG Iβ CNB-B bound to Rp-cGMPS displays an apo-like structure with its helical domain in an open conformation. Comparison with the cAMP-dependent protein kinase regulatory subunit (PKA RIα) showed that this conformation resembles the catalyticmore » subunit-bound inhibited state of PKA RIα more closely than the apo or Rp-cAMPS-bound conformations. Our results suggest that Rp-cGMPS inhibits PKG I by stabilizing the inactive conformation of CNB-B.« less

Crystal Structures of SlyA Protein, a Master Virulence Regulator of Salmonella, in Free and DNA-bound States

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

Dolan, Kyle T.; Duguid, Erica M.; He, Chuan

2011-11-17

SlyA is a master virulence regulator that controls the transcription of numerous genes in Salmonella enterica. We present here crystal structures of SlyA by itself and bound to a high-affinity DNA operator sequence in the slyA gene. SlyA interacts with DNA through direct recognition of a guanine base by Arg-65, as well as interactions between conserved Arg-86 and the minor groove and a large network of non-base-specific contacts with the sugar phosphate backbone. Our structures, together with an unpublished structure of SlyA bound to the small molecule effector salicylate (Protein Data Bank code 3DEU), reveal that, unlike many other MarRmore » family proteins, SlyA dissociates from DNA without large conformational changes when bound to this effector. We propose that SlyA and other MarR global regulators rely more on indirect readout of DNA sequence to exert control over many genes, in contrast to proteins (such as OhrR) that recognize a single operator.« less

Neutron diffraction of acetazolamide-bound human carbonic anhydrase II reveals atomic details of drug binding

PubMed Central

Fisher, S. Zoë; Aggarwal, Mayank; Kovalevsky, Andrey Y.; Silverman, David N.; McKenna, Robert

2012-01-01

Carbonic anhydrases (CAs) catalyze the hydration of CO2 forming HCO3− and a proton, an important reaction for many physiological processes including respiration, fluid secretion, and pH regulation. As such, CA isoforms are prominent clinical targets for treating various diseases. The clinically used acetazolamide (AZM) is a sulfonamide that binds with high affinity to human CA isoform II (HCA II). There are several X-ray structures available of AZM bound to various CA isoforms, but these complexes do not show the charged state of AZM, or hydrogen (H) atom positions of the protein and solvent. Neutron diffraction is a useful technique for directly observing H atoms and the mapping of H-bonding networks that can greatly contribute to rational drug design. To this end the neutron structure of H/D exchanged HCA II crystals in complex with AZM was determined. The structure reveals the molecular details of AZM binding and the charged state of the bound drug. This represents the first determined neutron structure of a clinically used drug bound to its target. PMID:22928733

Structural and Physical Basis for Anti-IgE Therapy

NASA Astrophysics Data System (ADS)

Wright, Jon D.; Chu, Hsing-Mao; Huang, Chun-Hsiang; Ma, Che; Wen Chang, Tse; Lim, Carmay

2015-06-01

Omalizumab, an anti-IgE antibody, used to treat severe allergic asthma and chronic idiopathic urticaria, binds to IgE in blood or membrane-bound on B lymphocytes but not to IgE bound to its high (FcɛRI) or low (CD23) affinity receptor. Mutagenesis studies indicate overlapping FcɛRI and omalizumab-binding sites in the Cɛ3 domain, but crystallographic studies show FcɛRI and CD23-binding sites that are far apart, so how can omalizumab block IgE from binding both receptors? We report a 2.42-Å omalizumab-Fab structure, a docked IgE-Fc/omalizumab-Fab structure consistent with available experimental data, and the free energy contributions of IgE residues to binding omalizumab, CD23, and FcɛRI. These results provide a structural and physical basis as to why omalizumab cannot bind receptor-bound IgE and why omalizumab-bound IgE cannot bind to CD23/FcɛRI. They reveal the key IgE residues and their roles in binding omalizumab, CD23, and FcɛRI.

Ensemble-based characterization of unbound and bound states on protein energy landscape

PubMed Central

Ruvinsky, Anatoly M; Kirys, Tatsiana; Tuzikov, Alexander V; Vakser, Ilya A

2013-01-01

Physicochemical description of numerous cell processes is fundamentally based on the energy landscapes of protein molecules involved. Although the whole energy landscape is difficult to reconstruct, increased attention to particular targets has provided enough structures for mapping functionally important subspaces associated with the unbound and bound protein structures. The subspace mapping produces a discrete representation of the landscape, further called energy spectrum. We compiled and characterized ensembles of bound and unbound conformations of six small proteins and explored their spectra in implicit solvent. First, the analysis of the unbound-to-bound changes points to conformational selection as the binding mechanism for four proteins. Second, results show that bound and unbound spectra often significantly overlap. Moreover, the larger the overlap the smaller the root mean square deviation (RMSD) between the bound and unbound conformational ensembles. Third, the center of the unbound spectrum has a higher energy than the center of the corresponding bound spectrum of the dimeric and multimeric states for most of the proteins. This suggests that the unbound states often have larger entropy than the bound states. Fourth, the exhaustively long minimization, making small intrarotamer adjustments (all-atom RMSD ≤ 0.7 Å), dramatically reduces the distance between the centers of the bound and unbound spectra as well as the spectra extent. It condenses unbound and bound energy levels into a thin layer at the bottom of the energy landscape with the energy spacing that varies between 0.8–4.6 and 3.5–10.5 kcal/mol for the unbound and bound states correspondingly. Finally, the analysis of protein energy fluctuations showed that protein vibrations itself can excite the interstate transitions, including the unbound-to-bound ones. PMID:23526684

Structural Studies on Intact Clostridium Botulinum Neurotoxins Complexed with Inhibitors Leading to Drug Design

DTIC Science & Technology

2006-02-01

6. Hanson, M. A., and Stevens, R. C. (2000) Cocrystal structure of synaptobrevin-ll bound to botulinum neurotoxin type B at 2.0 A resolution, Nature...Hanson, R.C. Stevens, Cocrystal structure of synaptobre- vin-ll bound to botulinum neurotoxin type B at 2.0 Å resolution, Nat. Struct. Biol. 7 (2000...of the Glu212 carboxylate in the catalytic pathway, Biochemistry 43, 6637-6644. 4. Hanson, M. A., and Stevens, R. C. (2000) Cocrystal structure of

Metal binding characterization and conformational studies using Raman microscopy of resin-bound poly(aspartic acid).

PubMed

Stair, Jacqueline L; Holcombe, James A

2007-03-01

The metal binding capacities, conditional stability constants, and secondary structure of immobilized polyaspartic acid (PLAsp) (n = 6, 20, and 30) on TentaGel resin were determined when binding Mg2+, Co2+, Cd2+, and Ni2+. Metal binding to the synthesized peptides was evaluated using breakthrough curves from a packed microcolumn and flame atomic absorption spectrophotometry (FAAS) detection. The metal capacities reached values of 590, 2160, and 3710 mumol of metal/g of resin for the 6-mer, 20-mer, and 30-mer, respectively, and this resulted in 2-3 residues per metal for all peptides and metals tested. Surprisingly, the concentrated environment of the resin along with the spatial distribution of attachment groups allowed for most residues to participate in metal binding regardless of the peptide length. Conditional stability constants calculated using single metal binding isotherms indicated that binding strength decreased as the chain length increased on the resin. Raman microscopy on single beads was used to determine PLAsp secondary structure, and all peptides were of a mixed conformation (i.e., beta-sheets, alpha-helices, random chain, etc.) during neutral conditioning and metal binding. Uniquely, the longer 20-mer and 30-mer peptides showed a distinct change from a mixed conformation to beta-sheets and alpha-helices during metal release with acid. This study confirms that metal release by longer immobilized peptides is often assisted by a conformational change, which easily spoils the binding cavity, while shorter peptides may release metal primarily by H+ displacement.

Effect of tin oxide nanoparticle binding on the structure and activity of α-amylase from Bacillus amyloliquefaciens

NASA Astrophysics Data System (ADS)

Jahir Khan, Mohammad; Qayyum, Shariq; Alam, Fahad; Husain, Qayyum

2011-11-01

Proteins adsorbed on nanoparticles (NPs) are being used in biotechnology, biosensors and drug delivery. However, understanding the effect of NPs on the structure of proteins is still in a nascent state. In the present paper tin oxide (SnO2) NPs were synthesized by the reaction of SnCl4·5H2O in methanol via the sol-gel method and characterized by x-ray diffraction (XRD), Fourier transform infrared spectroscopy (FT-IR) and transmission electron microscopy (TEM). The binding of these SnO2-NPs with α-amylase was investigated by using UV-vis, fluorescence and circular dichroism (CD) spectroscopic techniques. A strong quenching of tryptophan fluorescence intensity in α-amylase was observed due to formation of a ground state complex with SnO2-NPs. Far-UV CD spectra showed that the secondary structure of α-amylase was changed in the presence of NPs. The Michaelis-Menten constant (Km), was found to be 26.96 and 28.45 mg ml - 1, while Vmax was 4.173 and 3.116 mg ml - 1 min - 1 for free and NP-bound enzyme, respectively.

Fanconi anemia proteins in telomere maintenance.

PubMed

Sarkar, Jaya; Liu, Yie

2016-07-01

Mammalian chromosome ends are protected by nucleoprotein structures called telomeres. Telomeres ensure genome stability by preventing chromosome termini from being recognized as DNA damage. Telomere length homeostasis is inevitable for telomere maintenance because critical shortening or over-lengthening of telomeres may lead to DNA damage response or delay in DNA replication, and hence genome instability. Due to their repetitive DNA sequence, unique architecture, bound shelterin proteins, and high propensity to form alternate/secondary DNA structures, telomeres are like common fragile sites and pose an inherent challenge to the progression of DNA replication, repair, and recombination apparatus. It is conceivable that longer the telomeres are, greater is the severity of such challenges. Recent studies have linked excessively long telomeres with increased tumorigenesis. Here we discuss telomere abnormalities in a rare recessive chromosomal instability disorder called Fanconi Anemia and the role of the Fanconi Anemia pathway in telomere biology. Reports suggest that Fanconi Anemia proteins play a role in maintaining long telomeres, including processing telomeric joint molecule intermediates. We speculate that ablation of the Fanconi Anemia pathway would lead to inadequate aberrant structural barrier resolution at excessively long telomeres, thereby causing replicative burden on the cell. Published by Elsevier B.V.

Modeling ligand recognition at the P2Y12 receptor in light of X-ray structural information

NASA Astrophysics Data System (ADS)

Paoletta, Silvia; Sabbadin, Davide; von Kügelgen, Ivar; Hinz, Sonja; Katritch, Vsevolod; Hoffmann, Kristina; Abdelrahman, Aliaa; Straßburger, Jens; Baqi, Younis; Zhao, Qiang; Stevens, Raymond C.; Moro, Stefano; Müller, Christa E.; Jacobson, Kenneth A.

2015-08-01

The G protein-coupled P2Y12 receptor (P2Y12R) is an important antithrombotic target and of great interest for pharmaceutical discovery. Its recently solved, highly divergent crystallographic structures in complex either with nucleotides (full or partial agonist) or with a nonnucleotide antagonist raise the question of which structure is more useful to understand ligand recognition. Therefore, we performed extensive molecular modeling studies based on these structures and mutagenesis, to predict the binding modes of major classes of P2Y12R ligands previously reported. Various nucleotide derivatives docked readily to the agonist-bound P2Y12R, but uncharged nucleotide-like antagonist ticagrelor required a hybrid receptor resembling the agonist-bound P2Y12R except for the top portion of TM6. Supervised molecular dynamics (SuMD) of ticagrelor binding indicated interactions with the extracellular regions of P2Y12R, defining possible meta-binding sites. Ureas, sulfonylureas, sulfonamides, anthraquinones and glutamic acid piperazines docked readily to the antagonist-bound P2Y12R. Docking dinucleotides at both agonist- and antagonist-bound structures suggested interactions with two P2Y12R pockets. Thus, our structure-based approach consistently rationalized the main structure-activity relationships within each ligand class, giving useful information for designing improved ligands.

Interacting quantum walkers: two-body bosonic and fermionic bound states

NASA Astrophysics Data System (ADS)

Krapivsky, P. L.; Luck, J. M.; Mallick, K.

2015-11-01

We investigate the dynamics of bound states of two interacting particles, either bosons or fermions, performing a continuous-time quantum walk on a one-dimensional lattice. We consider the situation where the distance between both particles has a hard bound, and the richer situation where the particles are bound by a smooth confining potential. The main emphasis is on the velocity characterizing the ballistic spreading of these bound states, and on the structure of the asymptotic distribution profile of their center-of-mass coordinate. The latter profile generically exhibits many internal fronts.

Electronic structure of the metal center in the Cd(2+), Zn(2+), and Cu(2+) substituted forms of KDO8P synthase: implications for catalysis.

PubMed

Kona, Fathima; Tao, Peng; Martin, Philip; Xu, Xingjue; Gatti, Domenico L

2009-04-28

Aquifex aeolicus 3-deoxy-d-manno-octulosonate 8-phosphate synthase (KDO8PS) is active with a variety of different divalent metal ions bound in the active site. The Cd(2+), Zn(2+), and Cu(2+) substituted enzymes display similar values of k(cat) and similar dependence of K(m)(PEP) and K(m)(A5P) on both substrate and product concentrations. However, the flux-control coefficients for some of the catalytically relevant reaction steps are different in the presence of Zn(2+) or Cu(2+), suggesting that the type of metal bound in the active site affects the behavior of the enzyme in vivo. The type of metal also affects the rate of product release in the crystal environment. For example, the crystal structure of the Cu(2+) enzyme incubated with phosphoenolpyruvate (PEP) and arabinose 5-phosphate (A5P) shows the formed product, 3-deoxy-d-manno-octulosonate 8-phosphate (KDO8P), still bound in the active site in its linear conformation. This observation completes our structural studies of the condensation reaction, which altogether have provided high-resolution structures for the reactants, the intermediate, and the product bound forms of KDO8PS. The crystal structures of the Cd(2+), Zn(2+), and Cu(2+) substituted enzymes show four residues (Cys-11, His-185, Glu-222, and Asp-233) and a water molecule as possible metal ligands. Combined quantum mechanics/molecular mechanics (QM/MM) geometry optimizations reveal that the metal centers have a delocalized electronic structure, and that their true geometry is square pyramidal for Cd(2+) and Zn(2+) and distorted octahedral or distorted tetrahedral for Cu(2+). These geometries are different from those obtained by QM optimization in the gas phase (tetrahedral for Cd(2+) and Zn(2+), distorted tetrahedral for Cu(2+)) and may represent conformations of the metal center that minimize the reorganization energy between the substrate-bound and product-bound states. The QM/MM calculations also show that when only PEP is bound to the enzyme the electronic structure of the metal center is optimized to prevent a wasteful reaction of PEP with water.

Pre-polishing on a CNC platform with bound abrasive contour tools

NASA Astrophysics Data System (ADS)

Schoeffer, Adrienne E.

2003-05-01

Deterministic micorgrinding (DMG) of optical glasses and ceramics is the commercial manufacturing process of choice to shape glass surfaces prior to final finishing. This process employs rigid bound matrix diamond tooling resulting in surface roughness values of 3-51.tm peak to valley and 100-400nm rms, as well as mid-spatial frequency tool marks that require subsequent removal in secondary finishing steps. The ability to pre-polish optical surfaces within the grinding platform would reduce final finishing process times. Bound abrasive contour wheels containing cerium oxide, alumina or zirconia abrasives were constructed with an epoxy matrix. The effects of abrasive type, composition, and erosion promoters were examined for tool hardness (Shore D), and tested with commercial optical glasses in an OptiproTM CNC grinding platform. Metrology protocols were developed to examine tool wear and subsequent surface roughness. Work is directed to demonstrating effective material removal, improved surface roughness and cutter mark removal.

Prepolishing on a CNC platform with bound abrasive contour tools

NASA Astrophysics Data System (ADS)

Schoeffler, Adrienne E.; Gregg, Leslie L.; Schoen, John M.; Fess, Edward M.; Hakiel, Michael; Jacobs, Stephen D.

2003-05-01

Deterministic microgrinding (DMG) of optical glasses and ceramics is the commercial manufacturing process of choice to shape glass surfaces prior to final finishing. This process employs rigid bound matrix diamond tooling resulting in surface roughness values of 3-5μm peak to valley and 100-400nm rms, as well as mid-spatial frequency tool marks that require subsequent removal in secondary finishing steps. The ability to pre-polish optical surfaces within the grinding platform would reduce final finishing process times. Bound abrasive contour wheels containing cerium oxide, alumina or zirconia abrasives were constructed with an epoxy matrix. The effects of abrasive type, composition, and erosion promoters were examined for tool hardness (Shore D), and tested with commercial optical glasses in an Optipro CNC grinding platform. Metrology protocols were developed to examine tool wear and subsequent surface roughness. Work is directed to demonstrating effective material removal, improved surface roughness and cutter mark removal.

Lifetime of a Chemically Bound Helium Compound

NASA Technical Reports Server (NTRS)

Chaban, Galina M.; Lundell, Jan; Gerber, R. Benny; Kwak, Dochan (Technical Monitor)

2001-01-01

The rare-gas atoms are chemically inert, to an extent unique among all elements. This is due to the stable electronic structure of the atoms. Stable molecules with chemically bound rare-gas atoms are, however, known. A first such compound, XePtF6, W2S prepared in 1962 and since then a range of molecules containing radon, xenon and krypton have been obtained. Most recently, a first stable chemically bound compound of argon was prepared, leaving neon and helium as the only elements for which stable chemically bound molecules are not yet known. Electronic structure calculations predict that a metastable species HHeF exists, but significance of the result depends on the unknown lifetime. Here we report quantum dynamics calculations of the lifetime of HHeF, using accurate interactions computed from electronic structure theory. HHeF is shown to disintegrate by tunneling through energy barriers into He + HF and H + He + F the first channel greatly dominating. The lifetime of HHeF is more than 120 picoseconds, that of DHeF is 14 nanoseconds. The relatively long lifetimes are encouraging for the preparation prospects of this first chemically bound helium compound.

The structure of the antimicrobial active center of lactoferricin B bound to sodium dodecyl sulfate micelles.

PubMed

Schibli, D J; Hwang, P M; Vogel, H J

1999-03-12

Lactoferricin B (LfcinB) is a 25-residue antimicrobial peptide released from bovine lactoferrin upon pepsin digestion. The antimicrobial center of LfcinB consists of six residues (RRWQWR-NH2), and it possesses similar bactericidal activity to LfcinB. The structure of the six-residue peptide bound to sodium dodecyl sulfate (SDS) micelles has been determined by NMR spectroscopy and molecular dynamics refinement. The peptide adopts a well defined amphipathic structure when bound to SDS micelles with the Trp sidechains separated from the Arg residues. Additional evidence demonstrates that the peptide is oriented in the micelle such that the Trp residues are more deeply buried in the micelle than the Arg and Gln residues.

NES consensus redefined by structures of PKI-type and Rev-type nuclear export signals bound to CRM1.

PubMed

Güttler, Thomas; Madl, Tobias; Neumann, Piotr; Deichsel, Danilo; Corsini, Lorenzo; Monecke, Thomas; Ficner, Ralf; Sattler, Michael; Görlich, Dirk

2010-11-01

Classic nuclear export signals (NESs) confer CRM1-dependent nuclear export. Here we present crystal structures of the RanGTP-CRM1 complex alone and bound to the prototypic PKI or HIV-1 Rev NESs. These NESs differ markedly in the spacing of their key hydrophobic (Φ) residues, yet CRM1 recognizes them with the same rigid set of five Φ pockets. The different Φ spacings are compensated for by different conformations of the bound NESs: in the case of PKI, an α-helical conformation, and in the case of Rev, an extended conformation with a critical proline docking into a Φ pocket. NMR analyses of CRM1-bound and CRM1-free PKI NES suggest that CRM1 selects NES conformers that pre-exist in solution. Our data lead to a new structure-based NES consensus, and explain why NESs differ in their affinities for CRM1 and why supraphysiological NESs bind the exportin so tightly.

Strong Dependence of Hydration State of F-Actin on the Bound Mg(2+)/Ca(2+) Ions.

PubMed

Suzuki, Makoto; Imao, Asato; Mogami, George; Chishima, Ryotaro; Watanabe, Takahiro; Yamaguchi, Takaya; Morimoto, Nobuyuki; Wazawa, Tetsuichi

2016-07-21

Understanding of the hydration state is an important issue in the chemomechanical energetics of versatile biological functions of polymerized actin (F-actin). In this study, hydration-state differences of F-actin by the bound divalent cations are revealed through precision microwave dielectric relaxation (DR) spectroscopy. G- and F-actin in Ca- and Mg-containing buffer solutions exhibit dual hydration components comprising restrained water with DR frequency f2 (fw). The hydration state of F-actin is strongly dependent on the ionic composition. In every buffer tested, the HMW signal Dhyme (≡ (f1 - fw)δ1/(fwδw)) of F-actin is stronger than that of G-actin, where δw is DR-amplitude of bulk solvent and δ1 is that of HMW in a fixed-volume ellipsoid containing an F-actin and surrounding water in solution. Dhyme value of F-actin in Ca2.0-buffer (containing 2 mM Ca(2+)) is markedly higher than in Mg2.0-buffer (containing 2 mM Mg(2+)). Moreover, in the presence of 2 mM Mg(2+), the hydration state of F-actin is changed by adding a small fraction of Ca(2+) (∼0.1 mM) and becomes closer to that of the Ca-bound form in Ca2.0-buffer. This is consistent with the results of the partial specific volume and the Cotton effect around 290 nm in the CD spectra, indicating a change in the tertiary structure and less apparent change in the secondary structure of actin. The number of restrained water molecules per actin (N2) is estimated to be 1600-2100 for Ca2.0- and F-buffer and ∼2500 for Mg2.0-buffer at 10-15 °C. These numbers are comparable to those estimated from the available F-actin atomic structures as in the first water layer. The number of HMW molecules is roughly explained by the volume between the equipotential surface of -kT/2e and the first water layer of the actin surface by solving the Poisson-Boltzmann equation using UCSF Chimera.

Mirror energy difference and the structure of loosely bound proton-rich nuclei around A =20

NASA Astrophysics Data System (ADS)

Yuan, Cenxi; Qi, Chong; Xu, Furong; Suzuki, Toshio; Otsuka, Takaharu

2014-04-01

The properties of loosely bound proton-rich nuclei around A =20 are investigated within the framework of the nuclear shell model. In these nuclei, the strength of the effective interactions involving the loosely bound proton s1/2 orbit is significantly reduced in comparison with that of those in their mirror nuclei. We evaluate the reduction of the effective interaction by calculating the monopole-based-universal interaction (VMU) in the Woods-Saxon basis. The shell-model Hamiltonian in the sd shell, such as USD, can thus be modified to reproduce the binding energies and energy levels of the weakly bound proton-rich nuclei around A =20. The effect of the reduction of the effective interaction on the structure and decay properties of these nuclei is also discussed.

The engine of microtubule dynamics comes into focus.

PubMed

Mitchison, T J

2014-05-22

In this issue, Alushin et al. report high-resolution structures of three states of the microtubule lattice: GTP-bound, which is stable to depolymerization; unstable GDP-bound; and stable Taxol and GDP-bound. By comparing these structures at near-atomic resolution, they are able to propose a detailed model for how GTP hydrolysis destabilizes the microtubule and thus powers dynamic instability and chromosome movement. Destabilization of cytoskeleton filaments by nucleotide hydrolysis is an important general principle in cell dynamics, and this work represents a major step forward on a problem with a long history. Copyright © 2014 Elsevier Inc. All rights reserved.

The Phosphorylation of Vascular Endothelial Growth Factor Receptor-2 (VEGFR-2) by Engineered Surfaces with Electrostatically or Covalently Immobilized VEGF

PubMed Central

Anderson, Sean M.; Chen, Tom T.; Iruela-Arispe, M. Luisa; Segura, Tatiana

2010-01-01

Growth factors are a class of signaling proteins that direct cell fate through interaction with cell surface receptors. Although a myriad of possible cell fates stem from a growth factor binding to its receptor, the signaling cascades that result in one fate over another are still being elucidated. One possible mechanism by which nature modulates growth factor signaling is through the method of presentation of the growth factor – soluble or immobilized (matrix bound). Here we present the methodology to study signaling of soluble versus immobilized VEGF through VEGFR-2. We have designed a strategy to covalently immobilize VEGF using its heparin-binding domain to orient the molecule (bind) and a secondary functional group to mediate covalent binding (lock). This bind-and-lock approach aims to allow VEGF to assume a bioactive orientation before covalent immobilization. Surface plasmon resonance (SPR) demonstrated heparin and VEGF binding with surface densities of 60 ng/cm2 and 100 pg/cm2, respectively. ELISA experiments confirmed VEGF surface density and showed that electrostatically bound VEGF releases in cell medium and heparin solutions while covalently bound VEGF remains immobilized. Electrostatically bound VEGF and covalently bound VEGF phosphorylate VEGFR-2 in both VEGFR-2 transfected cells and VEGFR-2 endogenously producing cells. HUVECs plated on VEGF functionalized surfaces showed different morphologies between surface-bound VEGF and soluble VEGF. The surfaces synthesized in these studies allow for the study of VEGF/VEGFR-2 signaling induced by covalently bound, electrostatically bound, and soluble VEGF and may provide further insight into the design of materials for the generation of a mature and stable vasculature. PMID:19540581

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

Liu Chao; Hong Fashui; Wu Kang

Neodymium (Nd), as a member of rare earth elements, proved to enhance the photosynthesis rate and organic substance accumulation of spinach through the increase in carboxylation activity of Rubisco. Although the oxygenase activity of spinach Rubisco was slightly changed with the Nd{sup 3+} treatment, the specific factor of Rubisco was greatly increased. It was partially due to the promotion of Rubisco activase (R-A) activity but mainly to the formation of Rubisco-Rubisco activase super-complex, a heavier molecular mass protein (about 1200 kD) comprising both Rubisco and Rubisco activase. This super-complex was found during the extraction procedure of Rubisco by the gelmore » electrophoresis and Western-blot studies. The formation of Rubisco-R-A super-complex suggested that the secondary structure of the protein purified from the Nd{sup 3+}-treated spinach was different from that of the control. Extended X-ray absorption fine structure study of the 'Rubisco' purified from the Nd{sup 3+}-treated spinach revealed that Nd was bound with four oxygen atoms and two sulfur atoms of amino acid residues at the Nd-O and Nd-S bond lengths of 2.46 and 2.89 A, respectively.« less

Cu(II) potentiation of Alzheimer Abeta1-40 cytotoxicity and transition on its secondary structure.

PubMed

Dai, Xue-Ling; Sun, Ya-Xuan; Jiang, Zhao-Feng

2006-11-01

Mounting evidence has shown that dyshomeostasis of the redox-active biometals such as Cu and Fe can lead to oxidative stress, which plays a key role in the neuropathology of Alzheimer' disease (AD). Here we demonstrate that with the formation of Cu(II).beta1-40 complexes, copper markedly potentiates the neurotoxicity exhibited by beta-amyloid peptide (Ab). A greater amount of hydrogen peroxide was released when Cu(II).beta1-40 complexes was added to the xanthine oxidase/xanthine system detected by potassium iodide spectrophotometry. Copper bound to Abeta1-40 was observed by electron paramagnetic resonance (EPR) spectroscopy. Circular dichroism (CD) studies indicated that copper chelation could cause a structural transition of Abeta. The addition of copper to Ab introduced an increase on beta-sheet as well as alpha-helix, which may be responsible for the aggregation of Abeta. We hypothesized that Abeta aggregation induced by copper may be responsible for local injury in AD. The interaction between Cu(2+) and Ab also provides a possible mechanism for the enrichment of metal ions in amyloid plaques in the AD brain.

Folding cooperativity in a three-stranded beta-sheet model.

PubMed

Roe, Daniel R; Hornak, Viktor; Simmerling, Carlos

2005-09-16

The thermodynamic behavior of a previously designed three-stranded beta-sheet was studied via several microseconds of standard and replica exchange molecular dynamics simulations. The system is shown to populate at least four thermodynamic minima, including two partially folded states in which only a single hairpin is formed. Simulated melting curves show different profiles for the C and N-terminal hairpins, consistent with differences in secondary structure content in published NMR and CD/FTIR measurements, which probed different regions of the chain. Individual beta-hairpins that comprise the three-stranded beta-sheet are observed to form cooperatively. Partial folding cooperativity between the component hairpins is observed, and good agreement between calculated and experimental values quantifying this cooperativity is obtained when similar analysis techniques are used. However, the structural detail in the ensemble of conformations sampled in the simulations permits a more direct analysis of this cooperativity than has been performed on the basis of experimental data. The results indicate the actual folding cooperativity perpendicular to strand direction is significantly larger than the lower bound obtained previously.

Folding cooperativity in a 3-stranded β-sheet model

PubMed Central

Roe, Daniel R.; Hornak, Viktor

2015-01-01

Summary The thermodynamic behavior of a previously designed three-stranded β-sheet was studied via several µs of standard and replica exchange molecular dynamics simulations. The system is shown to populate at least four thermodynamic minima, including 2 partially folded states in which only a single hairpin is formed. Simulated melting curves show different profiles for the C and N-terminal hairpins, consistent with differences in secondary structure content in published NMR and CD/FTIR measurements, which probed different regions of the chain. Individual β-hairpins that comprise the 3-stranded β-sheet are observed to form cooperatively. Partial folding cooperativity between the component hairpins is observed, and good agreement between calculated and experimental values quantifying this cooperativity is obtained when similar analysis techniques are used. However, the structural detail in the ensemble of conformations sampled in the simulations permits a more direct analysis of this cooperatively than has been performed based on experimental data. The results indicate the actual folding cooperativity perpendicular to strand direction is significantly larger than the lower bound obtained previously. PMID:16095612

Elucidating Oxygen Reduction Active Sites in Pyrolyzed Metal-Nitrogen Coordinated Non-Precious-Metal Electrocatalyst Systems.

PubMed

Tylus, Urszula; Jia, Qingying; Strickland, Kara; Ramaswamy, Nagappan; Serov, Alexey; Atanassov, Plamen; Mukerjee, Sanjeev

2014-05-01

Detailed understanding of the nature of the active centers in non-precious-metal-based electrocatalyst, and their role in oxygen reduction reaction (ORR) mechanistic pathways will have a profound effect on successful commercialization of emission-free energy devices such as fuel cells. Recently, using pyrolyzed model structures of iron porphyrins, we have demonstrated that a covalent integration of the Fe-N x sites into π-conjugated carbon basal plane modifies electron donating/withdrawing capability of the carbonaceous ligand, consequently improving ORR activity. Here, we employ a combination of in situ X-ray spectroscopy and electrochemical methods to identify the various structural and functional forms of the active centers in non-heme Fe/N/C catalysts. Both methods corroboratively confirm the single site 2e - × 2e - mechanism in alkaline media on the primary Fe 2+ -N 4 centers and the dual-site 2e - × 2e - mechanism in acid media with the significant role of the surface bound coexisting Fe/Fe x O y nanoparticles (NPs) as the secondary active sites.

Characterization of flavonoid-protein interactions using fluorescence spectroscopy: Binding of pelargonidin to dairy proteins.

PubMed

Arroyo-Maya, Izlia J; Campos-Terán, José; Hernández-Arana, Andrés; McClements, David Julian

2016-12-15

In this study, the interaction between the flavonoid pelargonidin and dairy proteins: β-lactoglobulin (β-LG), whey protein (WPI), and caseinate (CAS) was investigated. Fluorescence experiments demonstrated that pelargonidin quenched milk proteins fluorescence strongly. However, the protein secondary structure was not significantly affected by pelargonidin, as judged from far-UV circular dichroism. Analysis of fluorescence data indicated that pelargonidin-induced quenching does not arise from a dynamical mechanism, but instead is due to protein-ligand binding. Therefore, quenching data were analyzed using the model of independent binding sites. Both β-LG and CAS, but not WPI, showed hyperbolic binding isotherms indicating that these proteins firmly bound pelargonidin at both pH 7.0 and 3.0 (binding constants ca. 1.0×10(5) at 25.0°C). To investigate the underlying thermodynamics, binding constants were determined at 25.0, 35.0, and 45.0°C. These results pointed to binding processes that depend on the structural conformation of the milk proteins. Copyright © 2016 Elsevier Ltd. All rights reserved.

Elucidating Oxygen Reduction Active Sites in Pyrolyzed Metal–Nitrogen Coordinated Non-Precious-Metal Electrocatalyst Systems

PubMed Central

2015-01-01

Detailed understanding of the nature of the active centers in non-precious-metal-based electrocatalyst, and their role in oxygen reduction reaction (ORR) mechanistic pathways will have a profound effect on successful commercialization of emission-free energy devices such as fuel cells. Recently, using pyrolyzed model structures of iron porphyrins, we have demonstrated that a covalent integration of the Fe–Nx sites into π-conjugated carbon basal plane modifies electron donating/withdrawing capability of the carbonaceous ligand, consequently improving ORR activity. Here, we employ a combination of in situ X-ray spectroscopy and electrochemical methods to identify the various structural and functional forms of the active centers in non-heme Fe/N/C catalysts. Both methods corroboratively confirm the single site 2e– × 2e– mechanism in alkaline media on the primary Fe2+–N4 centers and the dual-site 2e– × 2e– mechanism in acid media with the significant role of the surface bound coexisting Fe/FexOy nanoparticles (NPs) as the secondary active sites. PMID:24817921

Nonlinear effects in the bounded dust-vortex flow in plasma

NASA Astrophysics Data System (ADS)

Laishram, Modhuchandra; Sharma, Devendra; Chattopdhyay, Prabal K.; Kaw, Predhiman K.

2017-03-01

The vortex structures in a cloud of electrically suspended dust in a streaming plasma constitutes a driven system with a rich nonlinear flow regime. Experimentally recovered toroidal formations of this system have motivated study of its volumetrically driven-dissipative vortex flow dynamics using two-dimensional hydrodynamics in the incompressible Navier-Stokes regime. Nonlinear equilibrium solutions are obtained for this system where a nonuniformly driven two-dimensional dust flow exhibits distinct regions of localized accelerations and strong friction caused by stationary fluids at the confining boundaries resisting the dust flow. In agreement with observations in experiments, it is demonstrated that the nonlinear effects appear in the limit of small viscosity, where the primary vortices form scaling with the most dominant spatial scales of the domain topology and develop separated virtual boundaries along their periphery. This separation is triggered beyond a critical dust viscosity that signifies a structural bifurcation. Emergence of uniform vorticity core and secondary vortices with a newer level of identical dynamics highlights the applicability of the studied dynamics to gigantic vortex flows, such as the Jovian great red spot, to microscopic biophysical intracellular activity.

Zebrafish AID is capable of deaminating methylated deoxycytidines

PubMed Central

Abdouni, Hala; King, Justin J.; Suliman, Mussa; Quinlan, Matthew; Fifield, Heather; Larijani, Mani

2013-01-01

Activation-induced cytidine deaminase (AID) deaminates deoxycytidine (dC) to deoxyuracil (dU) at immunoglobulin loci in B lymphocytes to mediate secondary antibody diversification. Recently, AID has been proposed to also mediate epigenetic reprogramming by demethylating methylated cytidines (mC) possibly through deamination. AID overexpression in zebrafish embryos was shown to promote genome demethylation through G:T lesions, implicating a deamination-dependent mechanism. We and others have previously shown that mC is a poor substrate for human AID. Here, we examined the ability of bony fish AID to deaminate mC. We report that zebrafish AID was unique among all orthologs in that it efficiently deaminates mC. Analysis of domain-swapped and mutant AID revealed that mC specificity is independent of the overall high-catalytic efficiency of zebrafish AID. Structural modeling with or without bound DNA suggests that efficient deamination of mC by zebrafish AID is likely not due to a larger catalytic pocket allowing for better fit of mC, but rather because of subtle differences in the flexibility of its structure. PMID:23585279

Resolvent analysis of shear flows using One-Way Navier-Stokes equations

NASA Astrophysics Data System (ADS)

Rigas, Georgios; Schmidt, Oliver; Towne, Aaron; Colonius, Tim

2017-11-01

For three-dimensional flows, questions of stability, receptivity, secondary flows, and coherent structures require the solution of large partial-derivative eigenvalue problems. Reduced-order approximations are thus required for engineering prediction since these problems are often computationally intractable or prohibitively expensive. For spatially slowly evolving flows, such as jets and boundary layers, the One-Way Navier-Stokes (OWNS) equations permit a fast spatial marching procedure that results in a huge reduction in computational cost. Here, an adjoint-based optimization framework is proposed and demonstrated for calculating optimal boundary conditions and optimal volumetric forcing. The corresponding optimal response modes are validated against modes obtained in terms of global resolvent analysis. For laminar base flows, the optimal modes reveal modal and non-modal transition mechanisms. For turbulent base flows, they predict the evolution of coherent structures in a statistical sense. Results from the application of the method to three-dimensional laminar wall-bounded flows and turbulent jets will be presented. This research was supported by the Office of Naval Research (N00014-16-1-2445) and Boeing Company (CT-BA-GTA-1).

RNA-Puzzles Round III: 3D RNA structure prediction of five riboswitches and one ribozyme

PubMed Central

Biesiada, Marcin; Boniecki, Michał J.; Chou, Fang-Chieh; Ferré-D'Amaré, Adrian R.; Das, Rhiju; Dunin-Horkawicz, Stanisław; Geniesse, Caleb; Kappel, Kalli; Kladwang, Wipapat; Krokhotin, Andrey; Łach, Grzegorz E.; Major, François; Mann, Thomas H.; Pachulska-Wieczorek, Katarzyna; Patel, Dinshaw J.; Piccirilli, Joseph A.; Popenda, Mariusz; Purzycka, Katarzyna J.; Ren, Aiming; Rice, Greggory M.; Santalucia, John; Tandon, Arpit; Trausch, Jeremiah J.; Wang, Jian; Weeks, Kevin M.; Williams, Benfeard; Xiao, Yi; Zhang, Dong; Zok, Tomasz

2017-01-01

RNA-Puzzles is a collective experiment in blind 3D RNA structure prediction. We report here a third round of RNA-Puzzles. Five puzzles, 4, 8, 12, 13, 14, all structures of riboswitch aptamers and puzzle 7, a ribozyme structure, are included in this round of the experiment. The riboswitch structures include biological binding sites for small molecules (S-adenosyl methionine, cyclic diadenosine monophosphate, 5-amino 4-imidazole carboxamide riboside 5′-triphosphate, glutamine) and proteins (YbxF), and one set describes large conformational changes between ligand-free and ligand-bound states. The Varkud satellite ribozyme is the most recently solved structure of a known large ribozyme. All puzzles have established biological functions and require structural understanding to appreciate their molecular mechanisms. Through the use of fast-track experimental data, including multidimensional chemical mapping, and accurate prediction of RNA secondary structure, a large portion of the contacts in 3D have been predicted correctly leading to similar topologies for the top ranking predictions. Template-based and homology-derived predictions could predict structures to particularly high accuracies. However, achieving biological insights from de novo prediction of RNA 3D structures still depends on the size and complexity of the RNA. Blind computational predictions of RNA structures already appear to provide useful structural information in many cases. Similar to the previous RNA-Puzzles Round II experiment, the prediction of non-Watson–Crick interactions and the observed high atomic clash scores reveal a notable need for an algorithm of improvement. All prediction models and assessment results are available at http://ahsoka.u-strasbg.fr/rnapuzzles/. PMID:28138060

CSI 3.0: a web server for identifying secondary and super-secondary structure in proteins using NMR chemical shifts

PubMed Central

Hafsa, Noor E.; Arndt, David; Wishart, David S.

2015-01-01

The Chemical Shift Index or CSI 3.0 (http://csi3.wishartlab.com) is a web server designed to accurately identify the location of secondary and super-secondary structures in protein chains using only nuclear magnetic resonance (NMR) backbone chemical shifts and their corresponding protein sequence data. Unlike earlier versions of CSI, which only identified three types of secondary structure (helix, β-strand and coil), CSI 3.0 now identifies total of 11 types of secondary and super-secondary structures, including helices, β-strands, coil regions, five common β-turns (type I, II, I′, II′ and VIII), β hairpins as well as interior and edge β-strands. CSI 3.0 accepts experimental NMR chemical shift data in multiple formats (NMR Star 2.1, NMR Star 3.1 and SHIFTY) and generates colorful CSI plots (bar graphs) and secondary/super-secondary structure assignments. The output can be readily used as constraints for structure determination and refinement or the images may be used for presentations and publications. CSI 3.0 uses a pipeline of several well-tested, previously published programs to identify the secondary and super-secondary structures in protein chains. Comparisons with secondary and super-secondary structure assignments made via standard coordinate analysis programs such as DSSP, STRIDE and VADAR on high-resolution protein structures solved by X-ray and NMR show >90% agreement between those made with CSI 3.0. PMID:25979265

Crystal structure of the adenosine A 2A receptor bound to an antagonist reveals a potential allosteric pocket

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

Sun, Bingfa; Bachhawat, Priti; Chu, Matthew Ling-Hon

The adenosine A2A receptor (A2AR) has long been implicated in cardiovascular disorders. As more selective A2AR ligands are being identified, its roles in other disorders, such as Parkinson’s disease, are starting to emerge, and A2AR antagonists are important drug candidates for nondopaminergic anti-Parkinson treatment. Here we report the crystal structure of A2A receptor bound to compound 1 (Cmpd-1), a novel A2AR/N-methyl D-aspartate receptor subtype 2B (NR2B) dual antagonist and potential anti-Parkinson candidate compound, at 3.5 Å resolution. The A2A receptor with a cytochrome b562-RIL (BRIL) fusion (A2AR–BRIL) in the intracellular loop 3 (ICL3) was crystallized in detergent micelles using vapor-phasemore » diffusion. Whereas A2AR–BRIL bound to the antagonist ZM241385 has previously been crystallized in lipidic cubic phase (LCP), structural differences in the Cmpd-1–bound A2AR–BRIL prevented formation of the lattice observed with the ZM241385–bound receptor. The crystals grew with a type II crystal lattice in contrast to the typical type I packing seen from membrane protein structures crystallized in LCP. Cmpd-1 binds in a position that overlaps with the native ligand adenosine, but its methoxyphenyl group extends to an exosite not previously observed in other A2AR structures. Structural analysis revealed that Cmpd-1 binding results in the unique conformations of two tyrosine residues, Tyr91.35 and Tyr2717.36, which are critical for the formation of the exosite. The structure reveals insights into antagonist binding that are not observed in other A2AR structures, highlighting flexibility in the binding pocket that may facilitate the development of A2AR-selective compounds for the treatment of Parkinson’s disease.« less

Testing ΛCDM cosmology at turnaround: where to look for violations of the bound?

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

Tanoglidis, D.; Pavlidou, V.; Tomaras, T.N., E-mail: dtanogl@physics.uoc.gr, E-mail: pavlidou@physics.uoc.gr, E-mail: tomaras@physics.uoc.gr

In ΛCDM cosmology, structure formation is halted shortly after dark energy dominates the mass/energy budget of the Universe. A manifestation of this effect is that in such a cosmology the turnaround radius—the non-expanding mass shell furthest away from the center of a structure— has an upper bound. Recently, a new, local, test for the existence of dark energy in the form of a cosmological constant was proposed based on this turnaround bound. Before designing an experiment that, through high-precision determination of masses and —independently— turnaround radii, will challenge ΛCDM cosmology, we have to answer two important questions: first, when turnaround-scalemore » structures are predicted to be close enough to their maximum size, so that a possible violation of the bound may be observable. Second, which is the best mass scale to target for possible violations of the bound. These are the questions we address in the present work. Using the Press-Schechter formalism, we find that turnaround structures have in practice already stopped forming, and consequently, the turnaround radius of structures must be very close to the maximum value today. We also find that the mass scale of ∼ 10{sup 13} M{sub ⊙} characterizes the turnaround structures that start to form in a statistically important number density today —and even at an infinite time in the future, since structure formation has almost stopped. This mass scale also separates turnaround structures with qualitatively different cosmological evolution: smaller structures are no longer readjusting their mass distribution inside the turnaround scale, they asymptotically approach their ultimate abundance from higher values, and they are common enough to have, at some epoch, experienced major mergers with structures of comparable mass; larger structures exhibit the opposite behavior. We call this mass scale the transitional mass scale and we argue that it is the optimal for the purpose outlined above. As a corollary, we explain the different accretion behavior of small and larger structures observed in already conducted numerical simulations.« less

Structural and biochemical characterisation of Archaeoglobus fulgidus esterase reveals a bound CoA molecule in the vicinity of the active site.

PubMed

Sayer, Christopher; Finnigan, William; Isupov, Michail N; Levisson, Mark; Kengen, Servé W M; van der Oost, John; Harmer, Nicholas J; Littlechild, Jennifer A

2016-05-10

A new carboxyl esterase, AF-Est2, from the hyperthermophilic archaeon Archaeoglobus fulgidus has been cloned, over-expressed in Escherichia coli and biochemically and structurally characterized. The enzyme has high activity towards short- to medium-chain p-nitrophenyl carboxylic esters with optimal activity towards the valerate ester. The AF-Est2 has good solvent and pH stability and is very thermostable, showing no loss of activity after incubation for 30 min at 80 °C. The 1.4 Å resolution crystal structure of AF-Est2 reveals Coenzyme A (CoA) bound in the vicinity of the active site. Despite the presence of CoA bound to the AF-Est2 this enzyme has no CoA thioesterase activity. The pantetheine group of CoA partially obstructs the active site alcohol pocket suggesting that this ligand has a role in regulation of the enzyme activity. A comparison with closely related α/β hydrolase fold enzyme structures shows that the AF-Est2 has unique structural features that allow CoA binding. A comparison of the structure of AF-Est2 with the human carboxyl esterase 1, which has CoA thioesterase activity, reveals that CoA is bound to different parts of the core domain in these two enzymes and approaches the active site from opposite directions.

Structure of ATP-Bound Human ATP:Cobalamin Adenosyltransferase

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

Schubert,H.; Hill, C.

Mutations in the gene encoding human ATP:cobalamin adenosyltransferase (hATR) can result in the metabolic disorder known as methylmalonic aciduria (MMA). This enzyme catalyzes the final step in the conversion of cyanocobalamin (vitamin B{sub 12}) to the essential human cofactor adenosylcobalamin. Here we present the 2.5 {angstrom} crystal structure of ATP bound to hATR refined to an R{sub free} value of 25.2%. The enzyme forms a tightly associated trimer, where the monomer comprises a five-helix bundle and the active sites lie on the subunit interfaces. Only two of the three active sites within the trimer contain the bound ATP substrate, therebymore » providing examples of apo- and substrate-bound-active sites within the same crystal structure. Comparison of the empty and occupied sites indicates that twenty residues at the enzyme's N-terminus become ordered upon binding of ATP to form a novel ATP-binding site and an extended cleft that likely binds cobalamin. The structure explains the role of 20 invariant residues; six are involved in ATP binding, including Arg190, which hydrogen bonds to ATP atoms on both sides of the scissile bond. Ten of the hydrogen bonds are required for structural stability, and four are in positions to interact with cobalamin. The structure also reveals how the point mutations that cause MMA are deficient in these functions.« less

Structural and biochemical characterisation of Archaeoglobus fulgidus esterase reveals a bound CoA molecule in the vicinity of the active site

PubMed Central

Sayer, Christopher; Finnigan, William; Isupov, Michail N.; Levisson, Mark; Kengen, Servé W. M.; van der Oost, John; Harmer, Nicholas J.; Littlechild, Jennifer A.

2016-01-01

A new carboxyl esterase, AF-Est2, from the hyperthermophilic archaeon Archaeoglobus fulgidus has been cloned, over-expressed in Escherichia coli and biochemically and structurally characterized. The enzyme has high activity towards short- to medium-chain p-nitrophenyl carboxylic esters with optimal activity towards the valerate ester. The AF-Est2 has good solvent and pH stability and is very thermostable, showing no loss of activity after incubation for 30 min at 80 °C. The 1.4 Å resolution crystal structure of AF-Est2 reveals Coenzyme A (CoA) bound in the vicinity of the active site. Despite the presence of CoA bound to the AF-Est2 this enzyme has no CoA thioesterase activity. The pantetheine group of CoA partially obstructs the active site alcohol pocket suggesting that this ligand has a role in regulation of the enzyme activity. A comparison with closely related α/β hydrolase fold enzyme structures shows that the AF-Est2 has unique structural features that allow CoA binding. A comparison of the structure of AF-Est2 with the human carboxyl esterase 1, which has CoA thioesterase activity, reveals that CoA is bound to different parts of the core domain in these two enzymes and approaches the active site from opposite directions. PMID:27160974

Precise small molecule recognition of a toxic CUG RNA repeat expansion

PubMed Central

Rzuczek, Suzanne G; Colgan, Lesley A; Nakai, Yoshio; Cameron, Michael D; Furling, Denis; Yasuda, Ryohei; Disney, Matthew D

2017-01-01

Excluding the ribosome and riboswitches, developing small molecules that selectively target RNA is a longstanding problem in chemical biology. A typical cellular RNA is difficult to target because it has little tertiary, but abundant secondary structure. We designed allele-selective compounds that target such an RNA, the toxic noncoding repeat expansion (r(CUG)exp) that causes myotonic dystrophy type 1 (DM1). We developed several strategies to generate allele-selective small molecules, including non-covalent binding, covalent binding, cleavage and on-site probe synthesis. Covalent binding and cleavage enabled target profiling in cells derived from individuals with DM1, showing precise recognition of r(CUG)exp. In the on-site probe synthesis approach, small molecules bound adjacent sites in r(CUG)exp and reacted to afford picomolar inhibitors via a proximity-based click reaction only in DM1-affected cells. We expanded this approach to image r(CUG)exp in its natural context. PMID:27941760

Precise small-molecule recognition of a toxic CUG RNA repeat expansion.

PubMed

Rzuczek, Suzanne G; Colgan, Lesley A; Nakai, Yoshio; Cameron, Michael D; Furling, Denis; Yasuda, Ryohei; Disney, Matthew D

2017-02-01

Excluding the ribosome and riboswitches, developing small molecules that selectively target RNA is a longstanding problem in chemical biology. A typical cellular RNA is difficult to target because it has little tertiary, but abundant secondary structure. We designed allele-selective compounds that target such an RNA, the toxic noncoding repeat expansion (r(CUG) exp ) that causes myotonic dystrophy type 1 (DM1). We developed several strategies to generate allele-selective small molecules, including non-covalent binding, covalent binding, cleavage and on-site probe synthesis. Covalent binding and cleavage enabled target profiling in cells derived from individuals with DM1, showing precise recognition of r(CUG) exp . In the on-site probe synthesis approach, small molecules bound adjacent sites in r(CUG) exp and reacted to afford picomolar inhibitors via a proximity-based click reaction only in DM1-affected cells. We expanded this approach to image r(CUG) exp in its natural context.

Analytic studies of local-severe-storm observables by satellites

NASA Technical Reports Server (NTRS)

Dergarabedian, P.; Fendell, F.

1977-01-01

Attention is concentrated on the exceptionally violet whirlwind, often characterized by a fairly vertical axis of rotation. For a cylindrical polar coordinate system with axis coincident with the axis of rotation, the secondary flow involves the radial and axial velocity components. The thesis advanced is, first, that a violent whirlwind is characterized by swirl speeds relative to the axis of rotation on the order of 90 m/s, with 100 m/s being close to an upper bound. This estimate is based on interpretation of funnel-cloud shape (which also suggests properties of the radial profile of swirl, as well as the maximum magnitude); an error assessment of the funnel-cloud interpretation procedure is developed. Second, computation of ground-level pressure deficits achievable from typical tornado-spawning ambients by idealized thermohydrostatic processes suggests that a two-cell structure is required to sustain such large speeds.

Characterization of a Novel Association between Two Trypanosome-Specific Proteins and 5S rRNA

PubMed Central

Ciganda, Martin; Williams, Noreen

2012-01-01

P34 and P37 are two previously identified RNA binding proteins in the flagellate protozoan Trypanosoma brucei. RNA interference studies have determined that the proteins are essential and are involved in ribosome biogenesis. Here, we show that these proteins interact in vitro with the 5S rRNA with nearly identical binding characteristics in the absence of other cellular factors. The T. brucei 5S rRNA has a complex secondary structure and presents four accessible loops (A to D) for interactions with RNA-binding proteins. In other eukaryotes, loop C is bound by the L5 ribosomal protein and loop A mainly by TFIIIA. The binding of P34 and P37 to T. brucei 5S rRNA involves the LoopA region of the RNA, but these proteins also protect the L5 binding site located on LoopC. PMID:22253864

Froissart bound and self-similarity based models of proton structure functions

NASA Astrophysics Data System (ADS)

Choudhury, D. K.; Saikia, Baishali

2018-03-01

Froissart bound implies that the total proton-proton cross-section (or equivalently proton structure function) cannot rise faster than log2s ˜log2 1 x. Compatibility of such behavior with the notion of self-similarity in proton structure function was suggested by us sometime back. In the present work, we generalize and improve it further by considering more recent self-similarity based models of proton structure functions and compare with recent data as well as with the model of Block, Durand, Ha and McKay.

Two Active Site Divalent Ions in the Crystal Structure of the Hammerhead Ribozyme Bound to a Transition State Analogue.

PubMed

Mir, Aamir; Golden, Barbara L

2016-02-02

The crystal structure of the hammerhead ribozyme bound to the pentavalent transition state analogue vanadate reveals significant rearrangements relative to the previously determined structures. The active site contracts, bringing G10.1 closer to the cleavage site and repositioning a divalent metal ion such that it could, ultimately, interact directly with the scissile phosphate. This ion could also position a water molecule to serve as a general acid in the cleavage reaction. A second divalent ion is observed coordinated to O6 of G12. This metal ion is well-placed to help tune the pKA of G12. On the basis of this crystal structure as well as a wealth of biochemical studies, we propose a mechanism in which G12 serves as the general base and a magnesium-bound water serves as a general acid.

Protonated sugars: vibrational spectroscopy and conformational structure of protonated O-methyl α-D-galactopyranoside

NASA Astrophysics Data System (ADS)

Rudić, Svemir; Xie, Hong-bin; Gerber, R. Benny; Simons, John P.

2012-08-01

'Bridging' protons provide a common structural motif in biological assemblies such as proton wires and proton-bound dimers. Here we present a 'proof-of-principle' computational and vibrational spectroscopic investigation of an 'intra-molecular proton-bound dimer,' O-methyl α-D-galactopyranoside (αMeGal-H+), generated in the gas phase through photo-ionisation of its complex with phenol in a molecular beam. Its vibrational spectrum corresponds well with a classical molecular dynamics simulation conducted 'on-the-fly' and also with the lowest-energy structures predicted by DFT and ab initio calculations. They reveal proton-bound structures that bridge neighbouring pairs of oxygen atoms, preferentially O6 and O4, linked together within the carbohydrate scaffold. Motivated by the possibility of an entry into the microscopic mechanism of its acid (or enzyme)-catalysed hydrolysis, we also report the corresponding predictions for its singly hydrated complex.

Two active site divalent ions in the crystal structure of the hammerhead ribozyme bound to a transition state analogue

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

Mir, Aamir; Golden, Barbara L.

2015-11-09

The crystal structure of the hammerhead ribozyme bound to the pentavalent transition state analogue vanadate reveals significant rearrangements relative to the previously determined structures. The active site contracts, bringing G10.1 closer to the cleavage site and repositioning a divalent metal ion such that it could, ultimately, interact directly with the scissile phosphate. This ion could also position a water molecule to serve as a general acid in the cleavage reaction. A second divalent ion is observed coordinated to O6 of G12. This metal ion is well-placed to help tune the p K A of G12. Finally, on the basis ofmore » this crystal structure as well as a wealth of biochemical studies, in this paper we propose a mechanism in which G12 serves as the general base and a magnesium-bound water serves as a general acid.« less

A Maltose-Binding Protein Fusion Construct Yields a Robust Crystallography Platform for MCL1

PubMed Central

Clifton, Matthew C.; Dranow, David M.; Leed, Alison; Fulroth, Ben; Fairman, James W.; Abendroth, Jan; Atkins, Kateri A.; Wallace, Ellen; Fan, Dazhong; Xu, Guoping; Ni, Z. J.; Daniels, Doug; Van Drie, John; Wei, Guo; Burgin, Alex B.; Golub, Todd R.; Hubbard, Brian K.; Serrano-Wu, Michael H.

2015-01-01

Crystallization of a maltose-binding protein MCL1 fusion has yielded a robust crystallography platform that generated the first apo MCL1 crystal structure, as well as five ligand-bound structures. The ability to obtain fragment-bound structures advances structure-based drug design efforts that, despite considerable effort, had previously been intractable by crystallography. In the ligand-independent crystal form we identify inhibitor binding modes not observed in earlier crystallographic systems. This MBP-MCL1 construct dramatically improves the structural understanding of well-validated MCL1 ligands, and will likely catalyze the structure-based optimization of high affinity MCL1 inhibitors. PMID:25909780

The Effects of Focused/Unfocused Audio-Appended Reading Tasks on Intermediate Female EFL Learners' Written Accuracy

ERIC Educational Resources Information Center

Alavinia, Parviz; Shafaei, Ali; Salimi, Asghar

2018-01-01

The current research study was conducted to examine the impacts of focused and unfocused audio-appended reading tasks on female EFL learners' acquisition of a rule-bound structure (passive voice) and a non-rule-bound structure (prepositions). The participants of this study involved ninety intermediate female English learners. They were assigned…

A shock spectra and impedance method to determine a bound for spacecraft structural loads

NASA Technical Reports Server (NTRS)

Bamford, R.; Trubert, M.

1974-01-01

A method to determine a bound of structural loads for a spacecraft mounted on a launch vehicle is developed. The method utilizes the interface shock spectra and the relative impedance of the spacecraft and launch vehicle. The method is developed for single-degree-of-freedom models and then generalized to multidegree-of-freedom models.

An analysis of the vertical structure equation for arbitrary thermal profiles

NASA Technical Reports Server (NTRS)

Cohn, Stephen E.; Dee, Dick P.

1989-01-01

The vertical structure equation is a singular Sturm-Liouville problem whose eigenfunctions describe the vertical dependence of the normal modes of the primitive equations linearized about a given thermal profile. The eigenvalues give the equivalent depths of the modes. The spectrum of the vertical structure equation and the appropriateness of various upper boundary conditions, both for arbitrary thermal profiles were studied. The results depend critically upon whether or not the thermal profile is such that the basic state atmosphere is bounded. In the case of a bounded atmosphere it is shown that the spectrum is always totally discrete, regardless of details of the thermal profile. For the barotropic equivalent depth, which corresponds to the lowest eigen value, upper and lower bounds which depend only on the surface temperature and the atmosphere height were obtained. All eigenfunctions are bounded, but always have unbounded first derivatives. It was proved that the commonly invoked upper boundary condition that vertical velocity must vanish as pressure tends to zero, as well as a number of alternative conditions, is well posed. It was concluded that the vertical structure equation always has a totally discrete spectrum under the assumptions implicit in the primitive equations.

An analysis of the vertical structure equation for arbitrary thermal profiles

NASA Technical Reports Server (NTRS)

Cohn, Stephen E.; Dee, Dick P.

1987-01-01

The vertical structure equation is a singular Sturm-Liouville problem whose eigenfunctions describe the vertical dependence of the normal modes of the primitive equations linearized about a given thermal profile. The eigenvalues give the equivalent depths of the modes. The spectrum of the vertical structure equation and the appropriateness of various upper boundary conditions, both for arbitrary thermal profiles were studied. The results depend critically upon whether or not the thermal profile is such that the basic state atmosphere is bounded. In the case of a bounded atmosphere it is shown that the spectrum is always totally discrete, regardless of details of the thermal profile. For the barotropic equivalent depth, which corresponds to the lowest eigen value, upper and lower bounds which depend only on the surface temperature and the atmosphere height were obtained. All eigenfunctions are bounded, but always have unbounded first derivatives. It was proved that the commonly invoked upper boundary condition that vertical velocity must vanish as pressure tends to zero, as well as a number of alternative conditions, is well posed. It was concluded that the vertical structure equation always has a totally discrete spectrum under the assumptions implicit in the primitive equations.

The secondary cell wall polysaccharide of Bacillus anthracis provides the specific binding ligand for the C-terminal cell wall-binding domain of two phage endolysins, PlyL and PlyG

PubMed Central

Ganguly, Jhuma; Low, Lieh Y; Kamal, Nazia; Saile, Elke; Forsberg, L Scott; Gutierrez-Sanchez, Gerardo; Hoffmaster, Alex R; Liddington, Robert; Quinn, Conrad P; Carlson, Russell W; Kannenberg, Elmar L

2013-01-01

Endolysins are bacteriophage enzymes that lyse their bacterial host for phage progeny release. They commonly contain an N-terminal catalytic domain that hydrolyzes bacterial peptidoglycan (PG) and a C-terminal cell wall-binding domain (CBD) that confers enzyme localization to the PG substrate. Two endolysins, phage lysin L (PlyL) and phage lysin G (PlyG), are specific for Bacillus anthracis. To date, the cell wall ligands for their C-terminal CBD have not been identified. We recently described structures for a number of secondary cell wall polysaccharides (SCWPs) from B. anthracis and B. cereus strains. They are covalently bound to the PG and are comprised of a -ManNAc-GlcNAc-HexNAc- backbone with various galactosyl or glucosyl substitutions. Surface plasmon resonance (SPR) showed that the endolysins PlyL and PlyG bind to the SCWP from B. anthracis (SCWPBa) with high affinity (i.e. in the μM range with dissociation constants ranging from 0.81 × 10−6 to 7.51 × 10−6 M). In addition, the PlyL and PlyG SCWPBa binding sites reside with their C-terminal domains. The dissociation constants for the interactions of these endolysins and their derived C-terminal domains with the SCWPBa were in the range reported for other protein–carbohydrate interactions. Our findings show that the SCWPBa is the ligand that confers PlyL and PlyG lysin binding and localization to the PG. PlyL and PlyG also bound the SCWP from B. cereus G9241 with comparable affinities to SCWPBa. No detectable binding was found to the SCWPs from B. cereus ATCC (American Type Culture Collection) 10987 and ATCC 14579, thus demonstrating specificity of lysin binding to SCWPs. PMID:23493680

Location of the antigenic determinants of conjugative F-like pili.

PubMed Central

Worobec, E A; Frost, L S; Pieroni, P; Armstrong, G D; Hodges, R S; Parker, J M; Finlay, B B; Paranchych, W

1986-01-01

The amino terminus of the pilin protein constitutes the major epitope of F-like conjugative pili studied to date (F, ColB2, R1-19, R100-1, and pED208). Anti-pED208 pilus antibodies were passed through a CNBr-Sepharose affinity column linked to bovine serum albumin which was conjugated to a synthetic peptide, AcP(1-12), containing the major epitope at the amino terminus of pED208 pilin. This allowed the separation of two classes of antibodies; one was specific for the amino terminus and bound to the column, while the other, which recognizes a second epitope on the pilus, did not bind to the column. In addition, antibodies were raised against two amino-terminal peptide-bovine serum albumin conjugates [AcP(1-8) and AcP(1-12)] to ensure a source of pure, high-titer antibodies directed against the amino terminus. The location of these antibodies on intact pili was assayed by immunoelectron microscopy with a protein A-gold technique. The amino terminus-specific antibodies did not bind to the sides of the pili but appeared to be associated with the pilus tip. In addition, these antibodies were found to bind to the vesicle-like structure at the base of the pilus. The anti-pilus antibodies not specific for the amino terminus (unbound immunoglobulin G) were found to bind to the sides of the pilus. Anti-F and anti-ColB2 pilus antibodies bound to the sides of F, ColB2, and R1-19 pili, which have only their secondary epitope in common. The carboxyl-terminal lysine of R1-19 pilin prevents the absorption of anti-F plus antiserum but not anti-ColB2 pilus antiserum to the sides of the pilus, presumably by interfering with the recognition of this secondary epitope. Images PMID:2426247

Location of the antigenic determinants of conjugative F-like pili.

PubMed

Worobec, E A; Frost, L S; Pieroni, P; Armstrong, G D; Hodges, R S; Parker, J M; Finlay, B B; Paranchych, W

1986-08-01

The amino terminus of the pilin protein constitutes the major epitope of F-like conjugative pili studied to date (F, ColB2, R1-19, R100-1, and pED208). Anti-pED208 pilus antibodies were passed through a CNBr-Sepharose affinity column linked to bovine serum albumin which was conjugated to a synthetic peptide, AcP(1-12), containing the major epitope at the amino terminus of pED208 pilin. This allowed the separation of two classes of antibodies; one was specific for the amino terminus and bound to the column, while the other, which recognizes a second epitope on the pilus, did not bind to the column. In addition, antibodies were raised against two amino-terminal peptide-bovine serum albumin conjugates [AcP(1-8) and AcP(1-12)] to ensure a source of pure, high-titer antibodies directed against the amino terminus. The location of these antibodies on intact pili was assayed by immunoelectron microscopy with a protein A-gold technique. The amino terminus-specific antibodies did not bind to the sides of the pili but appeared to be associated with the pilus tip. In addition, these antibodies were found to bind to the vesicle-like structure at the base of the pilus. The anti-pilus antibodies not specific for the amino terminus (unbound immunoglobulin G) were found to bind to the sides of the pilus. Anti-F and anti-ColB2 pilus antibodies bound to the sides of F, ColB2, and R1-19 pili, which have only their secondary epitope in common. The carboxyl-terminal lysine of R1-19 pilin prevents the absorption of anti-F plus antiserum but not anti-ColB2 pilus antiserum to the sides of the pilus, presumably by interfering with the recognition of this secondary epitope.

CSI 3.0: a web server for identifying secondary and super-secondary structure in proteins using NMR chemical shifts.

PubMed

Hafsa, Noor E; Arndt, David; Wishart, David S

2015-07-01

The Chemical Shift Index or CSI 3.0 (http://csi3.wishartlab.com) is a web server designed to accurately identify the location of secondary and super-secondary structures in protein chains using only nuclear magnetic resonance (NMR) backbone chemical shifts and their corresponding protein sequence data. Unlike earlier versions of CSI, which only identified three types of secondary structure (helix, β-strand and coil), CSI 3.0 now identifies total of 11 types of secondary and super-secondary structures, including helices, β-strands, coil regions, five common β-turns (type I, II, I', II' and VIII), β hairpins as well as interior and edge β-strands. CSI 3.0 accepts experimental NMR chemical shift data in multiple formats (NMR Star 2.1, NMR Star 3.1 and SHIFTY) and generates colorful CSI plots (bar graphs) and secondary/super-secondary structure assignments. The output can be readily used as constraints for structure determination and refinement or the images may be used for presentations and publications. CSI 3.0 uses a pipeline of several well-tested, previously published programs to identify the secondary and super-secondary structures in protein chains. Comparisons with secondary and super-secondary structure assignments made via standard coordinate analysis programs such as DSSP, STRIDE and VADAR on high-resolution protein structures solved by X-ray and NMR show >90% agreement between those made with CSI 3.0. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

The conservation and function of RNA secondary structure in plants

PubMed Central

Vandivier, Lee E.; Anderson, Stephen J.; Foley, Shawn W.; Gregory, Brian D.

2016-01-01

RNA transcripts fold into secondary structures via intricate patterns of base pairing. These secondary structures impart catalytic, ligand binding, and scaffolding functions to a wide array of RNAs, forming a critical node of biological regulation. Among their many functions, RNA structural elements modulate epigenetic marks, alter mRNA stability and translation, regulate alternative splicing, transduce signals, and scaffold large macromolecular complexes. Thus, the study of RNA secondary structure is critical to understanding the function and regulation of RNA transcripts. Here, we review the origins, form, and function of RNA secondary structure, focusing on plants. We then provide an overview of methods for probing secondary structure, from physical methods such as X-ray crystallography and nuclear magnetic resonance imaging (NMR) to chemical and nuclease probing methods. Marriage with high-throughput sequencing has enabled these latter methods to scale across whole transcriptomes, yielding tremendous new insights into the form and function of RNA secondary structure. PMID:26865341

Structures of riboswitch RNA reaction states by mix-and-inject XFEL serial crystallography

NASA Astrophysics Data System (ADS)

Stagno, J. R.; Liu, Y.; Bhandari, Y. R.; Conrad, C. E.; Panja, S.; Swain, M.; Fan, L.; Nelson, G.; Li, C.; Wendel, D. R.; White, T. A.; Coe, J. D.; Wiedorn, M. O.; Knoska, J.; Oberthuer, D.; Tuckey, R. A.; Yu, P.; Dyba, M.; Tarasov, S. G.; Weierstall, U.; Grant, T. D.; Schwieters, C. D.; Zhang, J.; Ferré-D'Amaré, A. R.; Fromme, P.; Draper, D. E.; Liang, M.; Hunter, M. S.; Boutet, S.; Tan, K.; Zuo, X.; Ji, X.; Barty, A.; Zatsepin, N. A.; Chapman, H. N.; Spence, J. C. H.; Woodson, S. A.; Wang, Y.-X.

2017-01-01

Riboswitches are structural RNA elements that are generally located in the 5‧ untranslated region of messenger RNA. During regulation of gene expression, ligand binding to the aptamer domain of a riboswitch triggers a signal to the downstream expression platform. A complete understanding of the structural basis of this mechanism requires the ability to study structural changes over time. Here we use femtosecond X-ray free electron laser (XFEL) pulses to obtain structural measurements from crystals so small that diffusion of a ligand can be timed to initiate a reaction before diffraction. We demonstrate this approach by determining four structures of the adenine riboswitch aptamer domain during the course of a reaction, involving two unbound apo structures, one ligand-bound intermediate, and the final ligand-bound conformation. These structures support a reaction mechanism model with at least four states and illustrate the structural basis of signal transmission. The three-way junction and the P1 switch helix of the two apo conformers are notably different from those in the ligand-bound conformation. Our time-resolved crystallographic measurements with a 10-second delay captured the structure of an intermediate with changes in the binding pocket that accommodate the ligand. With at least a 10-minute delay, the RNA molecules were fully converted to the ligand-bound state, in which the substantial conformational changes resulted in conversion of the space group. Such notable changes in crystallo highlight the important opportunities that micro- and nanocrystals may offer in these and similar time-resolved diffraction studies. Together, these results demonstrate the potential of ‘mix-and-inject’ time-resolved serial crystallography to study biochemically important interactions between biomacromolecules and ligands, including those that involve large conformational changes.

Structures of riboswitch RNA reaction states by mix-and-inject XFEL serial crystallography

PubMed Central

Stagno, J. R.; Liu, Y.; Bhandari, Y. R.; Conrad, C. E.; Panja, S.; Swain, M.; Fan, L.; Nelson, G.; Li, C.; Wendel, D. R.; White, T. A.; Coe, J. D.; Wiedorn, M. O.; Knoska, J.; Oberthuer, D.; Tuckey, R. A.; Yu, P.; Dyba, M.; Tarasov, S. G.; Weierstall, U.; Grant, T. D.; Schwieters, C. D.; Zhang, J.; Ferré-D’Amaré, A. R.; Fromme, P.; Draper, D. E.; Liang, M.; Hunter, M. S.; Boutet, S.; Tan, K.; Zuo, X.; Ji, X.; Barty, A.; Zatsepin, N. A.; Chapman, H. N.; Spence, J. C. H.; Woodson, S. A.; Wang, Y.-X.

2017-01-01

Riboswitches are structural RNA elements that are generally located in the 5′ untranslated region of messenger RNA. During regulation of gene expression, ligand binding to the aptamer domain of a riboswitch triggers a signal to the downstream expression platform1–3. A complete understanding of the structural basis of this mechanism requires the ability to study structural changes over time4. Here we use femtosecond X-ray free electron laser (XFEL) pulses5,6 to obtain structural measurements from crystals so small that diffusion of a ligand can be timed to initiate a reaction before diffraction. We demonstrate this approach by determining four structures of the adenine riboswitch aptamer domain during the course of a reaction, involving two unbound apo structures, one ligand-bound intermediate, and the final ligand-bound conformation. These structures support a reaction mechanism model with at least four states and illustrate the structural basis of signal transmission. The three-way junction and the P1 switch helix of the two apo conformers are notably different from those in the ligand-bound conformation. Our time-resolved crystallographic measurements with a 10-second delay captured the structure of an intermediate with changes in the binding pocket that accommodate the ligand. With at least a 10-minute delay, the RNA molecules were fully converted to the ligand-bound state, in which the substantial conformational changes resulted in conversion of the space group. Such notable changes in crystallo highlight the important opportunities that micro- and nanocrystals may offer in these and similar time-resolved diffraction studies. Together, these results demonstrate the potential of ‘mix-and-inject’ time-resolved serial crystallography to study biochemically important interactions between biomacromolecules and ligands, including those that involve large conformational changes. PMID:27841871

Structured Uncertainty Bound Determination From Data for Control and Performance Validation

NASA Technical Reports Server (NTRS)

Lim, Kyong B.

2003-01-01

This report attempts to document the broad scope of issues that must be satisfactorily resolved before one can expect to methodically obtain, with a reasonable confidence, a near-optimal robust closed loop performance in physical applications. These include elements of signal processing, noise identification, system identification, model validation, and uncertainty modeling. Based on a recently developed methodology involving a parameterization of all model validating uncertainty sets for a given linear fractional transformation (LFT) structure and noise allowance, a new software, Uncertainty Bound Identification (UBID) toolbox, which conveniently executes model validation tests and determine uncertainty bounds from data, has been designed and is currently available. This toolbox also serves to benchmark the current state-of-the-art in uncertainty bound determination and in turn facilitate benchmarking of robust control technology. To help clarify the methodology and use of the new software, two tutorial examples are provided. The first involves the uncertainty characterization of a flexible structure dynamics, and the second example involves a closed loop performance validation of a ducted fan based on an uncertainty bound from data. These examples, along with other simulation and experimental results, also help describe the many factors and assumptions that determine the degree of success in applying robust control theory to practical problems.

Structure and dynamics of proflavine association around DNA.

PubMed

Sasikala, Wilbee D; Mukherjee, Arnab

2016-04-21

Proflavine is a small molecule that intercalates into DNA and, thereby, acts as an anticancer agent. Intercalation of proflavine is shown to be a two-step process in which the first step is believed to be the formation of a pre-intercalative outside bound state. Experimental studies so far have been unable to capture the nature of the outside bound state. However, the sub-millisecond timescale observed in fluorescence kinetic experiments is often attributed to the binding of proflavine outside of DNA. Here, we have performed molecular dynamics simulations with multiple proflavine molecules to study the structure and dynamics of the formation of the outside bound state of DNA at different ion concentrations. We observed that the timescale of the outside bound state formation is, at least, five orders of magnitude faster (in nanoseconds) than the experimentally reported timescale (sub-milliseconds) attributed to binding outside DNA. Moreover, we also observed the stacked arrangement of proflavine all around DNA, which is different from the experimentally predicted stacking arrangement perpendicular to the helical axis of DNA in the close vicinity of the phosphate groups. This study, therefore, provides insight into the molecular structure and dynamics of the pre-intercalative outside bound state and will help in understanding the overall intercalation mechanism.

Sound-absorbing slabs and structures based on granular materials (bound and unbound). [energy absorbing efficiency of porous material

NASA Technical Reports Server (NTRS)

Petre-Lazar, S.; Popeea, G.

1974-01-01

Sound absorbing slabs and structures made up of bound or unbound granular materials are considered and how to manufacture these elements at the building site. The raw material is a single grain powder (sand, expanded blast furnace slag, etc.) that imparts to the end products an apparent porosity of 25-45% and an energy dissipation within the structure leading to absorption coefficients that can be compared with those of mineral wool and urethane.

Improving virtual screening of G protein-coupled receptors via ligand-directed modeling

PubMed Central

Simms, John; Christopoulos, Arthur; Wootten, Denise

2017-01-01

G protein-coupled receptors (GPCRs) play crucial roles in cell physiology and pathophysiology. There is increasing interest in using structural information for virtual screening (VS) of libraries and for structure-based drug design to identify novel agonist or antagonist leads. However, the sparse availability of experimentally determined GPCR/ligand complex structures with diverse ligands impedes the application of structure-based drug design (SBDD) programs directed to identifying new molecules with a select pharmacology. In this study, we apply ligand-directed modeling (LDM) to available GPCR X-ray structures to improve VS performance and selectivity towards molecules of specific pharmacological profile. The described method refines a GPCR binding pocket conformation using a single known ligand for that GPCR. The LDM method is a computationally efficient, iterative workflow consisting of protein sampling and ligand docking. We developed an extensive benchmark comparing LDM-refined binding pockets to GPCR X-ray crystal structures across seven different GPCRs bound to a range of ligands of different chemotypes and pharmacological profiles. LDM-refined models showed improvement in VS performance over origin X-ray crystal structures in 21 out of 24 cases. In all cases, the LDM-refined models had superior performance in enriching for the chemotype of the refinement ligand. This likely contributes to the LDM success in all cases of inhibitor-bound to agonist-bound binding pocket refinement, a key task for GPCR SBDD programs. Indeed, agonist ligands are required for a plethora of GPCRs for therapeutic intervention, however GPCR X-ray structures are mostly restricted to their inactive inhibitor-bound state. PMID:29131821

JNSViewer—A JavaScript-based Nucleotide Sequence Viewer for DNA/RNA secondary structures

PubMed Central

Dong, Min; Graham, Mitchell; Yadav, Nehul

2017-01-01

Many tools are available for visualizing RNA or DNA secondary structures, but there is scarce implementation in JavaScript that provides seamless integration with the increasingly popular web computational platforms. We have developed JNSViewer, a highly interactive web service, which is bundled with several popular tools for DNA/RNA secondary structure prediction and can provide precise and interactive correspondence among nucleotides, dot-bracket data, secondary structure graphs, and genic annotations. In JNSViewer, users can perform RNA secondary structure predictions with different programs and settings, add customized genic annotations in GFF format to structure graphs, search for specific linear motifs, and extract relevant structure graphs of sub-sequences. JNSViewer also allows users to choose a transcript or specific segment of Arabidopsis thaliana genome sequences and predict the corresponding secondary structure. Popular genome browsers (i.e., JBrowse and BrowserGenome) were integrated into JNSViewer to provide powerful visualizations of chromosomal locations, genic annotations, and secondary structures. In addition, we used StructureFold with default settings to predict some RNA structures for Arabidopsis by incorporating in vivo high-throughput RNA structure profiling data and stored the results in our web server, which might be a useful resource for RNA secondary structure studies in plants. JNSViewer is available at http://bioinfolab.miamioh.edu/jnsviewer/index.html. PMID:28582416

Serial lectin affinity chromatography with concavalin A and wheat germ agglutinin demonstrates altered asparagine-linked sugar-chain structures of prostatic acid phosphatase in human prostate carcinoma.

PubMed

Yoshida, K I; Honda, M; Arai, K; Hosoya, Y; Moriguchi, H; Sumi, S; Ueda, Y; Kitahara, S

1997-08-01

Differences between human prostate carcinoma (PCA, five cases) and benign prostatic hyperplasia (BPH, five cases) in asparagine-linked (Asn) sugar-chain structure of prostatic acid phosphatase (PAP) were investigated using lectin affinity chromatography with concanavalin A (Con A) and wheat germ agglutinin (WGA). PAP activities were significantly decreased in PCA-derived PAP, while no significant differences between the two PAP preparations were observed in the enzymatic properties (Michaelis-Menten value, optimal pH, thermal stability, and inhibition study). In these PAP preparations, all activities were found only in the fractions which bound strongly to the Con A column and were undetectable in the Con A unbound fractions and in the fractions which bound weakly to the Con A column. The relative amounts of PAP which bound strongly to the Con A column but passed through the WGA column, were significantly greater in BPH-derived PAP than in PCA-derived PAP. In contrast, the relative amounts of PAP which bound strongly to the Con A column and bound to the WGA column, were significantly greater in PCA-derived PAP than in BPH-derived PAP. The findings suggest that Asn-linked sugar-chain structures are altered during oncogenesis in human prostate and also suggest that studies of qualitative differences of sugar-chain structures of PAP might lead to a useful diagnostic tool for PCA.

Conformational phases of membrane bound cytoskeletal filaments

NASA Astrophysics Data System (ADS)

Quint, David A.; Grason, Gregory; Gopinathan, Ajay

2013-03-01

Membrane bound cytoskeletal filaments found in living cells are employed to carry out many types of activities including cellular division, rigidity and transport. When these biopolymers are bound to a membrane surface they may take on highly non-trivial conformations as compared to when they are not bound. This leads to the natural question; What are the important interactions which drive these polymers to particular conformations when they are bound to a surface? Assuming that there are binding domains along the polymer which follow a periodic helical structure set by the natural monomeric handedness, these bound conformations must arise from the interplay of the intrinsic monomeric helicity and membrane binding. To probe this question, we study a continuous model of an elastic filament with intrinsic helicity and map out the conformational phases of this filament for various mechanical and structural parameters in our model, such as elastic stiffness and intrinsic twist of the filament. Our model allows us to gain insight into the possible mechanisms which drive real biopolymers such as actin and tubulin in eukaryotes and their prokaryotic cousins MreB and FtsZ to take on their functional conformations within living cells.

Anthrax Toxin Receptor 1 / Tumor Endothelial Marker 8: Mutation of Conserved Inserted Domain Residues Overrides Cytosolic Control of Protective Antigen Binding†

PubMed Central

Ramey, Jordan D.; Villareal, Valerie A.; Ng, Charles; Ward, Sabrina; Xiong, Jian-Ping; Clubb, Robert T.; Bradley, Kenneth A.

2010-01-01

Anthrax toxin receptor 1 (ANTXR1) / tumor endothelial marker 8 (TEM8) is one of two known proteinaceous cell surface anthrax toxin receptors. A metal ion dependent adhesion site (MIDAS) present in the integrin-like inserted (I) domain of ANTXR1 mediates the binding of the anthrax toxin subunit, protective antigen (PA). Here we provide evidence that single point mutations in the I domain can override regulation of ANTXR1 ligand-binding activity mediated by intracellular signals. A previously reported MIDAS-mutant of ANTXR1 (T118A) was found to retain normal metal ion binding and secondary structure but failed to bind PA, consistent with a locked inactive state. Conversely, mutation of a conserved I domain phenylalanine residue to a tryptophan (F205W) increased the proportion of cell-surface ANTXR1 that bound PA, consistent with a locked active state. Interestingly, the KD and total amount of PA bound by the isolated ANTXR1 I domain was not affected by the F205W mutation, indicating that ANTXR1 is preferentially found in the active state in the absence of inside-out signaling. Circular dichroism (CD) spectroscopy and 1H-15N heteronuclear single quantum coherence (HSQC) nuclear magnetic resonance (NMR) revealed that structural changes between T118A, F205W and WT I domains were minor despite a greater than 103-fold difference in their abilities to bind toxin. Regulation of toxin binding has important implications for the design of toxin inhibitors and for the targeting of ANTXR1 for anti-tumor therapies. PMID:20690680

Rtools: a web server for various secondary structural analyses on single RNA sequences.

PubMed

Hamada, Michiaki; Ono, Yukiteru; Kiryu, Hisanori; Sato, Kengo; Kato, Yuki; Fukunaga, Tsukasa; Mori, Ryota; Asai, Kiyoshi

2016-07-08

The secondary structures, as well as the nucleotide sequences, are the important features of RNA molecules to characterize their functions. According to the thermodynamic model, however, the probability of any secondary structure is very small. As a consequence, any tool to predict the secondary structures of RNAs has limited accuracy. On the other hand, there are a few tools to compensate the imperfect predictions by calculating and visualizing the secondary structural information from RNA sequences. It is desirable to obtain the rich information from those tools through a friendly interface. We implemented a web server of the tools to predict secondary structures and to calculate various structural features based on the energy models of secondary structures. By just giving an RNA sequence to the web server, the user can get the different types of solutions of the secondary structures, the marginal probabilities such as base-paring probabilities, loop probabilities and accessibilities of the local bases, the energy changes by arbitrary base mutations as well as the measures for validations of the predicted secondary structures. The web server is available at http://rtools.cbrc.jp, which integrates software tools, CentroidFold, CentroidHomfold, IPKnot, CapR, Raccess, Rchange and RintD. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

Cytoskeleton and Cytoskeleton-Bound RNA Visualization in Frog and Insect Oocytes.

PubMed

Kloc, Malgorzata; Bilinski, Szczepan; Kubiak, Jacek Z

2016-01-01

The majority of oocyte functions involves and depends on the cytoskeletal elements, which include microtubules and actin and cytokeratin filaments. Various structures and molecules are temporarily or permanently bound to the cytoskeletal elements and their functions rely on cytoskeleton integrity and its timely assembly. Thus the accurate visualization of cytoskeleton is often crucial for studies and analyses of oocyte structure and functions. Here we describe several reliable methods for microtubule and/or microfilaments preservation and visualization in Xenopus oocyte extracts, and in situ in live and fixed insect and frog (Xenopus) oocytes. In addition, we describe visualization of cytoskeleton-bound RNAs using molecular beacons in live Xenopus oocytes.

X-ray structures define human P2X3 receptor gating cycle and antagonist action

PubMed Central

Mansoor, Steven E.; Lü, Wei; Oosterheert, Wout; Shekhar, Mrinal; Tajkhorshid, Emad; Gouaux, Eric

2016-01-01

Summary P2X receptors are trimeric, non-selective cation channels activated by ATP that play important roles in cardiovascular, neuronal and immune systems. Despite their central function in human physiology and as potential targets of therapeutic agents, there are no structures of human P2X receptors. Mechanisms of receptor desensitization and ion permeation, principles of antagonism, and complete structure of the pore-forming transmembrane domains remain unclear. We report x-ray crystal structures of human P2X3 receptor in apo/resting, agonist-bound/open-pore, agonist-bound/desensitized and antagonist-bound closed states. The open state structure harbors an intracellular motif we term the “cytoplasmic cap”, that stabilizes the open state of the ion channel pore and creates lateral, phospholipid-lined cytoplasmic fenestrations for water and ion egress. Competitive antagonists TNP-ATP and A-317491 stabilize the apo/resting state and reveal the interactions responsible for competitive inhibition. These structures illuminate the conformational rearrangements underpinning P2X receptor gating and provide a foundation for development of new pharmacologic agents. PMID:27626375

Structures of human ADAR2 bound to dsRNA reveal base-flipping mechanism and basis for site selectivity

DOE PAGES

Matthews, Melissa M.; Thomas, Justin M.; Zheng, Yuxuan; ...

2016-04-11

Adenosine deaminases acting on RNA (ADARs) are editing enzymes that convert adenosine to inosine in duplex RNA, a modification reaction with wide-ranging consequences in RNA function. Understanding of the ADAR reaction mechanism, the origin of editing-site selectivity, and the effect of mutations is limited by the lack of high-resolution structural data for complexes of ADARs bound to substrate RNAs. In this paper, we describe four crystal structures of the human ADAR2 deaminase domain bound to RNA duplexes bearing a mimic of the deamination reaction intermediate. These structures, together with structure-guided mutagenesis and RNA-modification experiments, explain the basis of the ADARmore » deaminase domain's dsRNA specificity, its base-flipping mechanism, and its nearest-neighbor preferences. In addition, we identified an ADAR2-specific RNA-binding loop near the enzyme active site, thus rationalizing differences in selectivity observed between different ADARs. In conclusion, our results provide a structural framework for understanding the effects of ADAR mutations associated with human disease.« less

Surface-immobilized DNAzyme-type biocatalysis

NASA Astrophysics Data System (ADS)

Stefan, Loic; Lavergne, Thomas; Spinelli, Nicolas; Defrancq, Eric; Monchaud, David

2014-02-01

The structure of the double helix of deoxyribonucleic acid (DNA, also called duplex-DNA) was elucidated sixty years ago by Watson, Crick, Wilkins and Franklin. Since then, DNA has continued to hold a fascination for researchers in diverse fields including medicine and nanobiotechnology. Nature has indeed excelled in diversifying the use of DNA: beyond its canonical role of repository of genetic information, DNA could also act as a nanofactory able to perform some complex catalytic tasks in an enzyme-mimicking manner. The catalytic capability of DNA was termed DNAzyme; in this context, a peculiar DNA structure, a quadruple helix also named quadruplex-DNA, has recently garnered considerable interest since its autonomous catalytic proficiency relies on its higher-order folding that makes it suitable to interact efficiently with hemin, a natural cofactor of many enzymes. Quadruplexes have thus been widely studied for their hemoprotein-like properties, chiefly peroxidase-like activity, i.e., their ability to perform hemin-mediated catalytic oxidation reactions. Recent literature is replete with applications of quadruplex-based peroxidase-mimicking DNAzyme systems. Herein, we take a further leap along the road to biochemical applications, assessing the actual efficiency of catalytic quadruplexes for the detection of picomolar levels of surface-bound analytes in an enzyme-linked immunosorbent (ELISA)-type assay. To this end, we exploit an innovative strategy based on the functionalization of DNA by a multitasking platform named RAFT (for regioselectivity addressable functionalized template), whose versatility enables the grafting of DNA whatever its nature (duplex-DNA, quadruplex-DNA, etc.). We demonstrate that the resulting biotinylated RAFT/quadruplex systems indeed acquire catalytic properties that allow for efficient luminescent detection of picomoles of surface-bound streptavidin. We also highlight some of the pitfalls that have to be faced during optimization, notably demonstrating that highly optimized experimental conditions can make DNA pre-catalysts catalytically competent whatever their secondary structures.

Why They Come: SSATB Survey of Parents of Incoming Students

ERIC Educational Resources Information Center

Independent School, 2015

2015-01-01

What motivates families to consider an independent private school education and ultimately make the commitment to enroll their children? The Secondary School Admission Test Board (SSATB) surveyed 2,300 parents of private school-bound students in 2014 to answer this question. Based on the results of the survey, this brief article highlights five of…

Interactive-GIS-Tutor (IGIST) Integration: Creating a Digital Space Gateway within a Textbook-Bound South African Geography Class

ERIC Educational Resources Information Center

Fleischmann, E. M-L.; van der Westhuizen, C. P.; Cilliers, D.

2015-01-01

Currently there is a global drive among political and educational institutions to implement geographic information system (GIS) practice in secondary schools. However, Geography teachers worldwide, including in South Africa, face significant practical challenges in this regard. Lack of curriculum aligned GIS resources, funds and teacher GIS…

Multiple Pathways to College: A Secondary Analysis of the 2004 College Applicant Survey. ACAATO Document

ERIC Educational Resources Information Center

Colleges Ontario, 2005

2005-01-01

The 2004 College Applicant Survey (CAS) describes the college-bound applicant pool by covering a broad range of areas including key demographics, factors influencing college selection, academic background and financial preparedness. It is the most comprehensive and the richest source of survey data to date on applicants to the Ontario Colleges of…

The Effects of Cooperative Learning on the Classroom Participation of Students Placed at Risk for Societal Failure

ERIC Educational Resources Information Center

Drakeford, William

2012-01-01

A multiple baseline design across two subjects was used to determine the effectiveness of cooperative learning techniques on increasing student participation. The study was conducted on two male secondary students attending the upward bound pre-college program. Each student worked in small groups with specific roles, and two observers documented…

Numerical studies of laminar and turbulent drag reduction, part 2

NASA Technical Reports Server (NTRS)

Balasubramanian, R.; Orszag, S. A.

1983-01-01

The flow over wave shaped surfaces is studied using a Navier Stokes solver. Detailed comparisons with theoretical results are presented, including the stability of a laminar flow over wavy surfaces. Drag characteristics of nonplanar surfaces are predicted using the Navier-Stokes solver. The secondary instabilities of wall bounded and free shear flows are also discussed.

Chlorogenic Acids Biosynthesis in Centella asiatica Cells Is not Stimulated by Salicylic Acid Manipulation.

PubMed

Ncube, E N; Steenkamp, P A; Madala, N E; Dubery, I A

2016-07-01

Exogenous application of synthetic and natural elicitors of plant defence has been shown to result in mass production of secondary metabolites with nutraceuticals properties in cultured cells. In particular, salicylic acid (SA) treatment has been reported to induce the production of phenylpropanoids, including cinnamic acid derivatives bound to quinic acid (chlorogenic acids). Centella asiatica is an important medicinal plant with several therapeutic properties owing to its wide spectrum of secondary metabolites. We investigated the effect of SA on C. asiatica cells by monitoring perturbation of chlorogenic acids in particular. Different concentrations of SA were used to treat C. asiatica cells, and extracts from both treated and untreated cells were analysed using an optimised UHPLC-QTOF-MS/MS method. Semi-targeted multivariate data analyses with the aid of principal component analysis (PCA) and orthogonal projection to latent structures-discriminant analysis (OPLS-DA) revealed a concentration-dependent metabolic response. Surprisingly, a range of chlorogenic acid derivatives were found to be downregulated as a consequence of SA treatment. Moreover, irbic acid (3,5-O-dicaffeoyl-4-O-malonilquinic acid) was found to be a dominant CGA in C. asiatica cells, although the SA treatment also had a negative effect on its concentration. Overall SA treatment was found to be an ineffective elicitor of CGA production in cultured C. asiatica cells.

Bound states in string nets

NASA Astrophysics Data System (ADS)

Schulz, Marc Daniel; Dusuel, Sébastien; Vidal, Julien

2016-11-01

We discuss the emergence of bound states in the low-energy spectrum of the string-net Hamiltonian in the presence of a string tension. In the ladder geometry, we show that a single bound state arises either for a finite tension or in the zero-tension limit depending on the theory considered. In the latter case, we perturbatively compute the binding energy as a function of the total quantum dimension. We also address this issue in the honeycomb lattice where the number of bound states in the topological phase depends on the total quantum dimension. Finally, the internal structure of these bound states is analyzed in the zero-tension limit.

X-ray Thomson Scattering in Warm Dense Matter without the Chihara Decomposition.

PubMed

Baczewski, A D; Shulenburger, L; Desjarlais, M P; Hansen, S B; Magyar, R J

2016-03-18

X-ray Thomson scattering is an important experimental technique used to measure the temperature, ionization state, structure, and density of warm dense matter (WDM). The fundamental property probed in these experiments is the electronic dynamic structure factor. In most models, this is decomposed into three terms [J. Chihara, J. Phys. F 17, 295 (1987)] representing the response of tightly bound, loosely bound, and free electrons. Accompanying this decomposition is the classification of electrons as either bound or free, which is useful for gapped and cold systems but becomes increasingly questionable as temperatures and pressures increase into the WDM regime. In this work we provide unambiguous first principles calculations of the dynamic structure factor of warm dense beryllium, independent of the Chihara form, by treating bound and free states under a single formalism. The computational approach is real-time finite-temperature time-dependent density functional theory (TDDFT) being applied here for the first time to WDM. We compare results from TDDFT to Chihara-based calculations for experimentally relevant conditions in shock-compressed beryllium.

Channel branching ratios in CH2CN- photodetachment: Rotational structure and vibrational energy redistribution in autodetachment

NASA Astrophysics Data System (ADS)

Lyle, Justin; Wedig, Olivia; Gulania, Sahil; Krylov, Anna I.; Mabbs, Richard

2017-12-01

We report photoelectron spectra of CH2CN-, recorded at photon energies between 13 460 and 15 384 cm-1, which show rapid intensity variations in particular detachment channels. The branching ratios for various spectral features reveal rotational structure associated with autodetachment from an intermediate anion state. Calculations using equation-of-motion coupled-cluster method with single and double excitations reveal the presence of two dipole-bound excited anion states (a singlet and a triplet). The computed oscillator strength for the transition to the singlet dipole-bound state provides an estimate of the autodetachment channel contribution to the total photoelectron yield. Analysis of the different spectral features allows identification of the dipole-bound and neutral vibrational levels involved in the autodetachment processes. For the most part, the autodetachment channels are consistent with the vibrational propensity rule and normal mode expectation. However, examination of the rotational structure shows that autodetachment from the ν3 (v = 1 and v = 2) levels of the dipole-bound state displays behavior counter to the normal mode expectation with the final state vibrational level belonging to a different mode.

Binding of ReO[subscript 4];#8722; with an engineered MoO[subscript 4 superscript 2];#8722;-binding protein: towards a new approach in radiopharmaceutical applications

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

Aryal, Baikuntha P.; Brugarolas, Pedro; He, Chuan

2012-05-25

Radiolabeled biomolecules are routinely used for clinical diagnostics. {sup 99m}Tc is the most commonly used radioactive tracer in radiopharmaceuticals. {sup 188}Re and {sup 186}Re are also commonly used as radioactive tracers in medicine. However, currently available methods for radiolabeling are lengthy and involve several steps in bioconjugation processes. In this work we present a strategy to engineer proteins that may selectively recognize the perrhenate (ReO{sub 4}{sup -}) ion as a new way to label proteins. We found that a molybdate (MoO{sub 4}{sup 2-})-binding protein (ModA) from Escherichia coli can bind perrhenate with high affinity. Using fluorescence and isothermal titration calorimetrymore » measurements, we determined the dissociation constant of ModA for ReO{sub 4}{sup -} to be 541 nM and we solved a crystal structure of ModA with a bound ReO{sub 4}{sup -}. On the basis of the structure we created a mutant protein containing a disulfide linkage, which exhibited increased affinity for perrhenate (K{sub d} = 104 nM). High-resolution crystal structures of ModA (1.7 {angstrom}) and A11C/R153C mutant (2.0 {angstrom}) were solved with bound perrhenate. Both structures show that a perrhenate ion occupies the molybdate binding site using the same amino acid residues that are involved in molybdate binding. The overall structure of the perrhenate-bound ModA is unchanged compared with that of the molybdate-bound form. In the mutant protein, the bound perrhenate is further stabilized by the engineered disulfide bond.« less

The Structure of the Neurotoxin- Associated Protein HA33/A from Clostridium botulinum Suggests a Reoccurring Beta-Trefoil Fold in the Progenitor Toxin Complex

DTIC Science & Technology

2004-12-16

mistletoe lectin I. The ricin and mistle- toe lectin I structures revealed a domain architec- ture that are similar to HA33/A with two b-trefoil...domains. The complex crystal structures of ricin bound to lactose and mistletoe lectin I bound to galactose revealed that only the 1a and 2g repeats of...H., Tonevitsky, A. G., Agapov, I. I., Saward, S., Pfuller, U. & Palmer, R. A. (2003). Crystal structure at 3 Å of mistletoe lectin I, a dimeric

Studies in turbulence

NASA Technical Reports Server (NTRS)

Gatski, Thomas B. (Editor); Sarkar, Sutanu (Editor); Speziale, Charles G. (Editor)

1992-01-01

Various papers on turbulence are presented. Individual topics addressed include: modeling the dissipation rate in rotating turbulent flows, mapping closures for turbulent mixing and reaction, understanding turbulence in vortex dynamics, models for the structure and dynamics of near-wall turbulence, complexity of turbulence near a wall, proper orthogonal decomposition, propagating structures in wall-bounded turbulence flows. Also discussed are: constitutive relation in compressible turbulence, compressible turbulence and shock waves, direct simulation of compressible turbulence in a shear flow, structural genesis in wall-bounded turbulence flows, vortex lattice structure of turbulent shear slows, etiology of shear layer vortices, trilinear coordinates in fluid mechanics.

Statistical analysis of EGFR structures' performance in virtual screening

NASA Astrophysics Data System (ADS)

Li, Yan; Li, Xiang; Dong, Zigang

2015-11-01

In this work the ability of EGFR structures to distinguish true inhibitors from decoys in docking and MM-PBSA is assessed by statistical procedures. The docking performance depends critically on the receptor conformation and bound state. The enrichment of known inhibitors is well correlated with the difference between EGFR structures rather than the bound-ligand property. The optimal structures for virtual screening can be selected based purely on the complex information. And the mixed combination of distinct EGFR conformations is recommended for ensemble docking. In MM-PBSA, a variety of EGFR structures have identically good performance in the scoring and ranking of known inhibitors, indicating that the choice of the receptor structure has little effect on the screening.

Structure and anti-structure in the culture-bound syndromes: The Malay case.

PubMed

Lee, R L

1981-09-01

Turner's concepts of structure and anti-structure are applied to the culture-bound syndromes to demonstrate that they are dialectical aspects of cultural reality, The Malay cases of amok, latah and possession hysteria are discussed as instances of anti-structural behaviors that dramatize role-reversals and role-enhancement. The performers of these behaviors are not subjected to the Malay code of moral conduct. The supernatural ethos in Malay culture plays an important role in shaping tolerance towards them. Although this tolerance has been gradually eroded as a result of the introduction of Western psychiatry, the anti-structural status of these syndromes has not faded away but has assumed new meanings in terms of psychopathology.

The role of amino acid electron-donor/acceptor atoms in host-cell binding peptides is associated with their 3D structure and HLA-binding capacity in sterile malarial immunity induction

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

Patarroyo, Manuel E., E-mail: mepatarr@mail.com; Universidad Nacional de Colombia, Bogota; Almonacid, Hannia

Highlights: Black-Right-Pointing-Pointer Fundamental residues located in some HABPs are associated with their 3D structure. Black-Right-Pointing-Pointer Electron-donor atoms present in {beta}-turn, random, distorted {alpha}-helix structures. Black-Right-Pointing-Pointer Electron-donor atoms bound to HLA-DR53. Black-Right-Pointing-Pointer Electron-acceptor atoms present in regular {alpha}-helix structure bound to HLA-DR52. -- Abstract: Plasmodium falciparum malaria continues being one of the parasitic diseases causing the highest worldwide mortality due to the parasite's multiple evasion mechanisms, such as immunological silence. Membrane and organelle proteins are used during invasion for interactions mediated by high binding ability peptides (HABPs); these have amino acids which establish hydrogen bonds between them in some of theirmore » critical binding residues. Immunisation assays in the Aotus model using HABPs whose critical residues had been modified have revealed a conformational change thereby enabling a protection-inducing response. This has improved fitting within HLA-DR{beta}1{sup Asterisk-Operator} molecules where amino acid electron-donor atoms present in {beta}-turn, random or distorted {alpha}-helix structures preferentially bound to HLA-DR53 molecules, whilst HABPs having amino acid electron-acceptor atoms present in regular {alpha}-helix structure bound to HLA-DR52. This data has great implications for vaccine development.« less

Secondary flow in turbulent ducts with increasing aspect ratio

NASA Astrophysics Data System (ADS)

Vinuesa, R.; Schlatter, P.; Nagib, H. M.

2018-05-01

Direct numerical simulations of turbulent duct flows with aspect ratios 1, 3, 5, 7, 10, and 14.4 at a center-plane friction Reynolds number Reτ,c≃180 , and aspect ratios 1 and 3 at Reτ,c≃360 , were carried out with the spectral-element code nek5000. The aim of these simulations is to gain insight into the kinematics and dynamics of Prandtl's secondary flow of the second kind and its impact on the flow physics of wall-bounded turbulence. The secondary flow is characterized in terms of the cross-plane component of the mean kinetic energy, and its variation in the spanwise direction of the flow. Our results show that averaging times of around 3000 convective time units (based on duct half-height h ) are required to reach a converged state of the secondary flow, which extends up to a spanwise distance of around ≃5 h measured from the side walls. We also show that if the duct is not wide enough to accommodate the whole extent of the secondary flow, then its structure is modified as reflected through a different spanwise distribution of energy. Another confirmation of the extent of the secondary flow is the decay rate of kinetic energy of any remnant secondary motions for zc/h >5 (where zc is the spanwise distance from the corner) in aspect ratios 7, 10, and 14.4, which exhibits a decreasing level of energy with increasing averaging time ta, and in its rapid rate of decay given by ˜ta-1 . This is the same rate of decay observed in a spanwise-periodic channel simulation, which suggests that at the core, the kinetic energy of the secondary flow integrated over the cross-sectional area, , behaves as a random variable with zero mean, with rate of decay consistent with central limit theorem. Long-time averages of statistics in a region of rectangular ducts extending about the width of a well-designed channel simulation (i.e., extending about ≃3 h on each side of the center plane) indicate that ducts or experimental facilities with aspect ratios larger than 10 may, if properly designed, exhibit good agreement with results obtained from spanwise-periodic channel computations.

Deposition and reentrainment of Brownian particles in porous media under unfavorable chemical conditions: some concepts and applications.

PubMed

Hahn, Melinda W; O'Meliae, Charles R

2004-01-01

The deposition and reentrainment of particles in porous media have been examined theoretically and experimentally. A Brownian Dynamics/Monte Carlo (MC/BD) model has been developed that simulates the movement of Brownian particles near a collector under "unfavorable" chemical conditions and allows deposition in primary and secondary minima. A simple Maxwell approach has been used to estimate particle attachment efficiency by assuming deposition in the secondary minimum and calculating the probability of reentrainment. The MC/BD simulations and the Maxwell calculations support an alternative view of the deposition and reentrainment of Brownian particles under unfavorable chemical conditions. These calculations indicate that deposition into and subsequent release from secondary minima can explain reported discrepancies between classic model predictions that assume irreversible deposition in a primary well and experimentally determined deposition efficiencies that are orders of magnitude larger than Interaction Force Boundary Layer (IFBL) predictions. The commonly used IFBL model, for example, is based on the notion of transport over an energy barrier into the primary well and does not address contributions of secondary minimum deposition. A simple Maxwell model based on deposition into and reentrainment from secondary minima is much more accurate in predicting deposition rates for column experiments at low ionic strengths. It also greatly reduces the substantial particle size effects inherent in IFBL models, wherein particle attachment rates are predicted to decrease significantly with increasing particle size. This view is consistent with recent work by others addressing the composition and structure of the first few nanometers at solid-water interfaces including research on modeling water at solid-liquid interfaces, surface speciation, interfacial force measurements, and the rheological properties of concentrated suspensions. It follows that deposition under these conditions will depend on the depth of the secondary minimum and that some transition between secondary and primary depositions should occur when the height of the energy barrier is on the order of several kT. When deposition in secondary minima predominates, observed deposition should increase with increasing ionic strength, particle size, and Hamaker constant. Since an equilibrium can develop between bound and bulk particles, the collision efficiency [alpha] can no longer be considered a constant for a given physical and chemical system. Rather, in many cases it can decrease over time until it eventually reaches zero as equilibrium is established.

Mechanism of endonuclease cleavage by the HigB toxin

PubMed Central

Schureck, Marc A.; Repack, Adrienne; Miles, Stacey J.; Marquez, Jhomar; Dunham, Christine M.

2016-01-01

Bacteria encode multiple type II toxin–antitoxin modules that cleave ribosome-bound mRNAs in response to stress. All ribosome-dependent toxin family members structurally characterized to date adopt similar microbial RNase architectures despite possessing low sequence identities. Therefore, determining which residues are catalytically important in this specialized RNase family has been a challenge in the field. Structural studies of RelE and YoeB toxins bound to the ribosome provided significant insights but biochemical experiments with RelE were required to clearly demonstrate which residues are critical for acid-base catalysis of mRNA cleavage. Here, we solved an X-ray crystal structure of the wild-type, ribosome-dependent toxin HigB bound to the ribosome revealing potential catalytic residues proximal to the mRNA substrate. Using cell-based and biochemical assays, we further determined that HigB residues His54, Asp90, Tyr91 and His92 are critical for activity in vivo, while HigB H54A and Y91A variants have the largest effect on mRNA cleavage in vitro. Comparison of X-ray crystal structures of two catalytically inactive HigB variants with 70S-HigB bound structures reveal that HigB active site residues undergo conformational rearrangements likely required for recognition of its mRNA substrate. These data support the emerging concept that ribosome-dependent toxins have diverse modes of mRNA recognition. PMID:27378776

Minimal determinants for binding activated G-alpha from the structure of a G-alpha-i1/peptide dimer†

PubMed Central

Johnston, Christopher A.; Lobanova, Ekaterina S.; Shavkunov, Alexander S.; Low, Justin; Ramer, J. Kevin; Blaesius, Rainer; Fredericks, Zoey; Willard, Francis S.; Kuhlman, Brian; Arshavsky, Vadim Y.; Siderovski, David P.

2008-01-01

G-proteins cycle between an inactive GDP-bound state and active GTP-bound state, serving as molecular switches that coordinate cellular signaling. We recently used phage-display to identify a series of peptides that bind Gα subunits in a nucleotide-dependent manner [Johnston, C. A., Willard, F. S., Jezyk, M. R., Fredericks, Z., Bodor, E. T., Jones, M. B., Blaesius, R., Watts, V. J., Harden, T. K., Sondek, J., Ramer, J. K., and Siderovski, D. P. (2005) Structure 13, 1069–1080]. Here we describe the structural features and functions of KB-1753, a peptide that binds selectively to GDP·AlF4−- and GTPγS-bound states of Gαi subunits. KB-1753 blocks interaction of Gαtransducin with its effector, cGMP phosphodiesterase, and inhibits transducin-mediated activation of cGMP degradation. Additionally, KB-1753 interferes with RGS protein binding and resultant GAP activity. A fluorescent KB-1753 variant was found to act as a sensor for activated Gα in vitro. The crystal structure of KB-1753 bound to Gαi1·GDP·AlF4− reveals binding to a conserved hydrophobic groove between switch II and α3 helices, and, along with supporting biochemical data and previous structural analyses, supports the notion that this is the site of effector interactions for Gαi subunits. PMID:16981699

Stability of proton-bound clusters of alkyl alcohols, aldehydes and ketones in Ion Mobility Spectrometry.

PubMed

Jurado-Campos, Natividad; Garrido-Delgado, Rocío; Martínez-Haya, Bruno; Eiceman, Gary A; Arce, Lourdes

2018-08-01

Significant substances in emerging applications of ion mobility spectrometry such as breath analysis for clinical diagnostics and headspace analysis for food purity include low molar mass alcohols, ketones, aldehydes and esters which produce mobility spectra containing protonated monomers and proton-bound dimers. Spectra for all n- alcohols, aldehydes and ketones from carbon number three to eight exhibited protonated monomers and proton-bound dimers with ion drift times of 6.5-13.3 ms at ambient pressure and from 35° to 80 °C in nitrogen. Only n-alcohols from 1-pentanol to 1-octanol produced proton-bound trimers which were sufficiently stable to be observed at these temperatures and drift times of 12.8-16.3 ms. Polar functional groups were protected in compact structures in ab initio models for proton-bound dimers of alcohols, ketones and aldehydes. Only alcohols formed a V-shaped arrangement for proton-bound trimers strengthening ion stability and lifetime. In contrast, models for proton-bound trimers of aldehydes and ketones showed association of the third neutral through weak, non-specific, long-range interactions consistent with ion dissociation in the ion mobility drift tube before arriving at the detector. Collision cross sections derived from reduced mobility coefficients in nitrogen gas atmosphere support the predicted ion structures and approximate degrees of hydration. Copyright © 2018 Elsevier B.V. All rights reserved.

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

Amdursky, Nadav; Gazit, Ehud; Rosenman, Gil, E-mail: gilr@eng.tau.ac.il

Highlights: Black-Right-Pointing-Pointer We observe lag-phase crystallization process in insulin. Black-Right-Pointing-Pointer The crystallization is a result of the formation of higher order oligomers. Black-Right-Pointing-Pointer The crystallization also changes the secondary structure of the protein. Black-Right-Pointing-Pointer The spectroscopic signature can be used for amyloid inhibitors assay. -- Abstract: Insulin, as other amyloid proteins, can form amyloid fibrils at certain conditions. The self-assembled aggregation process of insulin can result in a variety of conformations, starting from small oligomers, going through various types of protofibrils, and finishing with bundles of fibrils. One of the most common consensuses among the various self-assembly processes that aremore » suggested in the literature is the formation of an early stage nucleus conformation. Here we present an additional insight for the self-assembly process of insulin. We show that at the early lag phase of the process (prior to fibril formation) the insulin monomers self-assemble into ordered nanostructures. The most notable feature of this early self-assembly process is the formation of nanocrystalline nucleus regions with a strongly bound electron-hole confinement, which also change the secondary structure of the protein. Each step in the self-assembly process is characterized by an optical spectroscopic signature, and possesses a narrow size distribution. By following the spectroscopic signature we can measure the potency of amyloid fibrils inhibitors already at the lag phase. We further demonstrate it by the use of epigallocatechin gallate, a known inhibitor for insulin fibrils. The findings can result in a spectroscopic-based application for the analysis of amyloid fibrils inhibitors.« less

Extracting Loop Bounds for WCET Analysis Using the Instrumentation Point Graph

NASA Astrophysics Data System (ADS)

Betts, A.; Bernat, G.

2009-05-01

Every calculation engine proposed in the literature of Worst-Case Execution Time (WCET) analysis requires upper bounds on loop iterations. Existing mechanisms to procure this information are either error prone, because they are gathered from the end-user, or limited in scope, because automatic analyses target very specific loop structures. In this paper, we present a technique that obtains bounds completely automatically for arbitrary loop structures. In particular, we show how to employ the Instrumentation Point Graph (IPG) to parse traces of execution (generated by an instrumented program) in order to extract bounds relative to any loop-nesting level. With this technique, therefore, non-rectangular dependencies between loops can be captured, allowing more accurate WCET estimates to be calculated. We demonstrate the improvement in accuracy by comparing WCET estimates computed through our HMB framework against those computed with state-of-the-art techniques.

Structural rearrangements occurring upon cofactor binding in the Mycobacterium smegmatis β-ketoacyl-acyl carrier protein reductase MabA.

PubMed

Küssau, Tanja; Flipo, Marion; Van Wyk, Niel; Viljoen, Albertus; Olieric, Vincent; Kremer, Laurent; Blaise, Mickaël

2018-05-01

In mycobacteria, the ketoacyl-acyl carrier protein (ACP) reductase MabA (designated FabG in other bacteria) catalyzes the NADPH-dependent reduction of β-ketoacyl-ACP substrates to β-hydroxyacyl-ACP products. This first reductive step in the fatty-acid biosynthesis elongation cycle is essential for bacteria, which makes MabA/FabG an interesting drug target. To date, however, very few molecules targeting FabG have been discovered and MabA remains the only enzyme of the mycobacterial type II fatty-acid synthase that lacks specific inhibitors. Despite the existence of several MabA/FabG crystal structures, the structural rearrangement that occurs upon cofactor binding is still not fully understood. Therefore, unlocking this knowledge gap could help in the design of new inhibitors. Here, high-resolution crystal structures of MabA from Mycobacterium smegmatis in its apo, NADP + -bound and NADPH-bound forms are reported. Comparison of these crystal structures reveals the structural reorganization of the lid region covering the active site of the enzyme. The crystal structure of the apo form revealed numerous residues that trigger steric hindrance to the binding of NADPH and substrate. Upon NADPH binding, these residues are pushed away from the active site, allowing the enzyme to adopt an open conformation. The transition from an NADPH-bound to an NADP + -bound form is likely to facilitate release of the product. These results may be useful for subsequent rational drug design and/or for in silico drug-screening approaches targeting MabA/FabG.

The structures of non-CG-repeat Z-DNAs co-crystallized with the Z-DNA-binding domain, hZ alpha(ADAR1).

PubMed

Ha, Sung Chul; Choi, Jongkeun; Hwang, Hye-Yeon; Rich, Alexander; Kim, Yang-Gyun; Kim, Kyeong Kyu

2009-02-01

The Z-DNA conformation preferentially occurs at alternating purine-pyrimidine repeats, and is specifically recognized by Z alpha domains identified in several Z-DNA-binding proteins. The binding of Z alpha to foreign or chromosomal DNA in various sequence contexts is known to influence various biological functions, including the DNA-mediated innate immune response and transcriptional modulation of gene expression. For these reasons, understanding its binding mode and the conformational diversity of Z alpha bound Z-DNAs is of considerable importance. However, structural studies of Z alpha bound Z-DNA have been mostly limited to standard CG-repeat DNAs. Here, we have solved the crystal structures of three representative non-CG repeat DNAs, d(CACGTG)(2), d(CGTACG)(2) and d(CGGCCG)(2) complexed to hZ alpha(ADAR1) and compared those structures with that of hZ alpha(ADAR1)/d(CGCGCG)(2) and the Z alpha-free Z-DNAs. hZ alpha(ADAR1) bound to each of the three Z-DNAs showed a well conserved binding mode with very limited structural deviation irrespective of the DNA sequence, although varying numbers of residues were in contact with Z-DNA. Z-DNAs display less structural alterations in the Z alpha-bound state than in their free form, thereby suggesting that conformational diversities of Z-DNAs are restrained by the binding pocket of Z alpha. These data suggest that Z-DNAs are recognized by Z alpha through common conformational features regardless of the sequence and structural alterations.

RNA Secondary Structure Prediction by Using Discrete Mathematics: An Interdisciplinary Research Experience for Undergraduate Students

ERIC Educational Resources Information Center

Ellington, Roni; Wachira, James; Nkwanta, Asamoah

2010-01-01

The focus of this Research Experience for Undergraduates (REU) project was on RNA secondary structure prediction by using a lattice walk approach. The lattice walk approach is a combinatorial and computational biology method used to enumerate possible secondary structures and predict RNA secondary structure from RNA sequences. The method uses…

Fourier-based classification of protein secondary structures.

PubMed

Shu, Jian-Jun; Yong, Kian Yan

2017-04-15

The correct prediction of protein secondary structures is one of the key issues in predicting the correct protein folded shape, which is used for determining gene function. Existing methods make use of amino acids properties as indices to classify protein secondary structures, but are faced with a significant number of misclassifications. The paper presents a technique for the classification of protein secondary structures based on protein "signal-plotting" and the use of the Fourier technique for digital signal processing. New indices are proposed to classify protein secondary structures by analyzing hydrophobicity profiles. The approach is simple and straightforward. Results show that the more types of protein secondary structures can be classified by means of these newly-proposed indices. Copyright © 2017 Elsevier Inc. All rights reserved.

Giant Enhancement of Defect-Bound Exciton Luminescence and Suppression of Band-Edge Luminescence in Monolayer WSe2-Ag Plasmonic Hybrid Structures.

PubMed

Johnson, Alex D; Cheng, Fei; Tsai, Yutsung; Shih, Chih-Kang

2017-07-12

We have investigated how the photoluminescence (PL) of WSe 2 is modified when coupled to Ag plasmonic structures at low temperature. Chemical vapor deposition (CVD) grown monolayer WSe 2 flakes were transferred onto a Ag film and a Ag nanotriangle array that had a 1.5 nm Al 2 O 3 capping layer. Using low-temperature (7.5 K) micro-PL mapping, we simultaneously observed enhancement of the defect-bound exciton emission and quenching of the band edge exciton emission when the WSe 2 was on a plasmonic structure. The enhancement of the defect-bound exciton emission was significant with enhancement factors of up to ∼200 for WSe 2 on the nanotriangle array when compared to WSe 2 on a 1.5 nm Al 2 O 3 capped Si substrate with a 300 nm SiO 2 layer. The giant enhancement of the luminescence from the defect-bound excitons is understood in terms of the Purcell effect and increased light absorption. In contrast, the surprising result of luminescence quenching of the bright exciton state on the same plasmonic nanostructure is due to a rather unique electronic structure of WSe 2 : the existence of a dark state below the bright exciton state.

Structure and Function Study of HIV and Influenza Fusion Proteins

NASA Astrophysics Data System (ADS)

Liang, Shuang

Human immunodeficiency virus (HIV) and influenza virus are membrane-enveloped viruses causing acquired immunodeficiency syndrome (AIDS) and flu. The initial step of HIV and influenza virus infection is fusion between viral and host cell membrane catalyzed by the viral fusion protein gp41 and hemagglutinin (HA) respectively. However, the structure of gp41 and HA as well as the infection mechanism are still not fully understood. This work addresses (1) full length gp41 ectodomain and TM domain structure and function and (2) IFP membrane location and IFP-membrane interaction. My studies of gp41 protein and IFP can provide better understanding of the membrane fusion mechanism and may aid development of anti-viral therapeutics and vaccine. The full length ectodomain and transmembrane domain of gp41 and shorter constructs were expressed, purified and solubilized at physiology conditions. The constructs adopt overall alpha helical structure in SDS and DPC detergents, and showed hyperthermostability with Tm > 90 °C. The oligomeric states of these proteins vary in different detergent buffer: predominant trimer for all constructs and some hexamer fraction for HM and HM_TM protein in SDS at pH 7.4; and mixtures of monomer, trimer, and higher-order oligomer protein in DPC at pH 4.0 and 7.4. Substantial protein-induced vesicle fusion was observed, including fusion of neutral vesicles at neutral pH, which are the conditions similar HIV/cell fusion. Vesicle fusion by a gp41 ectodomain construct has rarely been observed under these conditions, and is aided by inclusion of both the FP and TM, and by protein which is predominantly trimer rather than monomer. Current data was integrated with existing data, and a structural model was proposed. Secondary structure and conformation of IFP is a helix-turn-helix structure in membrane. However, there has been arguments about the IFP membrane location. 13C-2H REDOR solid-state NMR is used to solve this problem. The IFP adopts major alpha helical, minor beta strand secondary structure in PC/PG membrane. The alpha helical IFP's with respectively 13CO labeled Leu-2, Ala-7 and Gly-16 all show close contacts with the lipid acyl chain tail, suggesting IFP has strong interaction with the membrane. By screening the current IFP topology models, it either has a membrane-spanning confirmation, or it promotes lipid trail protrusion. IFP bounded lipid membrane structure was studied by paramagnetic relaxation enhancement (PRE) solid-state NMR to provide more information about the detailed IFP membrane location model. The T2 relaxation time and rate were measured for membrane with or without IFP and with or without Mn2+ . Based on the results, it is concluded that IFP does not promote lipid protrusion at both gel phase and liquid phase, which is evidenced by that the R2 difference with and without Mn2+ is smaller for IFP free membrane than IFP bounded membrane, meaning IFP does not induce a smaller average distance between lipid acyl chain and aqueous layer. By integrating these results, a IFP membrane spanning model was proposed, in which IFP N-terminal helix adopts a 45° angle with respect to membrane normal.

DNA interactions with a Methylene Blue redox indicator depend on the DNA length and are sequence specific.

PubMed

Farjami, Elaheh; Clima, Lilia; Gothelf, Kurt V; Ferapontova, Elena E

2010-06-01

A DNA molecular beacon approach was used for the analysis of interactions between DNA and Methylene Blue (MB) as a redox indicator of a hybridization event. DNA hairpin structures of different length and guanine (G) content were immobilized onto gold electrodes in their folded states through the alkanethiol linker at the 5'-end. Binding of MB to the folded hairpin DNA was electrochemically studied and compared with binding to the duplex structure formed by hybridization of the hairpin DNA to a complementary DNA strand. Variation of the electrochemical signal from the DNA-MB complex was shown to depend primarily on the DNA length and sequence used: the G-C base pairs were the preferential sites of MB binding in the duplex. For short 20 nts long DNA sequences, the increased electrochemical response from MB bound to the duplex structure was consistent with the increased amount of bound and electrochemically readable MB molecules (i.e. MB molecules that are available for the electron transfer (ET) reaction with the electrode). With longer DNA sequences, the balance between the amounts of the electrochemically readable MB molecules bound to the hairpin DNA and to the hybrid was opposite: a part of the MB molecules bound to the long-sequence DNA duplex seem to be electrochemically mute due to long ET distance. The increasing electrochemical response from MB bound to the short-length DNA hybrid contrasts with the decreasing signal from MB bound to the long-length DNA hybrid and allows an "off"-"on" genosensor development.

Crystal structure of cbbF from Zymomonas mobilis and its functional implication

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

Hwang, Hyo-Jeong; Park, Suk-Youl; Kim, Jeong-Sun, E-mail: jsunkim@chonnam.ac.kr

2014-02-28

Highlights: • The crystal structure of one cbbF from Zymomonas mobilis was revealed. • Scores of residues form two secondary structures with a non-polar protruded residue. • It exists as a dimeric form in solution. - Abstract: A phosphate group at the C1-atom of inositol-monophosphate (IMP) and fructose-1,6-bisphosphate (FBP) is hydrolyzed by a phosphatase IMPase and FBPase in a metal-dependent way, respectively. The two enzymes are almost indiscernible from each other because of their highly similar sequences and structures. Metal ions are bound to residues on the β1- and β2-strands and one mobile loop. However, FBP has another phosphate andmore » FBPases exist as a higher oligomeric state, which may discriminate FBPases from IMPases. There are three genes annotated as FBPases in Zymomonas mobilis, termed also cbbF (ZmcbbF). The revealed crystal structure of one ZmcbbF shows a globular structure formed by five stacked layers. Twenty-five residues in the middle of the sequence form an α-helix and a β-strand, which occupy one side of the catalytic site. A non-polar Leu residue among them is protruded to the active site, pointing out unfavorable access of a bulky charged group to this side. In vitro assays have shown its dimeric form in solution. Interestingly, two β-strands of β1 and β2 are disordered in the ZmcbbF structure. These data indicate that ZmcbbF might structurally belong to IMPase, and imply that its active site would be reorganized in a yet unreported way.« less

Molecular basis for the wide range of affinity found in Csr/Rsm protein-RNA recognition.

PubMed

Duss, Olivier; Michel, Erich; Diarra dit Konté, Nana; Schubert, Mario; Allain, Frédéric H-T

2014-04-01

The carbon storage regulator/regulator of secondary metabolism (Csr/Rsm) type of small non-coding RNAs (sRNAs) is widespread throughout bacteria and acts by sequestering the global translation repressor protein CsrA/RsmE from the ribosome binding site of a subset of mRNAs. Although we have previously described the molecular basis of a high affinity RNA target bound to RsmE, it remains unknown how other lower affinity targets are recognized by the same protein. Here, we have determined the nuclear magnetic resonance solution structures of five separate GGA binding motifs of the sRNA RsmZ of Pseudomonas fluorescens in complex with RsmE. The structures explain how the variation of sequence and structural context of the GGA binding motifs modulate the binding affinity for RsmE by five orders of magnitude (∼10 nM to ∼3 mM, Kd). Furthermore, we see that conformational adaptation of protein side-chains and RNA enable recognition of different RNA sequences by the same protein contributing to binding affinity without conferring specificity. Overall, our findings illustrate how the variability in the Csr/Rsm protein-RNA recognition allows a fine-tuning of the competition between mRNAs and sRNAs for the CsrA/RsmE protein.

Characterization of an RNA aptamer against HPV-16 L1 virus-like particles.

PubMed

Leija-Montoya, Ana Gabriela; Benítez-Hess, María Luisa; Toscano-Garibay, Julia Dolores; Alvarez-Salas, Luis Marat

2014-10-01

The human papillomavirus (HPV) capsid is mainly composed of the L1 protein that can self-assemble into virus-like particles (VLPs) that are structurally and immunologically similar to the infectious virions. We report here the characterization of RNA aptamers that recognize baculovirus-produced HPV-16 L1 VLPs. Interaction and slot-blot binding assays showed that all isolated aptamers efficiently bound HPV-16 VLPs, although the Sc5-c3 aptamer showed the highest specificity and affinity (Kd=0.05 pM). Sc5-c3 secondary structure consisted of a hairpin with a symmetric bubble and an unstructured 3'end. Biochemical and genetic analyses showed that the Sc5-c3 main loop is directly involved on VLPs binding. In particular, binding specificity appeared mediated by five non-consecutive nucleotide positions. Experiments using bacterial-produced HPV-16 L1 resulted in low Sc5-c3 binding, suggesting that recognition of HPV-16 L1 VLPs relies on quaternary structure features not present in bacteria-produced L1 protein. Sc5-c3 produced specific and stable binding to HPV-16 L1 VLPs even in biofluid protein mixes and thus it may provide a potential diagnostic tool for active HPV infection.

Characterization of an RNA Aptamer Against HPV-16 L1 Virus-Like Particles

PubMed Central

Leija-Montoya, Ana Gabriela; Benítez-Hess, María Luisa; Toscano-Garibay, Julia Dolores

2014-01-01

The human papillomavirus (HPV) capsid is mainly composed of the L1 protein that can self-assemble into virus-like particles (VLPs) that are structurally and immunologically similar to the infectious virions. We report here the characterization of RNA aptamers that recognize baculovirus-produced HPV-16 L1 VLPs. Interaction and slot-blot binding assays showed that all isolated aptamers efficiently bound HPV-16 VLPs, although the Sc5-c3 aptamer showed the highest specificity and affinity (Kd=0.05 pM). Sc5-c3 secondary structure consisted of a hairpin with a symmetric bubble and an unstructured 3′end. Biochemical and genetic analyses showed that the Sc5-c3 main loop is directly involved on VLPs binding. In particular, binding specificity appeared mediated by five non-consecutive nucleotide positions. Experiments using bacterial-produced HPV-16 L1 resulted in low Sc5-c3 binding, suggesting that recognition of HPV-16 L1 VLPs relies on quaternary structure features not present in bacteria-produced L1 protein. Sc5-c3 produced specific and stable binding to HPV-16 L1 VLPs even in biofluid protein mixes and thus it may provide a potential diagnostic tool for active HPV infection. PMID:25111024

Understanding the interactions of different substrates with wild-type and mutant acylaminoacyl peptidase using molecular dynamics simulations.

PubMed

Zhu, Jingxuan; Wang, Yan; Li, Xin; Han, Weiwei; Zhao, Li

2017-12-20

Acylaminoacylpeptidase (AAP) belongs to peptidase protein family, which can degrade amyloid β-peptide forms in the brains of patients, and hence leads to Alzheimer's disease. And so, AAP is considered to be a novel target in the design of drugs against Alzheimer's disease. In this investigation, six molecular dynamics simulations were used to find that the interaction between the wild-type and R526V AAP with two different substrates (p-nitrophenylcaprylate and Ac-Leu-p-nitroanilide). Our results were as follows: firstly, Ac-Leu-p-nitroanilide bound to R526V AAP to form a more disordered loop (residues 552-562) in the α/β-hydrolase fold like of AAP, which caused an open and inactive AAP domain form, secondly, binding p-nitrophenylcaprylate and Ac-Leu-p-nitroanilide to AAP can decrease the flexibility of residues 225-250, 260-270, and 425-450, in which the ordered secondary structures may contain the suitable geometrical structure and so it is useful to serine attack. Our theoretical results showed that the binding of the two substrates can induce specific conformational changes responsible for the diverse AAP catalytic specificity. These theoretical substrate-induced structural diversities can help explain the abilities of AAPs to recognize and hydrolyze extremely different substrates.

SHARP ENTRYWISE PERTURBATION BOUNDS FOR MARKOV CHAINS.

PubMed

Thiede, Erik; VAN Koten, Brian; Weare, Jonathan

For many Markov chains of practical interest, the invariant distribution is extremely sensitive to perturbations of some entries of the transition matrix, but insensitive to others; we give an example of such a chain, motivated by a problem in computational statistical physics. We have derived perturbation bounds on the relative error of the invariant distribution that reveal these variations in sensitivity. Our bounds are sharp, we do not impose any structural assumptions on the transition matrix or on the perturbation, and computing the bounds has the same complexity as computing the invariant distribution or computing other bounds in the literature. Moreover, our bounds have a simple interpretation in terms of hitting times, which can be used to draw intuitive but rigorous conclusions about the sensitivity of a chain to various types of perturbations.

Visualizing polynucleotide polymerase machines at work

PubMed Central

Steitz, Thomas A

2006-01-01

The structures of T7 RNA polymerase (T7 RNAP) captured in the initiation and elongation phases of transcription, that of φ29 DNA polymerase bound to a primer protein and those of the multisubunit RNAPs bound to initiating factors provide insights into how these proteins can initiate RNA synthesis and synthesize 6–10 nucleotides while remaining bound to the site of initiation. Structural insight into the translocation of the product transcript and the separation of the downstream duplex DNA is provided by the structures of the four states of nucleotide incorporation. Single molecule and biochemical studies show a distribution of primer terminus positions that is altered by the binding of NTP and PPi ligands. This article reviews the insights that imaging the structure of polynucleotide polymerases at different steps of the polymerization reaction has provided on the mechanisms of the polymerization reaction. Movies are shown that allow the direct visualization of the conformational changes that the polymerases undergo during the different steps of polymerization. PMID:16900098

Glycine receptor mechanism illuminated by electron cryo-microscopy

PubMed Central

Du, Juan; Lü, Wei; Wu, Shenping; Cheng, Yifan; Gouaux, Eric

2015-01-01

Summary The strychnine-sensitive glycine receptor (GlyR) mediates inhibitory synaptic transmission in the spinal cord and brainstem and is linked to neurological disorders including autism and hyperekplexia. Understanding of molecular mechanisms and pharmacology of GlyRs has been hindered by a dearth of high-resolution structures. Here we report electron cryo-microscopy structures of the α1 GlyR with strychnine, glycine, or glycine/ivermectin. Strychnine arrests the receptor in an antagonist-bound, closed ion channel state, glycine stabilizes the receptor in an agonist-bound open channel state, and the glycine/ivermectin complex adopts a potentially desensitized or partially open state. Relative to the glycine-bound state, strychnine expands the agonist-binding pocket via outward movement of the C loop, promotes rearrangement of the extracellular and transmembrane domain ‘wrist’ interface, and leads to rotation of the transmembrane domain toward the pore axis, occluding the ion conduction pathway. These structures illuminate GlyR mechanism and define a rubric to interpret structures of Cys-loop receptors. PMID:26344198

Glycine receptor mechanism elucidated by electron cryo-microscopy.

PubMed

Du, Juan; Lü, Wei; Wu, Shenping; Cheng, Yifan; Gouaux, Eric

2015-10-08

The strychnine-sensitive glycine receptor (GlyR) mediates inhibitory synaptic transmission in the spinal cord and brainstem and is linked to neurological disorders, including autism and hyperekplexia. Understanding of molecular mechanisms and pharmacology of glycine receptors has been hindered by a lack of high-resolution structures. Here we report electron cryo-microscopy structures of the zebrafish α1 GlyR with strychnine, glycine, or glycine and ivermectin (glycine/ivermectin). Strychnine arrests the receptor in an antagonist-bound closed ion channel state, glycine stabilizes the receptor in an agonist-bound open channel state, and the glycine/ivermectin complex adopts a potentially desensitized or partially open state. Relative to the glycine-bound state, strychnine expands the agonist-binding pocket via outward movement of the C loop, promotes rearrangement of the extracellular and transmembrane domain 'wrist' interface, and leads to rotation of the transmembrane domain towards the pore axis, occluding the ion conduction pathway. These structures illuminate the GlyR mechanism and define a rubric to interpret structures of Cys-loop receptors.

Structural studies of the nudix hydrolase DR1025 from deinococcus radiodurans and its ligand complexes

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

Ranatunga, Wasantha; Hill, Emma E.; Mooster, Jana L.

We have determined the crystal structure, at 1.4, of the Nudix hydrolase DR1025 from the extremely radiation resistant bacterium Deinococcus radiodurans. The protein forms an intertwined homodimer by exchanging N-terminal segments between chains. We have identified additional conserved elements of the Nudix fold, including the metal-binding motif, a kinked b-strand characterized by a proline two positions upstream of the Nudix consensus sequence, and participation of the N-terminal extension in the formation of the substrate-binding pocket. Crystal structures were also solved of DR1025 crystallized in the presence of magnesium and either a GTP analog or Ap4A (both at 1.6 resolution). Inmore » the Ap4Aco-crystal, the electron density indicated that the product of asymmetric hydrolysis, ATP, was bound to the enzyme. The GTP analog bound structure showed that GTP was bound almost identically as ATP. Neither nucleoside triphosphate was further cleaved.« less

Screening actuator locations for static shape control

NASA Technical Reports Server (NTRS)

Haftka, Raphael T.

1990-01-01

Correction of shape distortion due to zero-mean normally distributed errors in structural sizes which are random variables is examined. A bound on the maximum improvement in the expected value of the root-mean-square shape error is obtained. The shape correction associated with the optimal actuators is also characterized. An actuator effectiveness index is developed and shown to be helpful in screening actuator locations in the structure. The results are specialized to a simple form for truss structures composed of nominally identical members. The bound and effectiveness index are tested on a 55-m radiometer antenna truss structure. It is found that previously obtained results for optimum actuators had a performance close to the bound obtained here. Furthermore, the actuators associated with the optimum design are shown to have high effectiveness indices. Since only a small fraction of truss elements tend to have high effectiveness indices, the proposed screening procedure can greatly reduce the number of truss members that need to be considered as actuator sites.

Structural Dynamics of the Magnesium-bound Conformation of CorA in a lipid bilayer

PubMed Central

Dalmas, Olivier; Cuello, Luis G.; Jogini, Vishwanath; Cortes, D. Marien; Roux, Benoit; Perozo, Eduardo

2010-01-01

Summary The transmembrane conformation of Thermotoga maritima CorA, a Magnesium transport system, has been studied in it’s Mg2+-bound form by site-directed spin labeling and electron paramagnetic resonance spectroscopy. Probe mobility together with accessibility data were used to evaluate the overall dynamics and relative arrangement of individual transmembrane segments TM1 and TM2. TM1 extends toward the cytoplasmic side creating a water filled cavity, while TM2 is located in the periphery of the oligomer, contacting the lipid bilayer. A structural model for the conserved extracellular loop was generated based on EPR data and MD simulations, in which residue E316 is located towards the fivefold symmetry axis in position to electrostatically influence divalent ion translocation. Electrostatic analyses of our model suggest that, in agreement with the crystal structure, Mg2+ -bound CorA is in a close conformation. The present results suggest that long-range structural rearrangements are necessary to allow Mg2+ translocation. PMID:20637423

Probing Structural Changes among Analogous Inhibitor-Bound Forms of HIV-1 Protease and a Drug-Resistant Mutant in Solution by Nuclear Magnetic Resonance.

PubMed

Khan, Shahid N; Persons, John D; Paulsen, Janet L; Guerrero, Michel; Schiffer, Celia A; Kurt-Yilmaz, Nese; Ishima, Rieko

2018-03-13

In the era of state-of-the-art inhibitor design and high-resolution structural studies, detection of significant but small protein structural differences in the inhibitor-bound forms is critical to further developing the inhibitor. Here, we probed differences in HIV-1 protease (PR) conformation among darunavir and four analogous inhibitor-bound forms and compared them with a drug-resistant mutant using nuclear magnetic resonance chemical shifts. Changes in amide chemical shifts of wild-type (WT) PR among these inhibitor-bound forms, ΔCSP, were subtle but detectable and extended >10 Å from the inhibitor-binding site, asymmetrically between the two subunits of PR. Molecular dynamics simulations revealed differential local hydrogen bonding as the molecular basis of this remote asymmetric change. Inhibitor-bound forms of the drug-resistant mutant also showed a similar long-range ΔCSP pattern. Differences in ΔCSP values of the WT and the mutant (ΔΔCSPs) were observed at the inhibitor-binding site and in the surrounding region. Comparing chemical shift changes among highly analogous inhibitors and ΔΔCSPs effectively eliminated local environmental effects stemming from different chemical groups and enabled exploitation of these sensitive parameters to detect subtle protein conformational changes and to elucidate asymmetric and remote conformational effects upon inhibitor interaction.

Disorder-function relationships for the cell cycle regulatory proteins p21 and p27.

PubMed

Mitrea, Diana M; Yoon, Mi-Kyung; Ou, Li; Kriwacki, Richard W

2012-04-01

The classic structure-function paradigm has been challenged by a recently identified class of proteins: intrinsically disordered proteins (IDPs). Despite their lack of stable secondary or tertiary structure, IDPs are prevalent in all forms of life and perform myriad cellular functions, including signaling and regulation. Importantly, disruption of IDP homeostasis is associated with numerous human diseases, including cancer and neurodegeneration. Despite wide recognition of IDPs, the molecular mechanisms underlying their functions are not fully understood. Here we review the structural features and disorder-function relationships for p21 and p27, two cyclin-dependent kinase (Cdk) regulators involved in controlling cell division and fate. Studies of p21 bound to Cdk2/cyclin A revealed that a helix stretching mechanism mediates binding promiscuity. Further, investigations of Tyr88-phosphorylated p27 identified a signaling conduit that controls cell division and is disrupted in certain cancers. These mechanisms rely upon a balance between nascent structure in the free state, induced folding upon binding, and persistent flexibility within functional complexes. Although these disorder-function relationships are likely to be recapitulated in other IDPs, it is also likely that the vocabulary of their mechanisms is much more extensive than is currently understood. Further study of the physical properties of IDPs and elucidation of their links with function are needed to fully understand the mechanistic language of IDPs.

Asymmetric Preorganization of Inverted Pair Residues in the Sodium-Calcium Exchanger

PubMed Central

Giladi, Moshe; Almagor, Lior; van Dijk, Liat; Hiller, Reuben; Man, Petr; Forest, Eric; Khananshvili, Daniel

2016-01-01

In analogy with many other proteins, Na+/Ca2+ exchangers (NCX) adapt an inverted twofold symmetry of repeated structural elements, while exhibiting a functional asymmetry by stabilizing an outward-facing conformation. Here, structure-based mutant analyses of the Methanococcus jannaschii Na+/Ca2+ exchanger (NCX_Mj) were performed in conjunction with HDX-MS (hydrogen/deuterium exchange mass spectrometry) to identify the structure-dynamic determinants of functional asymmetry. HDX-MS identified hallmark differences in backbone dynamics at ion-coordinating residues of apo-NCX_Mj, whereas Na+or Ca2+ binding to the respective sites induced relatively small, but specific, changes in backbone dynamics. Mutant analysis identified ion-coordinating residues affecting the catalytic capacity (kcat/Km), but not the stability of the outward-facing conformation. In contrast, distinct “noncatalytic” residues (adjacent to the ion-coordinating residues) control the stability of the outward-facing conformation, but not the catalytic capacity. The helix-breaking signature sequences (GTSLPE) on the α1 and α2 repeats (at the ion-binding core) differ in their folding/unfolding dynamics, while providing asymmetric contributions to transport activities. The present data strongly support the idea that asymmetric preorganization of the ligand-free ion-pocket predefines catalytic reorganization of ion-bound residues, where secondary interactions with adjacent residues couple the alternating access. These findings provide a structure-dynamic basis for ion-coupled alternating access in NCX and similar proteins. PMID:26876271

Complete Mitochondrial Genome Sequence of Acrida cinerea (Acrididae: Orthoptera) and Comparative Analysis of Mitochondrial Genomes in Orthoptera

PubMed Central

Liu, Nian; Huang, Yuan

2010-01-01

The complete 15,599-bp mitogenome of Acrida cinerea was determined and compared with that of the other 20 orthopterans. It displays characteristic gene content, genome organization, nucleotide composition, and codon usage found in other Caelifera mitogenomes. Comparison of 21 orthopteran sequences revealed that the tRNAs encoded by the H-strand appear more conserved than those by the L-stand. All tRNAs form the typical clover-leaf structure except trnS (agn), and most of the size variation among tRNAs stemmed from the length variation in the arm and loop of TΨC and the loop of DHU. The derived secondary structure models of the rrnS and rrnL from 21 orthoptera species closely resemble those from other insects on CRW except a considerably enlarged loop of helix 1399 of rrnS in Caelifera, which is a potentially autapomorphy of Caelifera. In the A+T-rich region, tandem repeats are not only conserved in the closely related mitogenome but also share some conserved motifs in the same subfamily. A stem-loop structure, 16 bp or longer, is likely to be involved in replication initiation in Caelifera and Grylloidea. A long T-stretch (>17 bp) with conserved stem-loop structure next to rrnS on the H-strand, bounded by a purine at either end, exists in the three species from Tettigoniidae. PMID:21197069

Structural studies by X-ray diffraction on metal substituted desulforedoxin, a rubredoxin-type protein.

PubMed Central

Archer, M.; Carvalho, A. L.; Teixeira, S.; Moura, I.; Moura, J. J.; Rusnak, F.; Romão, M. J.

1999-01-01

Desulforedoxin (Dx), isolated from the sulfate reducing bacterium Desulfovibrio gigas, is a small homodimeric (2 x 36 amino acids) protein. Each subunit contains a high-spin iron atom tetrahedrally bound to four cysteinyl sulfur atoms, a metal center similar to that found in rubredoxin (Rd) type proteins. The simplicity of the active center in Dx and the possibility of replacing the iron by other metals make this protein an attractive case for the crystallographic analysis of metal-substituted derivatives. This study extends the relevance of Dx to the bioinorganic chemistry field and is important to obtain model compounds that can mimic the four sulfur coordination of metals in biology. Metal replacement experiments were carried out by reconstituting the apoprotein with In3+, Ga3+, Cd2+, Hg2+, and Ni2+ salts. The In3+ and Ga3+ derivatives are isomorphous with the iron native protein; whereas Cd2+, Hg2+, and Ni2+ substituted Dx crystallized under different experimental conditions, yielding two additional crystal morphologies; their structures were determined by the molecular replacement method. A comparison of the three-dimensional structures for all metal derivatives shows that the overall secondary and tertiary structures are maintained, while some differences in metal coordination geometry occur, namely, bond lengths and angles of the metal with the sulfur ligands. These data are discussed in terms of the entatic state theory. PMID:10422844

Structural mechanism underlying capsaicin binding and activation of TRPV1 ion channel

PubMed Central

Cheng, Wei; Yang, Wei; Yu, Peilin; Song, Zhenzhen; Yarov-Yarovoy, Vladimir; Zheng, Jie

2015-01-01

Capsaicin bestows spiciness by activating TRPV1 channel with exquisite potency and selectivity. Capsaicin-bound channel structure was previously resolved by cryo-EM at 4.2-to-4.5 Å resolution, however important details required for mechanistic understandings are unavailable: capsaicin was registered as a small electron density, reflecting neither its chemical structure nor specific ligand-channel interactions. We obtained the missing atomic-level details by iterative computation, which were confirmed by systematic site-specific functional tests. We observed that the bound capsaicin takes “tail-up, head-down” configurations. The vanillyl and amide groups form specific interactions to anchor its bound position, while the aliphatic tail may sample a range of conformations, making it invisible in cryo-EM images. Capsaicin stabilizes the open state by “pull-and-contact” interactions between the vanillyl group and the S4-S5 linker. Our study provided a structural mechanism for the agonistic function of capsaicin and its analogs, and demonstrated an effective approach to obtain atomic level information from cryo-EM structures. PMID:26053297

Crystal structures of substrate-free and retinoic acid-bound cyanobacterial cytochrome P450 CYP120A1.

PubMed

Kühnel, Karin; Ke, Na; Cryle, Max J; Sligar, Stephen G; Schuler, Mary A; Schlichting, Ilme

2008-06-24

The crystal structures of substrate-free and all-trans-retinoic acid-bound CYP120A1 from Synechocystis sp. PCC 6803 were determined at 2.4 and 2.1 A resolution, respectively, representing the first structural characterization of a cyanobacterial P450. Features of CYP120A1 not observed in other P450 structures include an aromatic ladder flanking the channel leading to the active site and a triple-glycine motif within SRS5. Using spectroscopic methods, CYP120A1 is shown to bind 13-cis-retinoic acid, 9-cis-retinoic acid, and retinal with high affinity and dissociation constants of less than 1 microM. Metabolism of retinoic acid by CYP120A1 suggests that CYP120A1 hydroxylates a variety of retinoid derivatives in vivo. On the basis of the retinoic acid-bound CYP120A1 crystal structure, we propose that either carbon 2 or the methyl groups (C16 or C17) of the beta-ionone ring are modified by CYP120A1.

Finding the Maximal Area of Bounded Polygons in a Circle

ERIC Educational Resources Information Center

Rokach, Arie

2005-01-01

The article deals with the area of polygons that are inscribed in a given circle. Naturally, the following question arises: Among all n-polygons that are inscribed in a given circle, which one has the biggest area? Intuitively, it may be guessed that is suitable for secondary students, and without any use id calculus, but only using very…

A Case Study of Best Practices of Implementing a One-to-One Technology Program

ERIC Educational Resources Information Center

Ackley, Stacy

2017-01-01

The purpose of this case study was to describe the experiences of rural school district leaders who implemented and have sustained a one-to-one technology program at the secondary level so to provide school districts considering one-to-one implementation knowledge of best practices. This study was a qualitative, bounded case study that used a…

Academic Preparedness and College Preparation Efforts: A Comparative Analysis of Perception of High School Advanced Placement Teachers and College Admissions Personnel

ERIC Educational Resources Information Center

Driver, Jason Hughes

2014-01-01

This study examines the relationship between high school efforts at post-secondary preparation for college bound students and the expectations of college and university admissions personnel. The Advanced Placement (AP) teachers and administrators at a rural Northwest Florida high school were administered a quantitative survey in order to collect…

Boosting Student Comprehension of Informational Materials at the Secondary Level at Benjamin Holt College Preparatory Academy

ERIC Educational Resources Information Center

Sampley, Jolyne K.

2008-01-01

Without proficiency in content reading skills, seventh grade students were at risk of not being able to achieve understanding of what they read in non-fiction texts, a skill that is absolutely necessary for college-bound students. The purpose of this action research study was to investigate whether an approach that involved specific and focused…

A Cognitive Analysis of Developmental Mathematics Students' Errors and Misconceptions in Real Number Computations and Evaluating Algebraic Expressions

ERIC Educational Resources Information Center

Titus, Freddie

2010-01-01

Fifty percent of college-bound students graduate from high school underprepared for mathematics at the post-secondary level. As a result, thirty-five percent of college students take developmental mathematics courses. What is even more shocking is the high failure rate (ranging from 35 to 42 percent) of students enrolled in developmental…

A Study of the National Upward Bound and Talent Search Programs. Final Report. Volume III: Descriptive Study of the Talent Search Program.

ERIC Educational Resources Information Center

Pyecha, J. N.; And Others

The Office of Education's Talent Search program is designed to: (a) identify needy youths with exceptional potential and encourage them to complete secondary school and undertake further education; (b) publicize student financial aid; and (c) encourage dropouts of demonstrated aptitude to reenter educational programs. The Talent Search program…

Influence of thermodynamically unfavorable secondary structures on DNA hybridization kinetics

PubMed Central

Hata, Hiroaki; Kitajima, Tetsuro

2018-01-01

Abstract Nucleic acid secondary structure plays an important role in nucleic acid–nucleic acid recognition/hybridization processes, and is also a vital consideration in DNA nanotechnology. Although the influence of stable secondary structures on hybridization kinetics has been characterized, unstable secondary structures, which show positive ΔG° with self-folding, can also form, and their effects have not been systematically investigated. Such thermodynamically unfavorable secondary structures should not be ignored in DNA hybridization kinetics, especially under isothermal conditions. Here, we report that positive ΔG° secondary structures can change the hybridization rate by two-orders of magnitude, despite the fact that their hybridization obeyed second-order reaction kinetics. The temperature dependence of hybridization rates showed non-Arrhenius behavior; thus, their hybridization is considered to be nucleation limited. We derived a model describing how ΔG° positive secondary structures affect hybridization kinetics in stopped-flow experiments with 47 pairs of oligonucleotides. The calculated hybridization rates, which were based on the model, quantitatively agreed with the experimental rate constant. PMID:29220504

piRNA-guided slicing of transposon transcripts enforces their transcriptional silencing via specifying the nuclear piRNA repertoire.

PubMed

Senti, Kirsten-André; Jurczak, Daniel; Sachidanandam, Ravi; Brennecke, Julius

2015-08-15

PIWI clade Argonaute proteins silence transposon expression in animal gonads. Their target specificity is defined by bound ∼23- to 30-nucleotide (nt) PIWI-interacting RNAs (piRNAs) that are processed from single-stranded precursor transcripts via two distinct pathways. Primary piRNAs are defined by the endonuclease Zucchini, while biogenesis of secondary piRNAs depends on piRNA-guided transcript cleavage and results in piRNA amplification. Here, we analyze the interdependencies between these piRNA biogenesis pathways in developing Drosophila ovaries. We show that secondary piRNA-guided target slicing is the predominant mechanism that specifies transcripts—including those from piRNA clusters—as primary piRNA precursors and defines the spectrum of Piwi-bound piRNAs in germline cells. Post-transcriptional silencing in the cytoplasm therefore enforces nuclear transcriptional target silencing, which ensures the tight suppression of transposons during oogenesis. As target slicing also defines the nuclear piRNA pool during mouse spermatogenesis, our findings uncover an unexpected conceptual similarity between the mouse and fly piRNA pathways. © 2015 Senti et al.; Published by Cold Spring Harbor Laboratory Press.

Robust Weighted Sum Harvested Energy Maximization for SWIPT Cognitive Radio Networks Based on Particle Swarm Optimization.

PubMed

Tuan, Pham Viet; Koo, Insoo

2017-10-06

In this paper, we consider multiuser simultaneous wireless information and power transfer (SWIPT) for cognitive radio systems where a secondary transmitter (ST) with an antenna array provides information and energy to multiple single-antenna secondary receivers (SRs) equipped with a power splitting (PS) receiving scheme when multiple primary users (PUs) exist. The main objective of the paper is to maximize weighted sum harvested energy for SRs while satisfying their minimum required signal-to-interference-plus-noise ratio (SINR), the limited transmission power at the ST, and the interference threshold of each PU. For the perfect channel state information (CSI), the optimal beamforming vectors and PS ratios are achieved by the proposed PSO-SDR in which semidefinite relaxation (SDR) and particle swarm optimization (PSO) methods are jointly combined. We prove that SDR always has a rank-1 solution, and is indeed tight. For the imperfect CSI with bounded channel vector errors, the upper bound of weighted sum harvested energy (WSHE) is also obtained through the S-Procedure. Finally, simulation results demonstrate that the proposed PSO-SDR has fast convergence and better performance as compared to the other baseline schemes.

Explicit formula for the Holevo bound for two-parameter qubit-state estimation problem

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

Suzuki, Jun, E-mail: junsuzuki@uec.ac.jp

The main contribution of this paper is to derive an explicit expression for the fundamental precision bound, the Holevo bound, for estimating any two-parameter family of qubit mixed-states in terms of quantum versions of Fisher information. The obtained formula depends solely on the symmetric logarithmic derivative (SLD), the right logarithmic derivative (RLD) Fisher information, and a given weight matrix. This result immediately provides necessary and sufficient conditions for the following two important classes of quantum statistical models; the Holevo bound coincides with the SLD Cramér-Rao bound and it does with the RLD Cramér-Rao bound. One of the important results ofmore » this paper is that a general model other than these two special cases exhibits an unexpected property: the structure of the Holevo bound changes smoothly when the weight matrix varies. In particular, it always coincides with the RLD Cramér-Rao bound for a certain choice of the weight matrix. Several examples illustrate these findings.« less

Bound states in the continuum on periodic structures: perturbation theory and robustness.

PubMed

Yuan, Lijun; Lu, Ya Yan

2017-11-01

On periodic structures, a bound state in the continuum (BIC) is a standing or propagating Bloch wave with a frequency in the radiation continuum. Some BICs (e.g., antisymmetric standing waves) are symmetry protected, since they have incompatible symmetry with outgoing waves in the radiation channels. The propagating BICs do not have this symmetry mismatch, but they still crucially depend on the symmetry of the structure. In this Letter, a perturbation theory is developed for propagating BICs on two-dimensional periodic structures. The Letter shows that these BICs are robust against structural perturbations that preserve the symmetry, indicating that these BICs, in fact, are implicitly protected by symmetry.

Conformational Transitions upon Ligand Binding: Holo-Structure Prediction from Apo Conformations

PubMed Central

Seeliger, Daniel; de Groot, Bert L.

2010-01-01

Biological function of proteins is frequently associated with the formation of complexes with small-molecule ligands. Experimental structure determination of such complexes at atomic resolution, however, can be time-consuming and costly. Computational methods for structure prediction of protein/ligand complexes, particularly docking, are as yet restricted by their limited consideration of receptor flexibility, rendering them not applicable for predicting protein/ligand complexes if large conformational changes of the receptor upon ligand binding are involved. Accurate receptor models in the ligand-bound state (holo structures), however, are a prerequisite for successful structure-based drug design. Hence, if only an unbound (apo) structure is available distinct from the ligand-bound conformation, structure-based drug design is severely limited. We present a method to predict the structure of protein/ligand complexes based solely on the apo structure, the ligand and the radius of gyration of the holo structure. The method is applied to ten cases in which proteins undergo structural rearrangements of up to 7.1 Å backbone RMSD upon ligand binding. In all cases, receptor models within 1.6 Å backbone RMSD to the target were predicted and close-to-native ligand binding poses were obtained for 8 of 10 cases in the top-ranked complex models. A protocol is presented that is expected to enable structure modeling of protein/ligand complexes and structure-based drug design for cases where crystal structures of ligand-bound conformations are not available. PMID:20066034

A Model for Determining Optimal Governance Structure in DoD Acquisition Projects in a Performance-Based Environment

DTIC Science & Technology

2010-04-30

combating market dynamism (Aldrich, 1979; Child, 1972), which is a result of evolving technology, shifting prices, or variance in product availability... principles : (1) human beings are bounded rationally, and (2), as a result of being rationally bound, will always choose to further their own self... principles to govern the relationship among the buyers and suppliers. Our conceptual model aligns the alternative governance structures derived

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

Ren, Aiming; Rajashankar, Kanagalaghatta R.; Patel, Dinshaw J.

Significant advances in our understanding of RNA architecture, folding and recognition have emerged from structure-function studies on riboswitches, non-coding RNAs whose sensing domains bind small ligands and whose adjacent expression platforms contain RNA elements involved in the control of gene regulation. We now report on the ligand-bound structure of the Thermotoga petrophila fluoride riboswitch, which adopts a higher-order RNA architecture stabilized by pseudoknot and long-range reversed Watson-Crick and Hoogsteen A {sm_bullet} U pair formation. The bound fluoride ion is encapsulated within the junctional architecture, anchored in place through direct coordination to three Mg{sup 2+} ions, which in turn are octahedrallymore » coordinated to water molecules and five inwardly pointing backbone phosphates. Our structure of the fluoride riboswitch in the bound state shows how RNA can form a binding pocket selective for fluoride, while discriminating against larger halide ions. The T. petrophila fluoride riboswitch probably functions in gene regulation through a transcription termination mechanism.« less

X-ray structures define human P2X(3) receptor gating cycle and antagonist action.

PubMed

Mansoor, Steven E; Lü, Wei; Oosterheert, Wout; Shekhar, Mrinal; Tajkhorshid, Emad; Gouaux, Eric

2016-10-06

P2X receptors are trimeric, non-selective cation channels activated by ATP that have important roles in the cardiovascular, neuronal and immune systems. Despite their central function in human physiology and although they are potential targets of therapeutic agents, there are no structures of human P2X receptors. The mechanisms of receptor desensitization and ion permeation, principles of antagonism, and complete structures of the pore-forming transmembrane domains of these receptors remain unclear. Here we report X-ray crystal structures of the human P2X 3 receptor in apo/resting, agonist-bound/open-pore, agonist-bound/closed-pore/desensitized and antagonist-bound/closed states. The open state structure harbours an intracellular motif we term the 'cytoplasmic cap', which stabilizes the open state of the ion channel pore and creates lateral, phospholipid-lined cytoplasmic fenestrations for water and ion egress. The competitive antagonists TNP-ATP and A-317491 stabilize the apo/resting state and reveal the interactions responsible for competitive inhibition. These structures illuminate the conformational rearrangements that underlie P2X receptor gating and provide a foundation for the development of new pharmacological agents.

Successful generation of structural information for fragment-based drug discovery.

PubMed

Öster, Linda; Tapani, Sofia; Xue, Yafeng; Käck, Helena

2015-09-01

Fragment-based drug discovery relies upon structural information for efficient compound progression, yet it is often challenging to generate structures with bound fragments. A summary of recent literature reveals that a wide repertoire of experimental procedures is employed to generate ligand-bound crystal structures successfully. We share in-house experience from setting up and executing fragment crystallography in a project that resulted in 55 complex structures. The ligands span five orders of magnitude in affinity and the resulting structures are made available to be of use, for example, for development of computational methods. Analysis of the results revealed that ligand properties such as potency, ligand efficiency (LE) and, to some degree, clogP influence the success of complex structure generation. Copyright © 2015 Elsevier Ltd. All rights reserved.

The dynamics of interacting salt structures and associated fluid flow in the western Norwegian-Danish Basin

NASA Astrophysics Data System (ADS)

Olsen, Mikkel S.; Clausen, Ole R.; Andresen, Katrine J.; Korstgård, John A.

2015-04-01

Minor secondary structures observed along the flanks of major salt structures in the Norwegian-Danish Basin appear to be generated mainly during the early stages of halokinesis. Seismic anomalies in the cover sediments at the flanks of the major salt structures and in relation to one of the secondary structures show several circular patterns. The circular patterns are generally interpreted as faults related to collapsing salt, indicating a subtle and dynamic cannibalization relationship between the secondary structure and the main diapir. High-amplitude reflections interpreted as either entrapped gas along the circular faults or diagenetic changes induced by the fluids originating from the salt-sediment interface generally enhances the seismic appearance of the circular faults, but potentially also disturb the seismic imaging of the faults. Other secondary salt structures, with a similar geometry, do not show sign of collapse, apparently due to a greater distance from the main salt structures and therefore not within the reach of being cannibalized by these. The observations furthermore suggest a trend showing a more advanced development of the main salt structures when the secondary structures are cannibalized. The lateral distribution of the main salt structures thus appears to be controlled not only by the initial thickness of the Zechstein salt, and possible underlying structures, but also by subtle variations in the location and evolution of secondary structures. The secondary structures have a major impact on the drainage of the deep Mesozoic succession as indicated by the fluid flow pattern also observed in the study, which emphasizes that a detailed mapping of salt structures including secondary structures at the flanks is of major importance during evaluation of petroleum systems in areas dominated by halokinesis.

Progesterone binding nano-carriers based on hydrophobically modified hyperbranched polyglycerols

NASA Astrophysics Data System (ADS)

Alizadeh Noghani, M.; Brooks, D. E.

2016-02-01

Progesterone (Pro) is a potent neurosteroid and promotes recovery from moderate Traumatic Brain Injury but its clinical application is severely impeded by its poor water solubility. Here we demonstrate that reversibly binding Pro within hydrophobically modified hyperbranched polyglycerol (HPG-Cn-MPEG) enhances its solubility, stability and bioavailability. Synthesis, characterization and Pro loading into HPG-Cn-MPEG is described. The release kinetics are correlated with structural properties and the results of Differential Scanning Calorimetry studies of a family of HPG-Cn-MPEGs of varying molecular weight and alkylation. While the maximum amount of Pro bound correlates well with the amount of alkyl carbon per molecule contributing to its hydrophobicity, the dominant first order rate constant for Pro release correlates strongly with the amount of structured or bound water in the dendritic domain of the polymer. The results provide evidence to justify more detailed studies of interactions with biological systems, both single cells and in animal models.Progesterone (Pro) is a potent neurosteroid and promotes recovery from moderate Traumatic Brain Injury but its clinical application is severely impeded by its poor water solubility. Here we demonstrate that reversibly binding Pro within hydrophobically modified hyperbranched polyglycerol (HPG-Cn-MPEG) enhances its solubility, stability and bioavailability. Synthesis, characterization and Pro loading into HPG-Cn-MPEG is described. The release kinetics are correlated with structural properties and the results of Differential Scanning Calorimetry studies of a family of HPG-Cn-MPEGs of varying molecular weight and alkylation. While the maximum amount of Pro bound correlates well with the amount of alkyl carbon per molecule contributing to its hydrophobicity, the dominant first order rate constant for Pro release correlates strongly with the amount of structured or bound water in the dendritic domain of the polymer. The results provide evidence to justify more detailed studies of interactions with biological systems, both single cells and in animal models. Electronic supplementary information (ESI) available: Fig. S-1: chemical structure of progesterone (Pro). Fig. S-2: 1H NMR spectrum of HPG-C8-MPEG. Fig. S-3: GPC chromatogram of HPG-C8-MPEG. Fig. S-4: 1H NMR spectrum of HPG-C12-MPEG. Fig. S-5: GPC chromatogram of HPG-C8-MPEG. Fig. S-6: FTIR spectrum of HPG-C8-MPEG. Fig. S-7: inverse-gated 13C NMR spectrum of HPG-C8-MPEG in methanol-d4. Fig. S-8: semi-log plot to determine initial rapid release kinetics for HPG-C8-MPEG/Pro in PBS. Fig. S-9: semi-log plot to determine secondary slow release kinetics for HPG-C8-MPEG/Pro in PBS. Fig. S-10: semi-log plot illustrating the kinetics of Pro release from HPG-C8-MPEG/Pro in plasma. Fig. S-11: dependence of k1 and Vp - Va. Fig. S-12: correlation between the maximum binding capacity of HPG-Cn-MPEG polymeric systems for binding Pro and their total mass of alkyl carbon external to the oxygen (R2 = 0.77 and p < 0.025). Table S-1: effect of loaded Pro on HPG-Cn-MPEG size. Fig. S-13. DLS size determination of HPG-C10-MPEG at 2 mg ml-1 (on the left) and HPG-C10-MPEG/Pro at 2 mg ml-1 of polymer and 25 μg ml-1 of Pro (on the right). The minor population of larger particles was reduced in diameter by Pro binding, illustrated above, consistent with an earlier report.11 See DOI: 10.1039/c5nr08175k

Regularization by Functions of Bounded Variation and Applications to Image Enhancement

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

Casas, E.; Kunisch, K.; Pola, C.

1999-09-15

Optimization problems regularized by bounded variation seminorms are analyzed. The optimality system is obtained and finite-dimensional approximations of bounded variation function spaces as well as of the optimization problems are studied. It is demonstrated that the choice of the vector norm in the definition of the bounded variation seminorm is of special importance for approximating subspaces consisting of piecewise constant functions. Algorithms based on a primal-dual framework that exploit the structure of these nondifferentiable optimization problems are proposed. Numerical examples are given for denoising of blocky images with very high noise.

Hydroxycinnamic acid bound arabinoxylans from millet brans-structural features and antioxidant activity.

PubMed

Bijalwan, Vandana; Ali, Usman; Kesarwani, Atul Kumar; Yadav, Kamalendra; Mazumder, Koushik

2016-07-01

Hydroxycinnamic acid bound arabinoxylans (HCA-AXs) were extracted from brans of five Indian millet varieties and response surface methodology was used to optimize the extraction conditions. The optimal condition to obtain highest yield of millet HCA-AXs was determined as follows: time 61min, temperature 66°C, ratio of solvent to sample 12ml/g. Linkage analysis indicated that hydroxycinnamic acid bound arabinoxylan from kodo millet (KM-HCA-AX) contained comparatively low branched arabinoxylan consisting of 14.6% mono-substituted, 1.2% di-substituted and 41.2% un-substituted Xylp residues. The HPLC analysis of millet HCA-AXs showed significant variation in the content of three major bound hydroxycinnamic acids (caffeic, p-coumaric and ferulic acid). The antioxidant activity of millet HCA-AXs were evaluated using three in vitro assay methods (DPPH, FRAP and β-carotene linoleate emulsion assays) which suggested both phenolic acid composition and structural characteristics of arabinoxylans could be correlated to their antioxidant potential, the detailed structural analysis revealed that low substituted KM-HCA-AX exhibited relatively higher antioxidant activity compared to other medium and highly substituted HCA-AXs from finger (FM), proso (PM), barnyard (BM) and foxtail (FOXM) millet. Copyright © 2016. Published by Elsevier B.V.

Influence of GTP/GDP and magnesium ion on the solvated structure of the protein FtsZ: a molecular dynamics study.

PubMed

Jamous, Carla; Basdevant, Nathalie; Ha-Duong, Tap

2014-01-01

We present here a structural analysis of ten extensive all-atom molecular dynamics simulations of the monomeric protein FtsZ in various binding states. Since the polymerization and GTPase activities of FtsZ depend on the nature of a bound nucleotide as well as on the presence of a magnesium ion, we studied the structural differences between the average conformations of the following five systems: FtsZ-Apo, FtsZ-GTP, FtsZ-GDP, FtsZ-GTP-Mg, and FtsZ-GDP-Mg. The in silico solvated average structure of FtsZ-Apo significantly differs from the crystallographic structure 1W59 of FtsZ which was crystallized in a dimeric form without nucleotide and magnesium. The simulated Apo form of the protein also clearly differs from the FtsZ structures when it is bound to its ligand, the most important discrepancies being located in the loops surrounding the nucleotide binding pocket. The three average structures of FtsZ-GTP, FtsZ-GDP, and FtsZ-GTP-Mg are overall similar, except for the loop T7 located at the opposite side of the binding pocket and whose conformation in FtsZ-GDP notably differs from the one in FtsZ-GTP and FtsZ-GTP-Mg. The presence of a magnesium ion in the binding pocket has no impact on the FtsZ conformation when it is bound to GTP. In contrast, when the protein is bound to GDP, the divalent cation causes a translation of the nucleotide outwards the pocket, inducing a significant conformational change of the loop H6-H7 and the top of helix H7.

Role of DNA secondary structures in fragile site breakage along human chromosome 10

PubMed Central

Dillon, Laura W.; Pierce, Levi C. T.; Ng, Maggie C. Y.; Wang, Yuh-Hwa

2013-01-01

The formation of alternative DNA secondary structures can result in DNA breakage leading to cancer and other diseases. Chromosomal fragile sites, which are regions of the genome that exhibit chromosomal breakage under conditions of mild replication stress, are predicted to form stable DNA secondary structures. DNA breakage at fragile sites is associated with regions that are deleted, amplified or rearranged in cancer. Despite the correlation, unbiased examination of the ability to form secondary structures has not been evaluated in fragile sites. Here, using the Mfold program, we predict potential DNA secondary structure formation on the human chromosome 10 sequence, and utilize this analysis to compare fragile and non-fragile DNA. We found that aphidicolin (APH)-induced common fragile sites contain more sequence segments with potential high secondary structure-forming ability, and these segments clustered more densely than those in non-fragile DNA. Additionally, using a threshold of secondary structure-forming ability, we refined legitimate fragile sites within the cytogenetically defined boundaries, and identified potential fragile regions within non-fragile DNA. In vitro detection of alternative DNA structure formation and a DNA breakage cell assay were used to validate the computational predictions. Many of the regions identified by our analysis coincide with genes mutated in various diseases and regions of copy number alteration in cancer. This study supports the role of DNA secondary structures in common fragile site instability, provides a systematic method for their identification and suggests a mechanism by which DNA secondary structures can lead to human disease. PMID:23297364

Causal analysis of self-sustaining processes in the logarithmic layer of wall-bounded turbulence

NASA Astrophysics Data System (ADS)

Bae, H. J.; Encinar, M. P.; Lozano-Durán, A.

2018-04-01

Despite the large amount of information provided by direct numerical simulations of turbulent flows, their underlying dynamics remain elusive even in the most simple and canonical configurations. Most common approaches to investigate the turbulence phenomena do not provide a clear causal inference between events, which is essential to determine the dynamics of self-sustaining processes. In the present work, we examine the causal interactions between streaks, rolls and mean shear in the logarithmic layer of a minimal turbulent channel flow. Causality between structures is assessed in a non-intrusive manner by transfer entropy, i.e., how much the uncertainty of one structure is reduced by knowing the past states of the others. We choose to represent streaks by the first Fourier modes of the streamwise velocity, while rolls are defined by the wall-normal and spanwise velocity modes. The results show that the process is mainly unidirectional rather than cyclic, and that the log-layer motions are sustained by extracting energy from the mean shear which controls the dynamics and time-scales. The well-known lift-up effect is also identified, but shown to be of secondary importance in the causal network between shear, streaks and rolls.

Formation and internal structure of superdense dark matter clumps and ultracompact minihaloes

NASA Astrophysics Data System (ADS)

Berezinsky, V. S.; Dokuchaev, V. I.; Eroshenko, Yu. N.

2013-11-01

We discuss the formation mechanisms and structure of the superdense dark matter clumps (SDMC) and ultracompact minihaloes (UCMH), outlining the differences between these types of DM objects. We define as SDMC the gravitationally bounded DM objects which have come into virial equilibrium at the radiation-dominated (RD) stage of the universe evolution. Such objects can be formed from the isocurvature (entropy) density perturbations or from the peaks in the spectrum of curvature (adiabatic) perturbation. The axion miniclusters (Kolb and Tkachev 1994) are the example of the former model. The system of central compact mass (e.g. in the form of SDMC or primordial black hole (PBH)) with the outer DM envelope formed in the process of secondary accretion we refer to as UCMH. Therefore, the SDMC can serve as the seed for the UCMH in some scenarios. Recently, the SDMC and UCMH were considered in the many works, and we try to systematize them here. We consider also the effect of asphericity of the initial density perturbation in the gravitational evolution, which decreases the SDMC amount and, as the result, suppresses the gamma-ray signal from DM annihilation.

On the Evolution of Pulsatile Flow Subject to a Transverse Impulse Body Force

NASA Astrophysics Data System (ADS)

di Labbio, Giuseppe; Keshavarz-Motamed, Zahra; Kadem, Lyes

2014-11-01

In the event of an unexpected abrupt traffic stop or car accident, automotive passengers will experience an abrupt body deceleration. This may lead to tearing or dissection of the aortic wall known as Blunt Traumatic Aortic Rupture (BTAR). BTAR is the second leading cause of death in automotive accidents and, although quite frequent, the mechanisms leading to BTAR are still not clearly identified, particularly the contribution of the flow field. As such, this work is intended to provide a fundamental framework for the investigation of the flow contribution to BTAR. In this fundamental study, pulsatile flow in a three-dimensional, straight pipe of circular cross-section is subjected to a unidirectional, transverse, impulse body force applied on a strictly bounded volume of fluid. These models were simulated using the Computational Fluid Dynamics (CFD) software FLUENT. The evolution of fluid field characteristics was investigated during and after the application of the force. The application of the force significantly modified the flow field. The force induces a transverse pressure gradient causing the development of secondary flow structures that dissipate the energy added by the acceleration. Once the force ceases to act, these structures are carried downstream and gradually dissipate their excess energy.

On the hydration of subnanometric antifouling organosilane adlayers: a molecular dynamics simulation.

PubMed

Sheikh, Sonia; Blaszykowski, Christophe; Nolan, Robert; Thompson, Damien; Thompson, Michael

2015-01-01

The connection between antifouling and surface hydration is a fascinating but daunting question to answer. Herein, we use molecular dynamics (MD) computer simulations to gain further insight into the role of surface functionalities in the molecular-level structuration of water (surface kosmotropicity)--within and atop subnanometric organosilane adlayers that were shown in previous experimental work to display varied antifouling behavior. Our simulations support the hypothesized intimate link between surface hydration and antifouling, in particular the importance of both internal and interfacial hydrophilicity and kosmotropicity. The antifouling mechanism is also discussed in terms of surface dehydration energy and water dynamicity (lability and mobility), notably the crucial requirement for clustered water molecules to remain tightly bound for extensive periods of time--i.e. exhibit slow exchange dynamics. A substrate effect on surface hydration, which would also participate in endowing antifouling adlayers with hydrogel-like characteristics, is also proposed. In contrast, the role of adlayer flexibility, if any, is assigned a secondary role in these ultrathin structures made of short building blocks. The conclusions from this work are well in line with those previously drawn in the literature. Copyright © 2014 Elsevier Inc. All rights reserved.

ZrO2 nanoparticles labeled via a native protein corona: detection by fluorescence microscopy and Raman microspectroscopy in rat lungs.

PubMed

Silge, Anja; Bräutigam, Katharina; Bocklitz, Thomas; Rösch, Petra; Vennemann, Antje; Schmitz, Inge; Popp, Jürgen; Wiemann, Martin

2015-08-07

ZrO2 nanoparticles are frequently used in composite materials such as dental fillers from where they may be released and inhaled upon polishing and grinding. Since the overall distribution of ZrO2 NP inside the lung parenchyma can hardly be observed by routine histology, here a labeling with a fluorphore was used secondary to the adsorption of serum proteins. Particles were then intratracheally instilled into rat lungs. After 3 h fluorescent structures consisted of agglomerates scattered throughout the lung parenchyma, which were mainly concentrated in alveolar macrophages after 3 d. A detection method based on Raman microspectroscopy was established to investigate the chemical composition of those fluorescent structures in detail. Raman measurements were arranged such that no spectral interference with the protein-bound fluorescence label was evident. Applying chemometrical methods, Raman signals of the ZrO2 nanomaterial were co-localized with the fluorescence label, indicating the stability of the nanomaterial-protein-dye complex inside the rat lung. The combination of Raman microspectroscopy and adsorptive fluorescence labeling may, therefore, become a useful tool for studying the localization of protein-coated nanomaterials in cells and tissues.

Antimicrobial activities of amphiphilic peptides covalently bonded to a water-insoluble resin.

PubMed Central

Haynie, S L; Crum, G A; Doele, B A

1995-01-01

A series of polymer-bound antimicrobial peptides was prepared, and the peptides were tested for their antimicrobial activities. The immobilized peptides were prepared by a strategy that used solid-phase peptide synthesis that linked the carboxy-terminal amino acid with an ethylenediamine-modified polyamide resin (PepsynK). The acid-stable, permanent amide bond between the support and the nascent peptide renders the peptide resistant to cleavage from the support during the final acid-catalyzed deprotection step in the synthesis. Select immobilized peptides containing amino acid sequences that ranged from the naturally occurring magainin to simpler synthetic sequences with idealized secondary structures were excellent antimicrobial agents against several organisms. The immobilized peptides typically reduced the number of viable cells by > or = 5 log units. We show that the reduction in cell numbers cannot be explained by the action of a soluble component. We observed no leached or hydrolyzed peptide from the resin, nor did we observe any antimicrobial activity in soluble extracts from the immobilized peptide. The immobilized peptides were washed and reused for repeated microbial contact and killing. These results suggest that the surface actions by magainins and structurally related antimicrobial peptides are sufficient for their lethal activities. PMID:7726486

Carbon dioxide is tightly bound in the [Co(Pyridine)(CO2)]- anionic complex

NASA Astrophysics Data System (ADS)

Graham, Jacob D.; Buytendyk, Allyson M.; Zhang, Xinxing; Kim, Seong K.; Bowen, Kit H.

2015-11-01

The [Co(Pyridine)(CO2)]- anionic complex was studied through the combination of photoelectron spectroscopy and density functional theory calculations. This complex was envisioned as a primitive model system for studying CO2 binding to negatively charged sites in metal organic frameworks. The vertical detachment energy (VDE) measured via the photoelectron spectrum is 2.7 eV. Our calculations imply a structure for [Co(Pyridine)(CO2)]- in which a central cobalt atom is bound to pyridine and CO2 moieties on either sides. This structure was validated by acceptable agreement between the calculated and measured VDE values. Based on our calculations, we found CO2 to be bound within the anionic complex by 1.4 eV.

Carbon dioxide is tightly bound in the [Co(Pyridine)(CO2)](-) anionic complex.

PubMed

Graham, Jacob D; Buytendyk, Allyson M; Zhang, Xinxing; Kim, Seong K; Bowen, Kit H

2015-11-14

The [Co(Pyridine)(CO2)](-) anionic complex was studied through the combination of photoelectron spectroscopy and density functional theory calculations. This complex was envisioned as a primitive model system for studying CO2 binding to negatively charged sites in metal organic frameworks. The vertical detachment energy (VDE) measured via the photoelectron spectrum is 2.7 eV. Our calculations imply a structure for [Co(Pyridine)(CO2)](-) in which a central cobalt atom is bound to pyridine and CO2 moieties on either sides. This structure was validated by acceptable agreement between the calculated and measured VDE values. Based on our calculations, we found CO2 to be bound within the anionic complex by 1.4 eV.

Evidence for weakly bound electrons in non-irradiated alkane crystals: The electrons as a probe of structural differences in crystals.

PubMed

Pietrow, M; Gagoś, M; Misiak, L E; Kornarzyński, K; Szurkowski, J; Rochowski, P; Grzegorczyk, M

2015-02-14

It is generally assumed that weakly bound (trapped) electrons in organic solids come only from radiolytical (or photochemical) processes like ionization caused by an excited positron entering the sample. This paper presents evidence for the presence of these electrons in non-irradiated samples of docosane. This can be due to the triboelectrification process. We argue that these electrons can be located (trapped) either in interlamellar gaps or in spaces made by non-planar conformers. Electrons from the former ones are bound more weakly than electrons from the latter ones. The origin of Vis absorption for the samples is explained. These spectra can be used as a probe indicating differences in the solid structures of hydrocarbons.

[Study of "bound insulin" of the blood sera of blood/donors and patients with diabetes mellitus by circular dichroism].

PubMed

Gracheva, N K; Kharitonenkov, I G

1978-01-01

Circular dichroism was applied to the study of the structure of the insulin-transferrin complexes ("bound insulin") isolated from the blood sera of donors and patients suffering from diabetes mellitus of moderate severity. There proved to be a considerable (in comparison with the normal) reduction of the alpha-helix areas in the "bound insulin"molecule of the patients. A comparative study of the circular dichroism spectra in the area of absorption of aromatic amino acids permitted to suppose that the structural changes of the molecule of a complex isolated from the blood sera of patients could not be explained by alterations in the area of the aromatic amino acids.

Building a knowledge-based statistical potential by capturing high-order inter-residue interactions and its applications in protein secondary structure assessment.

PubMed

Li, Yaohang; Liu, Hui; Rata, Ionel; Jakobsson, Eric

2013-02-25

The rapidly increasing number of protein crystal structures available in the Protein Data Bank (PDB) has naturally made statistical analyses feasible in studying complex high-order inter-residue correlations. In this paper, we report a context-based secondary structure potential (CSSP) for assessing the quality of predicted protein secondary structures generated by various prediction servers. CSSP is a sequence-position-specific knowledge-based potential generated based on the potentials of mean force approach, where high-order inter-residue interactions are taken into consideration. The CSSP potential is effective in identifying secondary structure predictions with good quality. In 56% of the targets in the CB513 benchmark, the optimal CSSP potential is able to recognize the native secondary structure or a prediction with Q3 accuracy higher than 90% as best scored in the predicted secondary structures generated by 10 popularly used secondary structure prediction servers. In more than 80% of the CB513 targets, the predicted secondary structures with the lowest CSSP potential values yield higher than 80% Q3 accuracy. Similar performance of CSSP is found on the CASP9 targets as well. Moreover, our computational results also show that the CSSP potential using triplets outperforms the CSSP potential using doublets and is currently better than the CSSP potential using quartets.

Bounds on the information rate of quantum-secret-sharing schemes

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

Sarvepalli, Pradeep

An important metric of the performance of a quantum-secret-sharing scheme is its information rate. Beyond the fact that the information rate is upper-bounded by one, very little is known in terms of bounds on the information rate of quantum-secret-sharing schemes. Furthermore, not every scheme can be realized with rate one. In this paper we derive upper bounds for the information rates of quantum-secret-sharing schemes. We show that there exist quantum access structures on n players for which the information rate cannot be better than O((log{sub 2}n)/n). These results are the quantum analogues of the bounds for classical-secret-sharing schemes proved bymore » Csirmaz.« less

Crystal Structures of Apo and Metal-Bound Forms of the UreE Protein from Helicobacter pylori: Role of Multiple Metal Binding Sites

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

Shi, Rong; Munger, Christine; Asinas, Abdalin

2010-10-22

The crystal structure of the urease maturation protein UreE from Helicobacter pylori has been determined in its apo form at 2.1 {angstrom} resolution, bound to Cu{sup 2+} at 2.7 {angstrom} resolution, and bound to Ni{sup 2+} at 3.1 {angstrom} resolution. Apo UreE forms dimers, while the metal-bound enzymes are arranged as tetramers that consist of a dimer of dimers associated around the metal ion through coordination by His102 residues from each subunit of the tetramer. Comparison of independent subunits from different crystal forms indicates changes in the relative arrangement of the N- and C-terminal domains in response to metal binding.more » The improved ability of engineered versions of UreE containing hexahistidine sequences at either the N-terminal or C-terminal end to provide Ni{sup 2+} for the final metal sink (urease) is eliminated in the H102A version. Therefore, the ability of the improved Ni{sup 2+}-binding versions to deliver more nickel is likely an effect of an increased local concentration of metal ions that can rapidly replenish transferred ions bound to His102.« less

R-chie: a web server and R package for visualizing RNA secondary structures

PubMed Central

Lai, Daniel; Proctor, Jeff R.; Zhu, Jing Yun A.; Meyer, Irmtraud M.

2012-01-01

Visually examining RNA structures can greatly aid in understanding their potential functional roles and in evaluating the performance of structure prediction algorithms. As many functional roles of RNA structures can already be studied given the secondary structure of the RNA, various methods have been devised for visualizing RNA secondary structures. Most of these methods depict a given RNA secondary structure as a planar graph consisting of base-paired stems interconnected by roundish loops. In this article, we present an alternative method of depicting RNA secondary structure as arc diagrams. This is well suited for structures that are difficult or impossible to represent as planar stem-loop diagrams. Arc diagrams can intuitively display pseudo-knotted structures, as well as transient and alternative structural features. In addition, they facilitate the comparison of known and predicted RNA secondary structures. An added benefit is that structure information can be displayed in conjunction with a corresponding multiple sequence alignments, thereby highlighting structure and primary sequence conservation and variation. We have implemented the visualization algorithm as a web server R-chie as well as a corresponding R package called R4RNA, which allows users to run the software locally and across a range of common operating systems. PMID:22434875

Characterising RNA secondary structure space using information entropy

PubMed Central

2013-01-01

Comparative methods for RNA secondary structure prediction use evolutionary information from RNA alignments to increase prediction accuracy. The model is often described in terms of stochastic context-free grammars (SCFGs), which generate a probability distribution over secondary structures. It is, however, unclear how this probability distribution changes as a function of the input alignment. As prediction programs typically only return a single secondary structure, better characterisation of the underlying probability space of RNA secondary structures is of great interest. In this work, we show how to efficiently compute the information entropy of the probability distribution over RNA secondary structures produced for RNA alignments by a phylo-SCFG, and implement it for the PPfold model. We also discuss interpretations and applications of this quantity, including how it can clarify reasons for low prediction reliability scores. PPfold and its source code are available from http://birc.au.dk/software/ppfold/. PMID:23368905

Visualizing the global secondary structure of a viral RNA genome with cryo-electron microscopy

PubMed Central

Garmann, Rees F.; Gopal, Ajaykumar; Athavale, Shreyas S.; Knobler, Charles M.; Gelbart, William M.; Harvey, Stephen C.

2015-01-01

The lifecycle, and therefore the virulence, of single-stranded (ss)-RNA viruses is regulated not only by their particular protein gene products, but also by the secondary and tertiary structure of their genomes. The secondary structure of the entire genomic RNA of satellite tobacco mosaic virus (STMV) was recently determined by selective 2′-hydroxyl acylation analyzed by primer extension (SHAPE). The SHAPE analysis suggested a single highly extended secondary structure with much less branching than occurs in the ensemble of structures predicted by purely thermodynamic algorithms. Here we examine the solution-equilibrated STMV genome by direct visualization with cryo-electron microscopy (cryo-EM), using an RNA of similar length transcribed from the yeast genome as a control. The cryo-EM data reveal an ensemble of branching patterns that are collectively consistent with the SHAPE-derived secondary structure model. Thus, our results both elucidate the statistical nature of the secondary structure of large ss-RNAs and give visual support for modern RNA structure determination methods. Additionally, this work introduces cryo-EM as a means to distinguish between competing secondary structure models if the models differ significantly in terms of the number and/or length of branches. Furthermore, with the latest advances in cryo-EM technology, we suggest the possibility of developing methods that incorporate restraints from cryo-EM into the next generation of algorithms for the determination of RNA secondary and tertiary structures. PMID:25752599

The dynamics of aloof baby Skyrmions

DOE PAGES

Salmi, Petja; Sutcliffe, Paul

2016-01-25

The aloof baby Skyrme model is a (2+1)-dimensional theory with solitons that are lightly bound. It is a low-dimensional analogue of a similar Skyrme model in (3+1)- dimensions, where the lightly bound solitons have binding energies comparable to nuclei. A previous study of static solitons in the aloof baby Skyrme model revealed that multi-soliton bound states have a cluster structure, with constituents that preserve their individual identities due to the short-range repulsion and long-range attraction between solitons. Furthermore, there are many different local energy minima that are all well-described by a simple binary species particle model. In this paper wemore » present the first results on soliton dynamics in the aloof baby Skyrme model. Numerical field theory simulations reveal that the lightly bound cluster structure results in a variety of exotic soliton scattering events that are novel in comparison to standard Skyrmion scattering. A dynamical version of the binary species point particle model is shown to provide a good qualitative description of the dynamics.« less

Robust cooperation of connected vehicle systems with eigenvalue-bounded interaction topologies in the presence of uncertain dynamics

NASA Astrophysics Data System (ADS)

Li, Keqiang; Gao, Feng; Li, Shengbo Eben; Zheng, Yang; Gao, Hongbo

2017-12-01

This study presents a distributed H-infinity control method for uncertain platoons with dimensionally and structurally unknown interaction topologies provided that the associated topological eigenvalues are bounded by a predesigned range.With an inverse model to compensate for nonlinear powertrain dynamics, vehicles in a platoon are modeled by third-order uncertain systems with bounded disturbances. On the basis of the eigenvalue decomposition of topological matrices, we convert the platoon system to a norm-bounded uncertain part and a diagonally structured certain part by applying linear transformation. We then use a common Lyapunov method to design a distributed H-infinity controller. Numerically, two linear matrix inequalities corresponding to the minimum and maximum eigenvalues should be solved. The resulting controller can tolerate interaction topologies with eigenvalues located in a certain range. The proposed method can also ensure robustness performance and disturbance attenuation ability for the closed-loop platoon system. Hardware-in-the-loop tests are performed to validate the effectiveness of our method.

The dynamics of aloof baby Skyrmions

NASA Astrophysics Data System (ADS)

Salmi, Petja; Sutcliffe, Paul

2016-01-01

The aloof baby Skyrme model is a (2+1)-dimensional theory with solitons that are lightly bound. It is a low-dimensional analogue of a similar Skyrme model in (3+1)-dimensions, where the lightly bound solitons have binding energies comparable to nuclei. A previous study of static solitons in the aloof baby Skyrme model revealed that multi-soliton bound states have a cluster structure, with constituents that preserve their individual identities due to the short-range repulsion and long-range attraction between solitons. Furthermore, there are many different local energy minima that are all well-described by a simple binary species particle model. In this paper we present the first results on soliton dynamics in the aloof baby Skyrme model. Numerical field theory simulations reveal that the lightly bound cluster structure results in a variety of exotic soliton scattering events that are novel in comparison to standard Skyrmion scattering. A dynamical version of the binary species point particle model is shown to provide a good qualitative description of the dynamics.

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

Salmi, Petja; Sutcliffe, Paul

The aloof baby Skyrme model is a (2+1)-dimensional theory with solitons that are lightly bound. It is a low-dimensional analogue of a similar Skyrme model in (3+1)- dimensions, where the lightly bound solitons have binding energies comparable to nuclei. A previous study of static solitons in the aloof baby Skyrme model revealed that multi-soliton bound states have a cluster structure, with constituents that preserve their individual identities due to the short-range repulsion and long-range attraction between solitons. Furthermore, there are many different local energy minima that are all well-described by a simple binary species particle model. In this paper wemore » present the first results on soliton dynamics in the aloof baby Skyrme model. Numerical field theory simulations reveal that the lightly bound cluster structure results in a variety of exotic soliton scattering events that are novel in comparison to standard Skyrmion scattering. A dynamical version of the binary species point particle model is shown to provide a good qualitative description of the dynamics.« less

[The changes in contents and composition of phenolic acids during cell xylem growth in scots pine].

PubMed

Antonova, G F; Zheliznichenko, T V; Stasova, V V

2011-01-01

The contents and composition of alcohol soluble phenolic acids were studied during cell xylem growth in the course of wood annual increment formation in the stems of Scots pine. The cells of cambium zone, of two stages of expansion growth and the outset of secondary thickening zone (before lignification) were successively gathered from the stem segments of 25-old pine trees in the period of earlywood xylem formation with constant anatomical and histochemical control. The contents of free and bound forms of phenolic acids, isolated by 80% ethanol from tissues, as well as of their ethers and esters were calculated both per dry weight and per cell. The content and relation of the fractions and the composition of phenolic acid have been found to change significantly from cambium zone to the outset of tracheid secondary thickening. The character of the variations depends on a calculation method. According to the calculation per cell the amount of free and bound phenolic acids and in their composition of esters and especially ethers increased at the first step of expansion growth zone, decreased at the second one and rose again in the outset of secondary wall deposition. In dependence on the stage of cell development the pool of bound phenolic acids exceeded of free acid pool in 2-5 times. Sinapic and ferulic acids dominated in the composition of free hydroxycinnamic acids. The content and composition of hydroxycinnamic acids in ethers and esters depended on cell development phase. In cambium p-coumaric and sinapic acids were principal aglycons in ethers, at other stages these were sinapic and caffeic acids. The esters in cambium zone included essentially p-coumaric acid and at the other stages - sinapic and ferulic acids. At the first phase of growth benzoic acid was connected principally by ester bonds. The pool of these esters decreased from the first phase of growth to the outset of cell wall thickening and in proportion to this the level of free benzoic acid rose.

Crystal structure of a cytochrome P450 2B6 genetic variant in complex with the inhibitor 4-(4-chlorophenyl)imidazole at 2.0-A resolution.

PubMed

Gay, Sean C; Shah, Manish B; Talakad, Jyothi C; Maekawa, Keiko; Roberts, Arthur G; Wilderman, P Ross; Sun, Ling; Yang, Jane Y; Huelga, Stephanie C; Hong, Wen-Xu; Zhang, Qinghai; Stout, C David; Halpert, James R

2010-04-01

The structure of the K262R genetic variant of human cytochrome P450 2B6 in complex with the inhibitor 4-(4-chlorophenyl)imidazole (4-CPI) has been determined using X-ray crystallography to 2.0-A resolution. Production of diffraction quality crystals was enabled through a combination of protein engineering, chaperone coexpression, modifications to the purification protocol, and the use of unique facial amphiphiles during crystallization. The 2B6-4-CPI complex is virtually identical to the rabbit 2B4 structure bound to the same inhibitor with respect to the arrangement of secondary structural elements and the placement of active site residues. The structure supports prior P450 2B6 homology models based on other mammalian cytochromes P450 and is consistent with the limited site-directed mutagenesis studies on 2B6 and extensive studies on P450 2B4 and 2B1. Although the K262R genetic variant shows unaltered binding of 4-CPI, altered binding affinity, kinetics, and/or product profiles have been previously shown with several other ligands. On the basis of new P450 2B6 crystal structure and previous 2B4 structures, substitutions at residue 262 affect a hydrogen-bonding network connecting the G and H helices, where subtle differences could be transduced to the active site. Docking experiments indicate that the closed protein conformation allows smaller ligands such as ticlopidine to bind to the 2B6 active site in the expected orientation. However, it is unknown whether 2B6 undergoes structural reorganization to accommodate bulkier molecules, as previously inferred from multiple P450 2B4 crystal structures.

Retention and chemical speciation of uranium in an oxidized wetland sediment from the Savannah River Site.

PubMed

Li, Dien; Seaman, John C; Chang, Hyun-Shik; Jaffe, Peter R; Koster van Groos, Paul; Jiang, De-Tong; Chen, Ning; Lin, Jinru; Arthur, Zachary; Pan, Yuanming; Scheckel, Kirk G; Newville, Matthew; Lanzirotti, Antonio; Kaplan, Daniel I

2014-05-01

Uranium speciation and retention mechanisms onto Savannah River Site (SRS) wetland sediments was studied using batch (ad)sorption experiments, sequential extraction, U L3-edge X-ray absorption near-edge structure (XANES) spectroscopy, fluorescence mapping and μ-XANES. Under oxidized conditions, U was highly retained by the SRS wetland sediments. In contrast to other similar but much lower natural organic matter (NOM) sediments, significant sorption of U onto the SRS sediments was observed at pH < 4 and pH > 8. Sequential extraction indicated that the U species were primarily associated with the acid soluble fraction (weak acetic acid extractable) and organic fraction (Na-pyrophosphate extractable). Uranium L3-edge XANES spectra of the U-bound sediments were nearly identical to that of uranyl acetate. Based on fluorescence mapping, U and Fe distributions in the sediment were poorly correlated, U was distributed throughout the sample and did not appear as isolated U mineral phases. The primary oxidation state of U in these oxidized sediments was U(VI), and there was little evidence that the high sorptive capacity of the sediments could be ascribed to abiotic or biotic reduction to the less soluble U(IV) species or to secondary mineral formation. Collectively, this study suggests that U may be strongly bound to wetland sediments, not only under reducing conditions by reductive precipitation, but also under oxidizing conditions through NOM-uranium bonding. Published by Elsevier Ltd.

BeStSel: a web server for accurate protein secondary structure prediction and fold recognition from the circular dichroism spectra.

PubMed

Micsonai, András; Wien, Frank; Bulyáki, Éva; Kun, Judit; Moussong, Éva; Lee, Young-Ho; Goto, Yuji; Réfrégiers, Matthieu; Kardos, József

2018-06-11

Circular dichroism (CD) spectroscopy is a widely used method to study the protein secondary structure. However, for decades, the general opinion was that the correct estimation of β-sheet content is challenging because of the large spectral and structural diversity of β-sheets. Recently, we showed that the orientation and twisting of β-sheets account for the observed spectral diversity, and developed a new method to estimate accurately the secondary structure (PNAS, 112, E3095). BeStSel web server provides the Beta Structure Selection method to analyze the CD spectra recorded by conventional or synchrotron radiation CD equipment. Both normalized and measured data can be uploaded to the server either as a single spectrum or series of spectra. The originality of BeStSel is that it carries out a detailed secondary structure analysis providing information on eight secondary structure components including parallel-β structure and antiparallel β-sheets with three different groups of twist. Based on these, it predicts the protein fold down to the topology/homology level of the CATH protein fold classification. The server also provides a module to analyze the structures deposited in the PDB for BeStSel secondary structure contents in relation to Dictionary of Secondary Structure of Proteins data. The BeStSel server is freely accessible at http://bestsel.elte.hu.

Application of Ring-Closing Metathesis to Grb2 SH3 Domain-Binding Peptides | Center for Cancer Research

Cancer.gov

In silico-generated hypothetical interactions of a ring-closing metathesis-macrocylized peptide bound to the amino terminal SH3 domain of the growth factor receptor bound protein 2 (Grb2). The complex was derived from the NMR solution structure of the bound parent peptide, Ac-V-P-P-P-V-P-P-R-R-R-amide (Protein Data Bank: 3GBQ). The protein surface is shown as electrostatic

Structurally coloured secondary particles composed of black and white colloidal particles.

PubMed

Takeoka, Yukikazu; Yoshioka, Shinya; Teshima, Midori; Takano, Atsushi; Harun-Ur-Rashid, Mohammad; Seki, Takahiro

2013-01-01

This study investigated the colourful secondary particles formed by controlling the aggregation states of colloidal silica particles and the enhancement of the structural colouration of the secondary particles caused by adding black particles. We obtained glossy, partially structurally coloured secondary particles in the absence of NaCl, but matte, whitish secondary particles were obtained in the presence of NaCl. When a small amount of carbon black was incorporated into both types of secondary particles, the incoherent multiple scattering of light from the amorphous region was considerably reduced. However, the peak intensities in the reflection spectra, caused by Bragg reflection and by coherent single wavelength scattering, were only slightly decreased. Consequently, a brighter structural colour of these secondary particles was observed with the naked eye. Furthermore, when magnetite was added as a black particle, the coloured secondary particles could be moved and collected by applying an external magnetic field.

Structurally Coloured Secondary Particles Composed of Black and White Colloidal Particles

PubMed Central

Takeoka, Yukikazu; Yoshioka, Shinya; Teshima, Midori; Takano, Atsushi; Harun-Ur-Rashid, Mohammad; Seki, Takahiro

2013-01-01

This study investigated the colourful secondary particles formed by controlling the aggregation states of colloidal silica particles and the enhancement of the structural colouration of the secondary particles caused by adding black particles. We obtained glossy, partially structurally coloured secondary particles in the absence of NaCl, but matte, whitish secondary particles were obtained in the presence of NaCl. When a small amount of carbon black was incorporated into both types of secondary particles, the incoherent multiple scattering of light from the amorphous region was considerably reduced. However, the peak intensities in the reflection spectra, caused by Bragg reflection and by coherent single wavelength scattering, were only slightly decreased. Consequently, a brighter structural colour of these secondary particles was observed with the naked eye. Furthermore, when magnetite was added as a black particle, the coloured secondary particles could be moved and collected by applying an external magnetic field. PMID:23917891

Analysis of protein circular dichroism spectra for secondary structure using a simple matrix multiplication.

PubMed

Compton, L A; Johnson, W C

1986-05-15

Inverse circular dichroism (CD) spectra are presented for each of the five major secondary structures of proteins: alpha-helix, antiparallel and parallel beta-sheet, beta-turn, and other (random) structures. The fraction of the each secondary structure in a protein is predicted by forming the dot product of the corresponding inverse CD spectrum, expressed as a vector, with the CD spectrum of the protein digitized in the same way. We show how this method is based on the construction of the generalized inverse from the singular value decomposition of a set of CD spectra corresponding to proteins whose secondary structures are known from X-ray crystallography. These inverse spectra compute secondary structure directly from protein CD spectra without resorting to least-squares fitting and standard matrix inversion techniques. In addition, spectra corresponding to the individual secondary structures, analogous to the CD spectra of synthetic polypeptides, are generated from the five most significant CD eigenvectors.

Structure of Mandelate Racemase with Bound Intermediate Analogues Benzohydroxamate and Cupferron†

PubMed Central

Lietzan, Adam D.; Nagar, Mitesh; Pellmann, Elise A.; Bourque, Jennifer R.; Bearne, Stephen L.; St Maurice, Martin

2012-01-01

Mandelate racemase (MR, EC 5.1.2.2) from Pseudomonas putida catalyzes the Mg2+-dependent interconversion of the enantiomers of mandelate, stabilizing the altered substrate in the transition state by 26 kcal/mol relative to the substrate in the ground state. To understand the origins of this binding discrimination, we solved the X-ray crystal structures of wild-type MR complexed with two analogues of the putative aci-carboxylate intermediate, benzohydroxamate and cupferron, to 2.2-Å resolution. Benzohydroxamate is shown to be a reasonable mimic of the transition state/intermediate since its binding affinity to 21 MR variants correlates well with changes in the free energy of transition state stabilization afforded by these variants. Both benzohydroxamate and cupferron chelate the active site divalent metal ion and are bound in a conformation with the phenyl ring coplanar with the hydroxamate and diazeniumdiolate moieties, respectively. Structural overlays of MR complexed with benzohydroxamate, cupferron, and the ground state analogue (S)-atrolacatate reveal that the para-carbon of the substrate phenyl ring moves by 0.8–1.2 Å between the ground state and intermediate state, consistent with the proposal that the phenyl ring moves during MR catalysis while the polar groups remain relatively fixed. Although the overall protein structure of MR with bound intermediate analogues is very similar to MR with bound (S)-atrolactate, the intermediate-Mg2+ distance shortens, suggesting a tighter complex with the catalytic Mg2+. In addition, Tyr 54 moves nearer to the phenyl ring of the bound intermediate analogues, contributing to an overall constriction of the active site cavity. However, site-directed mutagenesis experiments revealed that the role of Tyr 54 in MR catalysis is relatively minor, suggesting that alterations in enzyme structure that contribute to discrimination between the altered substrate in the transition state and the ground state by this proficient enzyme are extremely subtle. PMID:22264153

A statistical learning approach to the modeling of chromatographic retention of oligonucleotides incorporating sequence and secondary structure data

PubMed Central

Sturm, Marc; Quinten, Sascha; Huber, Christian G.; Kohlbacher, Oliver

2007-01-01

We propose a new model for predicting the retention time of oligonucleotides. The model is based on ν support vector regression using features derived from base sequence and predicted secondary structure of oligonucleotides. Because of the secondary structure information, the model is applicable even at relatively low temperatures where the secondary structure is not suppressed by thermal denaturing. This makes the prediction of oligonucleotide retention time for arbitrary temperatures possible, provided that the target temperature lies within the temperature range of the training data. We describe different possibilities of feature calculation from base sequence and secondary structure, present the results and compare our model to existing models. PMID:17567619

Fatty acids bound to recombinant tear lipocalin and their role in structural stabilization.

PubMed

Tsukamoto, Seiichi; Fujiwara, Kazuo; Ikeguchi, Masamichi

2009-09-01

A variant of human tear lipocalin was expressed in Escherichia coli, and the bound fatty acids were analysed by gas chromatography, mass spectroscopy and nuclear magnetic resonance spectroscopy. Five major fatty acids were identified as hexadecanoic acid (palmitic acid, PA), cis-9-hexadecenoic acid (palmitoleic acid), 9,10-methylenehexadecanoic acid, cis-11-octadecenoic acid (vaccenic acid) and 11,12-methyleneoctadecanoic acid (lactobacillic acid). The composition of the bound fatty acids was similar to the fatty acid composition of E. coli extract, suggesting that the binding affinities are similar for these fatty acids. The urea-induced and thermal-unfolding transitions of the holoprotein (nondelipidated), apoprotein (delipidated) and PA-bound protein were observed by circular dichroism. Holoproteins and PA-bound proteins showed the same stability against urea and heat, and were more stable than apoprotein. These results show that each bound fatty acid stabilizes recombinant tear lipocalin to a similar extent.

Bounded Linear Stability Analysis - A Time Delay Margin Estimation Approach for Adaptive Control

NASA Technical Reports Server (NTRS)

Nguyen, Nhan T.; Ishihara, Abraham K.; Krishnakumar, Kalmanje Srinlvas; Bakhtiari-Nejad, Maryam

2009-01-01

This paper presents a method for estimating time delay margin for model-reference adaptive control of systems with almost linear structured uncertainty. The bounded linear stability analysis method seeks to represent the conventional model-reference adaptive law by a locally bounded linear approximation within a small time window using the comparison lemma. The locally bounded linear approximation of the combined adaptive system is cast in a form of an input-time-delay differential equation over a small time window. The time delay margin of this system represents a local stability measure and is computed analytically by a matrix measure method, which provides a simple analytical technique for estimating an upper bound of time delay margin. Based on simulation results for a scalar model-reference adaptive control system, both the bounded linear stability method and the matrix measure method are seen to provide a reasonably accurate and yet not too conservative time delay margin estimation.

Microstructure and phase composition characterization in a Co{sub 38}Ni{sub 33}Al{sub 29} ferromagnetic shape memory alloy

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

Lu, J.B.

2016-08-15

Transmission electron microscopy was performed to investigate the microstructures of a secondary phase and its surrounding matrix in a Co{sub 38}Ni{sub 33}Al{sub 29} ferromagnetic shape memory alloy. The secondary phase shows a γ′ L1{sub 2} structure exhibiting a dendritic morphology with enclosed B2 austenite regions while the matrix shows the L1{sub 0} martensitic structure. A secondary phase-austenite-martensite sandwich structure with residual austenite ranging from several hundred nanometers to several micrometers wide is observed at the secondary phase-martensite interface due to the depletion of Co and enrichment of Al in the chemical gradient zone and the effect of the strong martensiticmore » start temperature dependency of the element concentrations. The crystallographic orientation relationship of the secondary phase and the B2 austenite fits the Kurdjumov-Sachs relationship. - Highlights: •The secondary phase has a γ′ L1{sub 2} structure exhibiting a dendritic morphology. •A secondary phase-austenite-martensite sandwich structure is observed. •The structural sandwich structure is due to elemental composition variation. •The secondary phase and the B2 austenite fit the Kurdjumov-Sachs relationship.« less

Free and ATP-bound structures of Ap4A hydrolase from Aquifex aeolicus V5.

PubMed

Jeyakanthan, Jeyaraman; Kanaujia, Shankar Prasad; Nishida, Yuya; Nakagawa, Noriko; Praveen, Surendran; Shinkai, Akeo; Kuramitsu, Seiki; Yokoyama, Shigeyuki; Sekar, Kanagaraj

2010-02-01

Asymmetric diadenosine tetraphosphate (Ap(4)A) hydrolases degrade the metabolite Ap(4)A back into ATP and AMP. The three-dimensional crystal structure of Ap(4)A hydrolase (16 kDa) from Aquifex aeolicus has been determined in free and ATP-bound forms at 1.8 and 1.95 A resolution, respectively. The overall three-dimensional crystal structure of the enzyme shows an alphabetaalpha-sandwich architecture with a characteristic loop adjacent to the catalytic site of the protein molecule. The ATP molecule is bound in the primary active site and the adenine moiety of the nucleotide binds in a ring-stacking arrangement equivalent to that observed in the X-ray structure of Ap(4)A hydrolase from Caenorhabditis elegans. Binding of ATP in the active site induces local conformational changes which may have important implications in the mechanism of substrate recognition in this class of enzymes. Furthermore, two invariant water molecules have been identified and their possible structural and/or functional roles are discussed. In addition, modelling of the substrate molecule at the primary active site of the enzyme suggests a possible path for entry and/or exit of the substrate and/or product molecule.

Noncoding RNA. piRNA-guided transposon cleavage initiates Zucchini-dependent, phased piRNA production.

PubMed

Han, Bo W; Wang, Wei; Li, Chengjian; Weng, Zhiping; Zamore, Phillip D

2015-05-15

PIWI-interacting RNAs (piRNAs) protect the animal germ line by silencing transposons. Primary piRNAs, generated from transcripts of genomic transposon "junkyards" (piRNA clusters), are amplified by the "ping-pong" pathway, yielding secondary piRNAs. We report that secondary piRNAs, bound to the PIWI protein Ago3, can initiate primary piRNA production from cleaved transposon RNAs. The first ~26 nucleotides (nt) of each cleaved RNA becomes a secondary piRNA, but the subsequent ~26 nt become the first in a series of phased primary piRNAs that bind Piwi, allowing piRNAs to spread beyond the site of RNA cleavage. The ping-pong pathway increases only the abundance of piRNAs, whereas production of phased primary piRNAs from cleaved transposon RNAs adds sequence diversity to the piRNA pool, allowing adaptation to changes in transposon sequence. Copyright © 2015, American Association for the Advancement of Science.

Chromatin insulator bodies are nuclear structures that form in response to osmotic stress and cell death

PubMed Central

Schoborg, Todd; Rickels, Ryan; Barrios, Josh

2013-01-01

Chromatin insulators assist in the formation of higher-order chromatin structures by mediating long-range contacts between distant genomic sites. It has been suggested that insulators accomplish this task by forming dense nuclear foci termed insulator bodies that result from the coalescence of multiple protein-bound insulators. However, these structures remain poorly understood, particularly the mechanisms triggering body formation and their role in nuclear function. In this paper, we show that insulator proteins undergo a dramatic and dynamic spatial reorganization into insulator bodies during osmostress and cell death in a high osmolarity glycerol–p38 mitogen-activated protein kinase–independent manner, leading to a large reduction in DNA-bound insulator proteins that rapidly repopulate chromatin as the bodies disassemble upon return to isotonicity. These bodies occupy distinct nuclear territories and contain a defined structural arrangement of insulator proteins. Our findings suggest insulator bodies are novel nuclear stress foci that can be used as a proxy to monitor the chromatin-bound state of insulator proteins and provide new insights into the effects of osmostress on nuclear and genome organization. PMID:23878275

The turn of the screw: an exercise in protein secondary structure.

PubMed

Pikaart, Michael

2011-01-01

An exercise using simple paper strips to illustrate protein helical and sheet secondary structures is presented. Drawing on the rich historical context of the use of physical models in protein biochemistry by early practitioners, in particular Linus Pauling, the purpose of this activity is to cultivate in students a hands-on, intuitive sense of protein secondary structure and to complement the common computer-based structural portrayals often used in teaching biochemistry. As students fold these paper strips into model secondary structures, they will better grasp how intramolecular hydrogen bonds form in the folding of a polypeptide into secondary structure, and how these hydrogen bonds direct the overall shape of helical and sheet structures, including the handedness of the α-helix and the difference between right- and the left-handed twist. Copyright © 2010 Wiley Periodicals, Inc.

VMD-SS: A graphical user interface plug-in to calculate the protein secondary structure in VMD program.

PubMed

Yahyavi, Masoumeh; Falsafi-Zadeh, Sajad; Karimi, Zahra; Kalatarian, Giti; Galehdari, Hamid

2014-01-01

The investigation on the types of secondary structure (SS) of a protein is important. The evolution of secondary structures during molecular dynamics simulations is a useful parameter to analyze protein structures. Therefore, it is of interest to describe VMD-SS (a software program) for the identification of secondary structure elements and its trajectories during simulation for known structures available at the Protein Data Bank (PDB). The program helps to calculate (1) percentage SS, (2) SS occurrence in each residue, (3) percentage SS during simulation, and (4) percentage residues in all SS types during simulation. The VMD-SS plug-in was designed using TCL script and stride to calculate secondary structure features. The database is available for free at http://science.scu.ac.ir/HomePage.aspx?TabID=13755.

Conservation of mRNA secondary structures may filter out mutations in Escherichia coli evolution

PubMed Central

Chursov, Andrey; Frishman, Dmitrij; Shneider, Alexander

2013-01-01

Recent reports indicate that mutations in viral genomes tend to preserve RNA secondary structure, and those mutations that disrupt secondary structural elements may reduce gene expression levels, thereby serving as a functional knockout. In this article, we explore the conservation of secondary structures of mRNA coding regions, a previously unknown factor in bacterial evolution, by comparing the structural consequences of mutations in essential and nonessential Escherichia coli genes accumulated over 40 000 generations in the course of the ‘long-term evolution experiment’. We monitored the extent to which mutations influence minimum free energy (MFE) values, assuming that a substantial change in MFE is indicative of structural perturbation. Our principal finding is that purifying selection tends to eliminate those mutations in essential genes that lead to greater changes of MFE values and, therefore, may be more disruptive for the corresponding mRNA secondary structures. This effect implies that synonymous mutations disrupting mRNA secondary structures may directly affect the fitness of the organism. These results demonstrate that the need to maintain intact mRNA structures imposes additional evolutionary constraints on bacterial genomes, which go beyond preservation of structure and function of the encoded proteins. PMID:23783573

Upward Bound: In the Beginning.

ERIC Educational Resources Information Center

Groutt, John; Hill, Calvin

2001-01-01

Describes the early history of the Upward Bound program, including the role of President Johnson's vision, the Task Force on Poverty, the Office of Economic Opportunity, and Community Action Programs; influences on the development of the program; establishment of the program's administrative structure; pilot programs; and early problems leading to…

Size-dependent impact of CNTs on dynamic properties of calmodulin

NASA Astrophysics Data System (ADS)

Gao, Jian; Wang, Liming; Kang, Seung-Gu; Zhao, Lina; Ji, Mingjuan; Chen, Chunying; Zhao, Yuliang; Zhou, Ruhong; Li, Jingyuan

2014-10-01

There are growing concerns about the biosafety of nanomaterials such as carbon nanotubes (CNTs) as their applications become more widespread. We report here a theoretical and experimental study of the binding of various sizes of CNTs [CNT (4,4), (5,5), (6,6) and (7,7)] to calmodulin (CaM) protein and, in particular, their impact on the Ca2+-dependent dynamic properties of CaM. Our simulations show that all the CNTs can plug into the hydrophobic binding pocket of Ca2+-bound CaM with binding affinities comparable with the native substrate M13 peptide. Even though CNT (4,4) shows a similar behavior to the M13 peptide in its dissociation from Ca2+-free CaM, wider CNTs still bind firmly to CaM, indicating a potential failure of Ca2+ regulation. Such a size-dependent impact of CNTs on the dynamic properties of CaM is a result of the excessively strong hydrophobic interactions between the wider CNTs and CaM. These simulation results were confirmed by circular dichroism spectroscopy, which showed that the secondary structures of CaM become insensitive to Ca2+ concentrations after the addition of CNTs. Our findings indicate that the cytotoxicity of nanoparticles to proteins arises not only from the inhibition of static protein structures (binding pockets), but also from impacts on their dynamic properties.There are growing concerns about the biosafety of nanomaterials such as carbon nanotubes (CNTs) as their applications become more widespread. We report here a theoretical and experimental study of the binding of various sizes of CNTs [CNT (4,4), (5,5), (6,6) and (7,7)] to calmodulin (CaM) protein and, in particular, their impact on the Ca2+-dependent dynamic properties of CaM. Our simulations show that all the CNTs can plug into the hydrophobic binding pocket of Ca2+-bound CaM with binding affinities comparable with the native substrate M13 peptide. Even though CNT (4,4) shows a similar behavior to the M13 peptide in its dissociation from Ca2+-free CaM, wider CNTs still bind firmly to CaM, indicating a potential failure of Ca2+ regulation. Such a size-dependent impact of CNTs on the dynamic properties of CaM is a result of the excessively strong hydrophobic interactions between the wider CNTs and CaM. These simulation results were confirmed by circular dichroism spectroscopy, which showed that the secondary structures of CaM become insensitive to Ca2+ concentrations after the addition of CNTs. Our findings indicate that the cytotoxicity of nanoparticles to proteins arises not only from the inhibition of static protein structures (binding pockets), but also from impacts on their dynamic properties. Electronic supplementary information (ESI) available. See DOI: 10.1039/c4nr01623h

Structured and Unstructured Binding of an Intrinsically Disordered Protein as Revealed by Atomistic Simulations.

PubMed

Ithuralde, Raúl Esteban; Roitberg, Adrián Enrique; Turjanski, Adrián Gustavo

2016-07-20

Intrinsically disordered proteins (IDPs) are a set of proteins that lack a definite secondary structure in solution. IDPs can acquire tertiary structure when bound to their partners; therefore, the recognition process must also involve protein folding. The nature of the transition state (TS), structured or unstructured, determines the binding mechanism. The characterization of the TS has become a major challenge for experimental techniques and molecular simulations approaches since diffusion, recognition, and binding is coupled to folding. In this work we present atomistic molecular dynamics (MD) simulations that sample the free energy surface of the coupled folding and binding of the transcription factor c-myb to the cotranscription factor CREB binding protein (CBP). This process has been recently studied and became a model to study IDPs. Despite the plethora of available information, we still do not know how c-myb binds to CBP. We performed a set of atomistic biased MD simulations running a total of 15.6 μs. Our results show that c-myb folds very fast upon binding to CBP with no unique pathway for binding. The process can proceed through both structured or unstructured TS's with similar probabilities. This finding reconciles previous seemingly different experimental results. We also performed Go-type coarse-grained MD of several structured and unstructured models that indicate that coupled folding and binding follows a native contact mechanism. To the best of our knowledge, this is the first atomistic MD simulation that samples the free energy surface of the coupled folding and binding processes of IDPs.

Soluble expression, purification and characterization of the full length IS2 Transposase.

PubMed

Lewis, Leslie A; Astatke, Mekbib; Umekubo, Peter T; Alvi, Shaheen; Saby, Robert; Afrose, Jehan

2011-10-27

The two-step transposition pathway of insertion sequences of the IS3 family, and several other families, involves first the formation of a branched figure-of-eight (F-8) structure by an asymmetric single strand cleavage at one optional donor end and joining to the flanking host DNA near the target end. Its conversion to a double stranded minicircle precedes the second insertional step, where both ends function as donors. In IS2, the left end which lacks donor function in Step I acquires it in Step II. The assembly of two intrinsically different protein-DNA complexes in these F-8 generating elements has been intuitively proposed, but a barrier to testing this hypothesis has been the difficulty of isolating a full length, soluble and active transposase that creates fully formed synaptic complexes in vitro with protein bound to both binding and catalytic domains of the ends. We address here a solution to expressing, purifying and structurally analyzing such a protein. A soluble and active IS2 transposase derivative with GFP fused to its C-terminus functions as efficiently as the native protein in in vivo transposition assays. In vitro electrophoretic mobility shift assay data show that the partially purified protein prepared under native conditions binds very efficiently to cognate DNA, utilizing both N- and C-terminal residues. As a precursor to biophysical analyses of these complexes, a fluorescence-based random mutagenesis protocol was developed that enabled a structure-function analysis of the protein with good resolution at the secondary structure level. The results extend previous structure-function work on IS3 family transposases, identifying the binding domain as a three helix H + HTH bundle and explaining the function of an atypical leucine zipper-like motif in IS2. In addition gain- and loss-of-function mutations in the catalytic active site define its role in regional and global binding and identify functional signatures that are common to the three dimensional catalytic core motif of the retroviral integrase superfamily. Intractably insoluble transposases, such as the IS2 transposase, prepared by solubilization protocols are often refractory to whole protein structure-function studies. The results described here have validated the use of GFP-tagging and fluorescence-based random mutagenesis in overcoming this limitation at the secondary structure level.

Soluble expression, purification and characterization of the full length IS2 Transposase

PubMed Central

2011-01-01

Background The two-step transposition pathway of insertion sequences of the IS3 family, and several other families, involves first the formation of a branched figure-of-eight (F-8) structure by an asymmetric single strand cleavage at one optional donor end and joining to the flanking host DNA near the target end. Its conversion to a double stranded minicircle precedes the second insertional step, where both ends function as donors. In IS2, the left end which lacks donor function in Step I acquires it in Step II. The assembly of two intrinsically different protein-DNA complexes in these F-8 generating elements has been intuitively proposed, but a barrier to testing this hypothesis has been the difficulty of isolating a full length, soluble and active transposase that creates fully formed synaptic complexes in vitro with protein bound to both binding and catalytic domains of the ends. We address here a solution to expressing, purifying and structurally analyzing such a protein. Results A soluble and active IS2 transposase derivative with GFP fused to its C-terminus functions as efficiently as the native protein in in vivo transposition assays. In vitro electrophoretic mobility shift assay data show that the partially purified protein prepared under native conditions binds very efficiently to cognate DNA, utilizing both N- and C-terminal residues. As a precursor to biophysical analyses of these complexes, a fluorescence-based random mutagenesis protocol was developed that enabled a structure-function analysis of the protein with good resolution at the secondary structure level. The results extend previous structure-function work on IS3 family transposases, identifying the binding domain as a three helix H + HTH bundle and explaining the function of an atypical leucine zipper-like motif in IS2. In addition gain- and loss-of-function mutations in the catalytic active site define its role in regional and global binding and identify functional signatures that are common to the three dimensional catalytic core motif of the retroviral integrase superfamily. Conclusions Intractably insoluble transposases, such as the IS2 transposase, prepared by solubilization protocols are often refractory to whole protein structure-function studies. The results described here have validated the use of GFP-tagging and fluorescence-based random mutagenesis in overcoming this limitation at the secondary structure level. PMID:22032517

Between timelessness and historiality: on the dynamics of the epistemic objects of mathematics.

PubMed

Epple, Moritz

2011-09-01

In order to discuss the temporal structure of mathematical research, this essay offers four related definitions of a mathematical object from different times and places. It is argued that in order to appreciate the differences between these definitions, the historian needs to understand that none of them made sense in mathematical practice without a technical framework, referred to but not explained in the definitions themselves (an "epistemic configuration of research"); that the dynamics of the epistemic objects of mathematical research are secondary to the dynamics of these epistemic configurations as a whole; and that the dynamics of epistemic configurations of mathematical research do not follow law-like processes. Very different types of change may happen, and some of them link the dynamics of epistemic configurations with events and developments far beyond the bounds of the research field in question. These insights have historiographical consequences that require us to rethink the kind of temporality ascribed to mathematics.

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

Tian, Shiliang; Liu, Jing; Cowley, Ryan E.

Here, S-Nitrosothiols are known as reagents for NO storage and transportation and as regulators in many physiological processes. Although the S-nitrosylation catalysed by haem proteins is well known, no direct evidence of S-nitrosylation in copper proteins has been reported. Here, we report reversible insertion of NO into a copper–thiolate bond in an engineered copper centre in Pseudomonas aeruginosa azurin by rational design of the primary coordination sphere and tuning its reduction potential by deleting a hydrogen bond in the secondary coordination sphere. The results not only provide the first direct evidence of S-nitrosylation of Cu(II)-bound cysteine in metalloproteins, but alsomore » shed light on the reaction mechanism and structural features responsible for stabilizing the elusive Cu(I)–S(Cys)NO species. The fast, efficient and reversible S-nitrosylation reaction is used to demonstrate its ability to prevent NO inhibition of cytochrome bo 3 oxidase activity by competing for NO binding with the native enzyme under physiologically relevant conditions.« less

Membrane Destruction and DNA Binding of Staphylococcus aureus Cells Induced by Carvacrol and Its Combined Effect with a Pulsed Electric Field.

PubMed

Wang, Lang-Hong; Wang, Man-Sheng; Zeng, Xin-An; Zhang, Zhi-Hong; Gong, De-Ming; Huang, Yan-Bo

2016-08-17

Carvacrol (5-isopropyl-2-methylphenol, CAR) is an antibacterial ingredient that occurs naturally in the leaves of the plant Origanum vulgare. The antimicrobial mechanism of CAR against Staphylococcus aureus ATCC 43300 was investigated in the study. Analysis of the membrane fatty acids by gas chromatography-mass spectrometry (GC-MS) showed that exposure to CAR at low concentrations induced a marked increase in the level of unbranched fatty acids (from 34.90 ± 1.77% to 62.37 ± 4.26%). Moreover, CAR at higher levels severely damaged the integrity and morphologies of the S. aureus cell membrane. The DNA-binding properties of CAR were also investigated using fluorescence, circular dichroism, molecular modeling, and atomic-force microscopy. The results showed that CAR bound to DNA via the minor-groove mode, mildly perturbed the DNA secondary structure, and induced DNA molecules to be aggregated. Furthermore, a combination of CAR with a pulsed-electric field was found to exhibit strong synergistic effects on S. aureus.

New potentially active pyrazinamide derivatives synthesized under microwave conditions.

PubMed

Jandourek, Ondrej; Dolezal, Martin; Kunes, Jiri; Kubicek, Vladimir; Paterova, Pavla; Pesko, Matus; Buchta, Vladimir; Kralova, Katarina; Zitko, Jan

2014-07-03

A series of 18 N-alkyl substituted 3-aminopyrazine-2-carboxamides was prepared in this work according to previously experimentally set and proven conditions using microwave assisted synthesis methodology. This approach for the aminodehalogenation reaction was chosen due to higher yields and shorter reaction times compared to organic reactions with conventional heating. Antimycobacterial, antibacterial, antifungal and photosynthetic electron transport (PET) inhibiting in vitro activities of these compounds were investigated. Experiments for the determination of lipophilicity were also performed. Only a small number of substances with alicyclic side chain showed activity against fungi which was the same or higher than standards and the biological efficacy of the compounds increased with rising lipophilicity. Nine pyrazinamide derivatives also inhibited PET in spinach chloroplasts and the IC50 values of these compounds varied in the range from 14.3 to 1590.0 μmol/L. The inhibitory activity was connected not only with the lipophilicity, but also with the presence of secondary amine fragment bounded to the pyrazine ring. Structure-activity relationships are discussed as well.

Designer lipid-like peptides: a class of detergents for studying functional olfactory receptors using commercial cell-free systems.

PubMed

Corin, Karolina; Baaske, Philipp; Ravel, Deepali B; Song, Junyao; Brown, Emily; Wang, Xiaoqiang; Wienken, Christoph J; Jerabek-Willemsen, Moran; Duhr, Stefan; Luo, Yuan; Braun, Dieter; Zhang, Shuguang

2011-01-01

A crucial bottleneck in membrane protein studies, particularly G-protein coupled receptors, is the notorious difficulty of finding an optimal detergent that can solubilize them and maintain their stability and function. Here we report rapid production of 12 unique mammalian olfactory receptors using short designer lipid-like peptides as detergents. The peptides were able to solubilize and stabilize each receptor. Circular dichroism showed that the purified olfactory receptors had alpha-helical secondary structures. Microscale thermophoresis suggested that the receptors were functional and bound their odorants. Blot intensity measurements indicated that milligram quantities of each olfactory receptor could be produced with at least one peptide detergent. The peptide detergents' capability was comparable to that of the detergent Brij-35. The ability of 10 peptide detergents to functionally solubilize 12 olfactory receptors demonstrates their usefulness as a new class of detergents for olfactory receptors, and possibly other G-protein coupled receptors and membrane proteins.

Designer Lipid-Like Peptides: A Class of Detergents for Studying Functional Olfactory Receptors Using Commercial Cell-Free Systems

PubMed Central

Corin, Karolina; Baaske, Philipp; Ravel, Deepali B.; Song, Junyao; Brown, Emily; Wang, Xiaoqiang; Wienken, Christoph J.; Jerabek-Willemsen, Moran; Duhr, Stefan; Luo, Yuan; Braun, Dieter; Zhang, Shuguang

2011-01-01

A crucial bottleneck in membrane protein studies, particularly G-protein coupled receptors, is the notorious difficulty of finding an optimal detergent that can solubilize them and maintain their stability and function. Here we report rapid production of 12 unique mammalian olfactory receptors using short designer lipid-like peptides as detergents. The peptides were able to solubilize and stabilize each receptor. Circular dichroism showed that the purified olfactory receptors had alpha-helical secondary structures. Microscale thermophoresis suggested that the receptors were functional and bound their odorants. Blot intensity measurements indicated that milligram quantities of each olfactory receptor could be produced with at least one peptide detergent. The peptide detergents' capability was comparable to that of the detergent Brij-35. The ability of 10 peptide detergents to functionally solubilize 12 olfactory receptors demonstrates their usefulness as a new class of detergents for olfactory receptors, and possibly other G-protein coupled receptors and membrane proteins. PMID:22132066

Interaction between Vaccinium bracteatum Thunb. leaf pigment and rice proteins.

PubMed

Wang, Li; Xu, Yuan; Zhou, Sumei; Qian, Haifeng; Zhang, Hui; Qi, Xiguang; Fan, Meihua

2016-03-01

In this study, we investigated the interaction of Vaccinium bracteatum Thunb. leaf (VBTL) pigment and rice proteins. In the presence of rice protein, VBTL pigment antioxidant activity and free polyphenol content decreased by 67.19% and 68.11%, respectively, and L(∗) of the protein-pigment complex decreased significantly over time. L(∗) values of albumin, globulin and glutelin during 60-min pigment exposure decreased by 55.00, 57.14, and 54.30%, respectively, indicating that these proteins had bound to the pigment. A significant difference in protein surface hydrophobicity was observed between rice proteins and pigment-protein complexes, indicating that hydrophobic interaction is a major binding mechanism between VBTL pigment and rice proteins. A significant difference in secondary structures between proteins and protein-pigment complexes was also uncovered, indicating that hydrogen bonding may be another mode of interaction between VBTL pigment and rice proteins. Our results indicate that VBTL pigment can stain rice proteins with hydrophobic and hydrogen interactions. Copyright © 2015 Elsevier Ltd. All rights reserved.

Interactions of cinnamaldehyde and its metabolite cinnamic acid with human serum albumin and interference of other food additives.

PubMed

Sun, Qiaomei; Yang, Hongqin; Tang, Peixiao; Liu, Jiuyang; Wang, Wan; Li, Hui

2018-03-15

Considering the adverse effect of food additives on humans, thorough research of their physiological effects at the molecular level is important. The interactions of cinnamaldehyde (CNMA), a food perfume, and its major metabolite cinnamic acid (CA) with human serum albumin (HSA) were examined by multiple-spectroscopies. NMR analysis revealed CNMA and CA both bound to HSA, and STD-NMR experiments established CNMA and CA primarily interacted with site I and site II of HSA, respectively. The ligands caused strong quenching of HSA fluorescence through a static quenching mechanism, with hydrophobic and electrostatic interaction between CNMA/CA and HSA, respectively. UV-vis absorption and CD results showed ligands induced secondary structure changes of HSA. Binding configurations were proved by docking method. Furthermore, binding constants of CNMA/CA-HSA systems were influenced by the addition of four other food additives. These studies have increased our knowledge regarding the safety and biological action of CNMA and CA. Copyright © 2017 Elsevier Ltd. All rights reserved.

Reversible S-nitrosylation in an engineered azurin

DOE PAGES

Tian, Shiliang; Liu, Jing; Cowley, Ryan E.; ...

2016-04-25

Here, S-Nitrosothiols are known as reagents for NO storage and transportation and as regulators in many physiological processes. Although the S-nitrosylation catalysed by haem proteins is well known, no direct evidence of S-nitrosylation in copper proteins has been reported. Here, we report reversible insertion of NO into a copper–thiolate bond in an engineered copper centre in Pseudomonas aeruginosa azurin by rational design of the primary coordination sphere and tuning its reduction potential by deleting a hydrogen bond in the secondary coordination sphere. The results not only provide the first direct evidence of S-nitrosylation of Cu(II)-bound cysteine in metalloproteins, but alsomore » shed light on the reaction mechanism and structural features responsible for stabilizing the elusive Cu(I)–S(Cys)NO species. The fast, efficient and reversible S-nitrosylation reaction is used to demonstrate its ability to prevent NO inhibition of cytochrome bo 3 oxidase activity by competing for NO binding with the native enzyme under physiologically relevant conditions.« less

Interaction between Pin1 and its natural product inhibitor epigallocatechin-3-gallate by spectroscopy and molecular dynamics simulations

NASA Astrophysics Data System (ADS)

Xi, Lei; Wang, Yu; He, Qing; Zhang, Qingyan; Du, Linfang

2016-12-01

The binding of epigallocatechin-3-gallate (EGCG) to wild type Pin1 in solution was studied by spectroscopic methods and molecular dynamics simulations in this research to explore the binding mode and inhibition mechanism. The binding constants and number of binding sites per Pin1 for EGCG were calculated through the Stern-Volmer equation. The values of binding free energy and thermodynamic parameters were calculated and indicated that hydrogen bonds, electrostatic interaction and Van der Waals interaction played the major role in the binding process. The alterations of Pin1 secondary structure in the presence of EGCG were confirmed by far-UV circular dichroism spectra. The binding model at atomic-level revealed that EGCG was bound to the Glu12, Lys13, Arg14, Met15 and Arg17 in WW domain. Furthermore, EGCG could also interact with Arg69, Asp112, Cys113 and Ser114 in PPIase domain.

An analytical study of nitrogen oxides and carbon monoxide emissions in hydrocarbon combustion with added nitrogen, preliminary results

NASA Technical Reports Server (NTRS)

Bittker, D. A.

1979-01-01

The effect of combustor operating conditions on the conversion of fuel-bound nitrogen (FBN) to nitrogen oxides NO sub x was analytically determined. The effect of FBN and of operating conditions on carbon monoxide (CO) formation was also studied. For these computations, the combustor was assumed to be a two stage, adiabatic, perfectly-stirred reactor. Propane-air was used as the combustible mixture and fuel-bound nitrogen was simulated by adding nitrogen atoms to the mixture. The oxidation of propane and formation of NO sub x and CO were modeled by a fifty-seven reaction chemical mechanism. The results for NO sub x and CO formation are given as functions of primary and secondary stage equivalence ratios and residence times.

Mineralogical basis for the interpretation of multi-element (ICP-AES), oxalic acid, and aqua regia partial digestions of stream sediments for reconnaissance exploration geochemistry

USGS Publications Warehouse

Church, S.E.; Mosier, E.L.; Motooka, J.M.

1987-01-01

We have applied partial digestion procedures, primarily oxalic acid and aqua regia leaches, to several regional geochemical reconnaissance studies carried out using Inductively Coupled Plasma-Atomic Emission Spectroscopy (ICP-AES) analytical methods. We have chosen to use these two acids because the oxalic acid primarily attacks those compounds formed during secondary geochemical processes, whereas aqua regia will digest the primary sulfide phases as well as secondary phases. Application of the partial digestion technique has proven superior to total digestion because the concentration of metals in hydromorphic compounds and the sulfides is enhanced relative to the metals bound in the unattacked silicate phases. The aqua regia digestion attacks and leaches metals from the mafic chain silicates and the phyllosilicates (coordination number of VI or more), yielding a characteristic geochemical signature, but does not leach appreciable metal from many other silicates. In order to interpret the results from these leach studies, we have initiated an investigation of a large suite of hand-picked mineral separates. The study includes analyses of about two hundred minerals representing the common rock-forming minerals as well as end-member compositions of various silicates, oxides, sulfides, carbonates, sulfates, and some vanadates, molybdates, tungstates, and phosphates. The objective of this study is to evaluate the effect of leaching by acids of particular lattice sites in specific mineral structures. ?? 1987.

Efficient organ localization using multi-label convolutional neural networks in thorax-abdomen CT scans

NASA Astrophysics Data System (ADS)

Efrain Humpire-Mamani, Gabriel; Arindra Adiyoso Setio, Arnaud; van Ginneken, Bram; Jacobs, Colin

2018-04-01

Automatic localization of organs and other structures in medical images is an important preprocessing step that can improve and speed up other algorithms such as organ segmentation, lesion detection, and registration. This work presents an efficient method for simultaneous localization of multiple structures in 3D thorax-abdomen CT scans. Our approach predicts the location of multiple structures using a single multi-label convolutional neural network for each orthogonal view. Each network takes extra slices around the current slice as input to provide extra context. A sigmoid layer is used to perform multi-label classification. The output of the three networks is subsequently combined to compute a 3D bounding box for each structure. We used our approach to locate 11 structures of interest. The neural network was trained and evaluated on a large set of 1884 thorax-abdomen CT scans from patients undergoing oncological workup. Reference bounding boxes were annotated by human observers. The performance of our method was evaluated by computing the wall distance to the reference bounding boxes. The bounding boxes annotated by the first human observer were used as the reference standard for the test set. Using the best configuration, we obtained an average wall distance of 3.20~+/-~7.33 mm in the test set. The second human observer achieved 1.23~+/-~3.39 mm. For all structures, the results were better than those reported in previously published studies. In conclusion, we proposed an efficient method for the accurate localization of multiple organs. Our method uses multiple slices as input to provide more context around the slice under analysis, and we have shown that this improves performance. This method can easily be adapted to handle more organs.

Structural basis for the ligand-binding specificity of fatty acid-binding proteins (pFABP4 and pFABP5) in gentoo penguin.

PubMed

Lee, Chang Woo; Kim, Jung Eun; Do, Hackwon; Kim, Ryeo-Ok; Lee, Sung Gu; Park, Hyun Ho; Chang, Jeong Ho; Yim, Joung Han; Park, Hyun; Kim, Il-Chan; Lee, Jun Hyuck

2015-09-11

Fatty acid-binding proteins (FABPs) are involved in transporting hydrophobic fatty acids between various aqueous compartments of the cell by directly binding ligands inside their β-barrel cavities. Here, we report the crystal structures of ligand-unbound pFABP4, linoleate-bound pFABP4, and palmitate-bound pFABP5, obtained from gentoo penguin (Pygoscelis papua), at a resolution of 2.1 Å, 2.2 Å, and 2.3 Å, respectively. The pFABP4 and pFABP5 proteins have a canonical β-barrel structure with two short α-helices that form a cap region and fatty acid ligand binding sites in the hydrophobic cavity within the β-barrel structure. Linoleate-bound pFABP4 and palmitate-bound pFABP5 possess different ligand-binding modes and a unique ligand-binding pocket due to several sequence dissimilarities (A76/L78, T30/M32, underlining indicates pFABP4 residues) between the two proteins. Structural comparison revealed significantly different conformational changes in the β3-β4 loop region (residues 57-62) as well as the flipped Phe60 residue of pFABP5 than that in pFABP4 (the corresponding residue is Phe58). A ligand-binding study using fluorophore displacement assays shows that pFABP4 has a relatively strong affinity for linoleate as compared to pFABP5. In contrast, pFABP5 exhibits higher affinity for palmitate than that for pFABP4. In conclusion, our high-resolution structures and ligand-binding studies provide useful insights into the ligand-binding preferences of pFABPs based on key protein-ligand interactions. Copyright © 2015 Elsevier Inc. All rights reserved.

Local Conformational Stability of HIV-1 gp120 in Unliganded and CD4-Bound States as Defined by Amide Hydrogen/Deuterium Exchange▿ †

PubMed Central

Kong, Leopold; Huang, Chih-chin; Coales, Stephen J.; Molnar, Kathleen S.; Skinner, Jeff; Hamuro, Yoshitomo; Kwong, Peter D.

2010-01-01

The binding reaction of the HIV-1 gp120 envelope glycoprotein to the CD4 receptor involves exceptional changes in enthalpy and entropy. Crystal structures of gp120 in unliganded and various ligand-bound states, meanwhile, reveal an inner domain able to fold into diverse conformations, a structurally invariant outer domain, and, in the CD4-bound state, a bridging sheet minidomain. These studies, however, provide only hints as to the flexibility of each state. Here we use amide hydrogen/deuterium exchange coupled to mass spectrometry to provide quantifications of local conformational stability for HIV-1 gp120 in unliganded and CD4-bound states. On average, unliganded core gp120 displayed >10,000-fold slower exchange of backbone-amide hydrogens than a theoretically unstructured protein of the same composition, with binding by CD4 reducing the rate of gp120 amide exchange a further 10-fold. For the structurally constant CD4, alterations in exchange correlated well with alterations in binding surface (P value = 0.0004). For the structurally variable gp120, however, reductions in flexibility extended outside the binding surface, and regions of expected high structural diversity (inner domain/bridging sheet) displayed roughly 20-fold more rapid exchange in the unliganded state than regions of low diversity (outer domain). Thus, despite an extraordinary reduction in entropy, neither unliganded gp120 nor free CD4 was substantially unstructured, suggesting that most of the diverse conformations that make up the gp120 unliganded state are reasonably ordered. The results provide a framework for understanding how local conformational stability influences entropic change, conformational diversity, and structural rearrangements in the gp120-CD4 binding reaction. PMID:20660185

Monomerization alters the dynamics of the lid region in Campylobacter jejuni CstII: an MD simulation study.

PubMed

Prabhakar, Pradeep Kumar; Srivastava, Alka; Rao, K Krishnamurthy; Balaji, Petety V

2016-01-01

CstII, a bifunctional (α2,3/8) sialyltransferase from Campylobacter jejuni, is a homotetramer. It has been reported that mutation of the interface residues Phe121 (F121D) or Tyr125 (Y125Q) leads to monomerization and partial loss of enzyme activity, without any change in the secondary or tertiary structures. MD simulations of both tetramer and monomer, with and without bound donor substrate, were performed for the two mutants and WT to understand the reasons for partial loss of activity due to monomerization since the active site is located within each monomer. RMSF values were found to correlate with the crystallographic B-factor values indicating that the simulations are able to capture the flexibility of the molecule effectively. There were no gross changes in either the secondary or tertiary structure of the proteins during MD simulations. However, interface is destabilized by the mutations, and more importantly the flexibility of the lid region (Gly152-Lys190) is affected. The lid region accesses three major conformations named as open, intermediate, and closed conformations. In both Y121Q and F121D mutants, the closed conformation is accessed predominantly. In this conformation, the catalytic base His188 is also displaced. Normal mode analysis also revealed differences in the lid movement in tetramer and monomer. This provides a possible explanation for the partial loss of enzyme activity in both interface mutants. The lid region controls the traffic of substrates and products in and out of the active site, and the dynamics of this region is regulated by tetramerization. Thus, this study provides valuable insights into the role of loop dynamics in enzyme activity of CstII.

MiR-146a polymorphism correlates with lung cancer risk in Chinese nonsmoking females

PubMed Central

Yin, Zhihua; Cui, Zhigang; Ren, Yangwu; Xia, Lingzi; Li, Hang; Zhou, Baosen

2017-01-01

This study provides evidence that the common rs2910164 polymorphism in miR-146a strongly correlates with lung cancer risk in nonsmoking females in northeast China. The genotypes of miR-146a rs2910164 were determined in 1131 patients with lung cancer and 1003 healthy control subjects. Tissue samples were used to evaluate the association between miRNA expression and lung cancer risk as well as the correlation between rs2910164 genotypes and miR-146a expression. The secondary structures of the wild-type and variant miR-146a sequences were predicted, and luciferase-based target assays were used to test whether miR-146a bound to tumor necrosis factor receptor associated factor 6 (TRAF6) mRNA. Individuals carrying heterozygous CG genotype of miR-146a rs2910164 had less risk of lung cancer than those carrying homozygous wild CC genotype (OR = 0.76, 95% CI = 0.60-0.98, P = 0.032). We found no significant association between miR-146a expression and lung cancer risk. MiR-146a expression differed in those carrying the CC genotype as compared with the CG or the GG genotype (P = 0.032 and 0.001), and the secondary structure of the C allele differed slightly from the G allele. Significantly lower levels of luciferase activity were observed when the TRAF6 3′UTR was cotransfected with miR-146a-3p carrying the rs2910164 C allele (P = 0.001). Thus, miR-146a rs2910164 polymorphism may influence susceptibility to lung cancer in Chinese nonsmoking females through targeting TRAF6. PMID:27911870

Properties of Protein Drug Target Classes

PubMed Central

Bull, Simon C.; Doig, Andrew J.

2015-01-01

Accurate identification of drug targets is a crucial part of any drug development program. We mined the human proteome to discover properties of proteins that may be important in determining their suitability for pharmaceutical modulation. Data was gathered concerning each protein’s sequence, post-translational modifications, secondary structure, germline variants, expression profile and drug target status. The data was then analysed to determine features for which the target and non-target proteins had significantly different values. This analysis was repeated for subsets of the proteome consisting of all G-protein coupled receptors, ion channels, kinases and proteases, as well as proteins that are implicated in cancer. Machine learning was used to quantify the proteins in each dataset in terms of their potential to serve as a drug target. This was accomplished by first inducing a random forest that could distinguish between its targets and non-targets, and then using the random forest to quantify the drug target likeness of the non-targets. The properties that can best differentiate targets from non-targets were primarily those that are directly related to a protein’s sequence (e.g. secondary structure). Germline variants, expression levels and interactions between proteins had minimal discriminative power. Overall, the best indicators of drug target likeness were found to be the proteins’ hydrophobicities, in vivo half-lives, propensity for being membrane bound and the fraction of non-polar amino acids in their sequences. In terms of predicting potential targets, datasets of proteases, ion channels and cancer proteins were able to induce random forests that were highly capable of distinguishing between targets and non-targets. The non-target proteins predicted to be targets by these random forests comprise the set of the most suitable potential future drug targets, and should therefore be prioritised when building a drug development programme. PMID:25822509

The Role of Histone Tails in the Nucleosome: A Computational Study

PubMed Central

Erler, Jochen; Zhang, Ruihan; Petridis, Loukas; Cheng, Xiaolin; Smith, Jeremy C.; Langowski, Jörg

2014-01-01

Histone tails play an important role in gene transcription and expression. We present here a systematic computational study of the role of histone tails in the nucleosome, using replica exchange molecular dynamics simulations with an implicit solvent model and different well-established force fields. We performed simulations for all four histone tails, H4, H3, H2A, and H2B, isolated and with inclusion of the nucleosome. The results confirm predictions of previous theoretical studies for the secondary structure of the isolated tails but show a strong dependence on the force field used. In the presence of the entire nucleosome for all force fields, the secondary structure of the histone tails is destabilized. Specific contacts are found between charged lysine and arginine residues and DNA phosphate groups and other binding sites in the minor and major DNA grooves. Using cluster analysis, we found a single dominant configuration of binding to DNA for the H4 and H2A histone tails, whereas H3 and H2B show multiple binding configurations with an equal probability. The leading stabilizing contribution for those binding configurations is the attractive interaction between the positively charged lysine and arginine residues and the negatively charged phosphate groups, and thus the resulting charge neutralization. Finally, we present results of molecular dynamics simulations in explicit solvent to confirm our conclusions. Results from both implicit and explicit solvent models show that large portions of the histone tails are not bound to DNA, supporting the complex role of these tails in gene transcription and expression and making them possible candidates for binding sites of transcription factors, enzymes, and other proteins. PMID:25517156

Dual Role of OhrR as a Repressor and an Activator in Response to Organic Hydroperoxides in Streptomyces coelicolor▿

PubMed Central

Oh, So-Young; Shin, Jung-Ho; Roe, Jung-Hye

2007-01-01

Organic hydroperoxide resistance in bacteria is achieved primarily through reducing oxidized membrane lipids. The soil-inhabiting aerobic bacterium Streptomyces coelicolor contains three paralogous genes for organic hydroperoxide resistance: ohrA, ohrB, and ohrC. The ohrA gene is transcribed divergently from ohrR, which encodes a putative regulator of MarR family. Both the ohrA and ohrR genes were induced highly by various organic hydroperoxides. The ohrA gene was induced through removal of repression by OhrR, whereas the ohrR gene was induced through activation by OhrR. Reduced OhrR bound to the ohrA-ohrR intergenic region, which contains a central (primary) and two adjacent (secondary) inverted-repeat motifs that overlap with promoter elements. Organic peroxide decreased the binding affinity of OhrR for the primary site, with a concomitant decrease in cooperative binding to the adjacent secondary sites. The single cysteine C28 in OhrR was involved in sensing oxidants, as determined by substitution mutagenesis. The C28S mutant of OhrR bound to the intergenic region without any change in binding affinity in response to organic peroxides. These results lead us to propose a model for the dual action of OhrR as a repressor and an activator in S. coelicolor. Under reduced conditions, OhrR binds cooperatively to the intergenic region, repressing transcription from both genes. Upon oxidation, the binding affinity of OhrR decreases, with a concomitant loss of cooperative binding, which allows RNA polymerase to bind to both the ohrA and ohrR promoters. The loosely bound oxidized OhrR can further activate transcription from the ohrR promoter. PMID:17586628

A generalized analysis of hydrophobic and loop clusters within globular protein sequences

PubMed Central

Eudes, Richard; Le Tuan, Khanh; Delettré, Jean; Mornon, Jean-Paul; Callebaut, Isabelle

2007-01-01

Background Hydrophobic Cluster Analysis (HCA) is an efficient way to compare highly divergent sequences through the implicit secondary structure information directly derived from hydrophobic clusters. However, its efficiency and application are currently limited by the need of user expertise. In order to help the analysis of HCA plots, we report here the structural preferences of hydrophobic cluster species, which are frequently encountered in globular domains of proteins. These species are characterized only by their hydrophobic/non-hydrophobic dichotomy. This analysis has been extended to loop-forming clusters, using an appropriate loop alphabet. Results The structural behavior of hydrophobic cluster species, which are typical of protein globular domains, was investigated within banks of experimental structures, considered at different levels of sequence redundancy. The 294 more frequent hydrophobic cluster species were analyzed with regard to their association with the different secondary structures (frequencies of association with secondary structures and secondary structure propensities). Hydrophobic cluster species are predominantly associated with regular secondary structures, and a large part (60 %) reveals preferences for α-helices or β-strands. Moreover, the analysis of the hydrophobic cluster amino acid composition generally allows for finer prediction of the regular secondary structure associated with the considered cluster within a cluster species. We also investigated the behavior of loop forming clusters, using a "PGDNS" alphabet. These loop clusters do not overlap with hydrophobic clusters and are highly associated with coils. Finally, the structural information contained in the hydrophobic structural words, as deduced from experimental structures, was compared to the PSI-PRED predictions, revealing that β-strands and especially α-helices are generally over-predicted within the limits of typical β and α hydrophobic clusters. Conclusion The dictionary of hydrophobic clusters described here can help the HCA user to interpret and compare the HCA plots of globular protein sequences, as well as provides an original fundamental insight into the structural bricks of protein folds. Moreover, the novel loop cluster analysis brings additional information for secondary structure prediction on the whole sequence through a generalized cluster analysis (GCA), and not only on regular secondary structures. Such information lays the foundations for developing a new and original tool for secondary structure prediction. PMID:17210072

A novel type of light-harvesting antenna protein of red algal origin in algae with secondary plastids.

PubMed

Sturm, Sabine; Engelken, Johannes; Gruber, Ansgar; Vugrinec, Sascha; Kroth, Peter G; Adamska, Iwona; Lavaud, Johann

2013-07-30

Light, the driving force of photosynthesis, can be harmful when present in excess; therefore, any light harvesting system requires photoprotection. Members of the extended light-harvesting complex (LHC) protein superfamily are involved in light harvesting as well as in photoprotection and are found in the red and green plant lineages, with a complex distribution pattern of subfamilies in the different algal lineages. Here, we demonstrate that the recently discovered "red lineage chlorophyll a/b-binding-like proteins" (RedCAPs) form a monophyletic family within this protein superfamily. The occurrence of RedCAPs was found to be restricted to the red algal lineage, including red algae (with primary plastids) as well as cryptophytes, haptophytes and heterokontophytes (with secondary plastids of red algal origin). Expression of a full-length RedCAP:GFP fusion construct in the diatom Phaeodactylum tricornutum confirmed the predicted plastid localisation of RedCAPs. Furthermore, we observed that similarly to the fucoxanthin chlorophyll a/c-binding light-harvesting antenna proteins also RedCAP transcripts in diatoms were regulated in a diurnal way at standard light conditions and strongly repressed at high light intensities. The absence of RedCAPs from the green lineage implies that RedCAPs evolved in the red lineage after separation from the the green lineage. During the evolution of secondary plastids, RedCAP genes therefore must have been transferred from the nucleus of the endocytobiotic alga to the nucleus of the host cell, a process that involved complementation with pre-sequences allowing import of the gene product into the secondary plastid bound by four membranes. Based on light-dependent transcription and on localisation data, we propose that RedCAPs might participate in the light (intensity and quality)-dependent structural or functional reorganisation of the light-harvesting antennae of the photosystems upon dark to light shifts as regularly experienced by diatoms in nature. Remarkably, in plastids of the red lineage as well as in green lineage plastids, the phycobilisome based cyanobacterial light harvesting system has been replaced by light harvesting systems that are based on members of the extended LHC protein superfamily, either for one of the photosystems (PS I of red algae) or for both (diatoms). In their proposed function, the RedCAP protein family may thus have played a role in the evolutionary structural remodelling of light-harvesting antennae in the red lineage.

Binding of Dihydrostreptomycin to Escherichia coli Ribosomes: Characteristics and Equilibrium of the Reaction

PubMed Central

Chang, F. N.; Flaks, Joel G.

1972-01-01

The binding of dihydrostreptomycin to ribosomes and ribosomal subunits of a number of different Escherichia coli strains was studied, and the Mg2+ and pH dependence, as well as the effect of salts and polynucleotides, was determined. The only requirement for binding with ribosomes and subunits from susceptible strains was 10 mm Mg2+. Monovalent salts weakened the binding in a manner similar to the effects on ribonucleic acid secondary structure, and this was antagonized to some extent by increased amounts of Mg2+. Bound dihydrostreptomycin could be readily exchanged by streptomycin and any antibiotically active derivative, but not by fragments of the antibiotic or any other aminoglycoside. With native (run-off) 70S ribosomes from streptomycin-susceptible strains, the binding was rapid and relatively temperature independent over the range from 0 to 37 C. Polynucleotides did not stimulate the binding. With concentrations of dihydrostreptomycin up to 10−5m, greater than 95% of native 70S ribosomes bound exactly 1 molecule of the antibiotic tightly, with a Kdiss for the bound complex at 25 C of 9.4 × 10−8m. The following thermodynamic parameters were found for the binding with 70S ribosomes at 25 C:ΔG° = −9.6 kcal/mole, ΔH° = −6.2 kcal/mole, and ΔS° = +11.4 entropy units/mole. Differences in affinity for the antibiotic were found between ribosomes of K-12 strains and those of other E. coli strains. There was insignificant binding to 70S ribosomes or subunits from streptomycin-resistant or -dependent strains, and to 50S subunits from susceptible strains. The binding to 30S subunits from susceptible strains was weaker by an order of magnitude than that to the 70S particle, with a Kdiss at 25 C of 10−6m. Polyuridylic acid stimulated this binding slightly but did not influence the affinity of the bound molecule. At antibiotic concentrations above 10−5m, streptomycin-susceptible 70S and 30S particles bound additional molecules of the antibiotic, and binding also occurred to ribosomes from streptomycin-resistant and -dependent strains, as well as to 50S subunits from all strains. Kdiss for all of these binding equilibria were [Formula: see text] 10−4m. This weaker non-specific binding coincided with the beginning of aggregation phenomena involving the particles, and occurred at sites distinct from the single site which binds the antibiotic tightly. This latter site was completely lost after the one-step mutation to high-level resistance or dependence. PMID:4133236

Two-dimensional sum-frequency generation (2D SFG) reveals structure and dynamics of a surface-bound peptide

PubMed Central

Laaser, Jennifer E.; Skoff, David R.; Ho, Jia-Jung; Joo, Yongho; Serrano, Arnaldo L.; Steinkruger, Jay D.; Gopalan, Padma; Gellman, Samuel H.; Zanni, Martin T.

2014-01-01

Surface-bound polypeptides and proteins are increasingly used to functionalize inorganic interfaces such as electrodes, but their structural characterization is exceedingly difficult with standard technologies. In this paper, we report the first two-dimensional sum-frequency generation (2D SFG) spectra of a peptide monolayer, which is collected by adding a mid-IR pulse shaper to a standard femtosecond SFG spectrometer. On a gold surface, standard FTIR spectroscopy is inconclusive about the peptide structure because of solvation-induced frequency shifts, but the 2D lineshapes, anharmonic shifts, and lifetimes obtained from 2D SFG reveal that the peptide is largely α-helical and upright. Random coil residues are also observed, which do not themselves appear in SFG spectra due to their isotropic structural distribution, but which still absorb infrared light and so can be detected by cross-peaks in 2D SFG spectra. We discuss these results in the context of peptide design. Because of the similar way in which the spectra are collected, these 2D SFG spectra can be directly compared to 2D IR spectra, thereby enabling structural interpretations of surface-bound peptides and biomolecules based on the well-studied structure/2D IR spectra relationships established from soluble proteins. PMID:24372101

A new crystal structure of the bifunctional antibiotic simocyclinone D8 bound to DNA gyrase gives fresh insight into the mechanism of inhibition.

PubMed

Hearnshaw, Stephen J; Edwards, Marcus J; Stevenson, Clare E; Lawson, David M; Maxwell, Anthony

2014-05-15

Simocyclinone D8 (SD8) is an antibiotic produced by Streptomyces antibioticus that targets DNA gyrase. A previous structure of SD8 complexed with the N-terminal domain of the DNA gyrase A protein (GyrA) suggested that four SD8 molecules stabilized a tetramer of the protein; subsequent mass spectrometry experiments suggested that a protein dimer with two symmetry-related SD8s was more likely. This work describes the structures of a further truncated form of the GyrA N-terminal domain fragment with and without SD8 bound. The structure with SD8 has the two SD8 molecules bound within the same GyrA dimer. This new structure is entirely consistent with the mutations in GyrA that confer SD8 resistance and, by comparison with a new apo structure of the GyrA N-terminal domain, reveals the likely conformation changes that occur upon SD8 binding and the detailed mechanism of SD8 inhibition of gyrase. Isothermal titration calorimetry experiments are consistent with the crystallography results and further suggest that a previously observed complex between SD8 and GyrB is ~1000-fold weaker than the interaction with GyrA. Copyright © 2014. Published by Elsevier Ltd.

A New Crystal Structure of the Bifunctional Antibiotic Simocyclinone D8 Bound to DNA Gyrase Gives Fresh Insight into the Mechanism of Inhibition

PubMed Central

Hearnshaw, Stephen J.; Edwards, Marcus J.; Stevenson, Clare E.; Lawson, David M.; Maxwell, Anthony

2014-01-01

Simocyclinone D8 (SD8) is an antibiotic produced by Streptomyces antibioticus that targets DNA gyrase. A previous structure of SD8 complexed with the N-terminal domain of the DNA gyrase A protein (GyrA) suggested that four SD8 molecules stabilized a tetramer of the protein; subsequent mass spectrometry experiments suggested that a protein dimer with two symmetry-related SD8s was more likely. This work describes the structures of a further truncated form of the GyrA N-terminal domain fragment with and without SD8 bound. The structure with SD8 has the two SD8 molecules bound within the same GyrA dimer. This new structure is entirely consistent with the mutations in GyrA that confer SD8 resistance and, by comparison with a new apo structure of the GyrA N-terminal domain, reveals the likely conformation changes that occur upon SD8 binding and the detailed mechanism of SD8 inhibition of gyrase. Isothermal titration calorimetry experiments are consistent with the crystallography results and further suggest that a previously observed complex between SD8 and GyrB is ~ 1000-fold weaker than the interaction with GyrA. PMID:24594357

Structural insight into the substrate- and dioxygen-binding manner in the catalytic cycle of rieske nonheme iron oxygenase system, carbazole 1,9a-dioxygenase.

PubMed

Ashikawa, Yuji; Fujimoto, Zui; Usami, Yusuke; Inoue, Kengo; Noguchi, Haruko; Yamane, Hisakazu; Nojiri, Hideaki

2012-06-24

Dihydroxylation of tandemly linked aromatic carbons in a cis-configuration, catalyzed by multicomponent oxygenase systems known as Rieske nonheme iron oxygenase systems (ROs), often constitute the initial step of aerobic degradation pathways for various aromatic compounds. Because such RO reactions inherently govern whether downstream degradation processes occur, novel oxygenation mechanisms involving oxygenase components of ROs (RO-Os) is of great interest. Despite substantial progress in structural and physicochemical analyses, no consensus exists on the chemical steps in the catalytic cycles of ROs. Thus, determining whether conformational changes at the active site of RO-O occur by substrate and/or oxygen binding is important. Carbazole 1,9a-dioxygenase (CARDO), a RO member consists of catalytic terminal oxygenase (CARDO-O), ferredoxin (CARDO-F), and ferredoxin reductase. We have succeeded in determining the crystal structures of oxidized CARDO-O, oxidized CARDO-F, and both oxidized and reduced forms of the CARDO-O: CARDO-F binary complex. In the present study, we determined the crystal structures of the reduced carbazole (CAR)-bound, dioxygen-bound, and both CAR- and dioxygen-bound CARDO-O: CARDO-F binary complex structures at 1.95, 1.85, and 2.00 Å resolution. These structures revealed the conformational changes that occur in the catalytic cycle. Structural comparison between complex structures in each step of the catalytic mechanism provides several implications, such as the order of substrate and dioxygen bindings, the iron-dioxygen species likely being Fe(III)-(hydro)peroxo, and the creation of room for dioxygen binding and the promotion of dioxygen binding in desirable fashion by preceding substrate binding. The RO catalytic mechanism is proposed as follows: When the Rieske cluster is reduced, substrate binding induces several conformational changes (e.g., movements of the nonheme iron and the ligand residue) that create room for oxygen binding. Dioxygen bound in a side-on fashion onto nonheme iron is activated by reduction to the peroxo state [Fe(III)-(hydro)peroxo]. This state may react directly with the bound substrate, or O-O bond cleavage may occur to generate Fe(V)-oxo-hydroxo species prior to the reaction. After producing a cis-dihydrodiol, the product is released by reducing the nonheme iron. This proposed scheme describes the catalytic cycle of ROs and provides important information for a better understanding of the mechanism.

Structural features and dynamic investigations of the membrane-bound cytochrome P450 17A1.

PubMed

Cui, Ying-Lu; Xue, Qiao; Zheng, Qing-Chuan; Zhang, Ji-Long; Kong, Chui-Peng; Fan, Jing-Rong; Zhang, Hong-Xing

2015-10-01

Cytochrome P450 (CYP) 17A1 is a dual-function monooxygenase with a critical role in the synthesis of many human steroid hormones. The enzyme is an important target for treatment of breast and prostate cancers that proliferate in response to estrogens and androgens. Despite the crystallographic structures available for CYP17A1, no membrane-bound structural features of this enzyme at atomic level are available. Accumulating evidence has indicated that the interactions between bounded CYPs and membrane could contribute to the recruitment of lipophilic substrates. To this end, we have investigated the effects on structural characteristics in the presence of the membrane for CYP17A1. The MD simulation results demonstrate a spontaneous insertion process of the enzyme to the lipid. Two predominant modes of CYP17A1 in the membrane are captured, characterized by the depths of insertion and orientations of the enzyme to the membrane surface. The measured heme tilt angles show good consistence with experimental data, thereby verifying the validity of the structural models. Moreover, conformational changes induced by the membrane might have impact on the accessibility of the active site to lipophilic substrates. The dynamics of internal aromatic gate formed by Trp220 and Phe224 are suggested to regulate tunnel opening motions. The knowledge of the membrane binding characteristics could guide future experimental and computational works on membrane-bound CYPs so that various investigations of CYPs in their natural, lipid environment rather than in artificially solubilized forms may be achieved. Copyright © 2015. Published by Elsevier B.V.

A Bulky Rhodium Complex Bound to an Adenosine-Adenosine DNA Mismatch: General Architecture of the Metalloinsertion Binding Mode†

PubMed Central

Zeglis, Brian M.; Pierre, Valérie C.; Kaiser, Jens T.; Barton, Jacqueline K.

2009-01-01

Two crystal structures are determined for Δ-Rh(bpy)2(chrysi)3+ (chrysi = 5,6-chrysenequinone diimine) bound to the oligonucleotide duplex 5′-CGGAAATTACCG-3′ containing two adenosine-adenosine mismatches (italics) through metalloinsertion. Diffraction quality crystals with two different space groups (P3221 and P43212) were obtained under very similar crystallization conditions. In both structures, the bulky rhodium complex inserts into the two mismatched sites from the minor groove side, ejecting the mismatched bases into the major groove. The conformational changes are localized to the mismatched site; the metal complex replaces the mismatched base pair without an increase in base pair rise. The expansive metal complex is accommodated in the duplex by a slight opening in the phosphodiester backbone; all sugars retain a C2′-endo puckering, and flanking base pairs neither stretch nor shear. The structures differ, however, in that in one of the structures, an additional metal complex is bound by intercalation from the major groove at the central 5′-AT-3′ step. We conclude that this additional metal complex is intercalated into this central step because of crystal packing forces. The structures described here of Δ-Rh(bpy)2(chrysi)3+ bound to thermodynamically destabilized AA mismatches share critical features with binding by metalloinsertion in two other oligonucleotides containing different single base mismatches. These results underscore the generality of the metalloinsertion as a new mode of non-covalent binding by small molecules with a DNA duplex. PMID:19374348

Structural Insights into Bound Water in Crystalline Amino Acids: Experimental and Theoretical (17)O NMR.

PubMed

Michaelis, Vladimir K; Keeler, Eric G; Ong, Ta-Chung; Craigen, Kimberley N; Penzel, Susanne; Wren, John E C; Kroeker, Scott; Griffin, Robert G

2015-06-25

We demonstrate here that the (17)O NMR properties of bound water in a series of amino acids and dipeptides can be determined with a combination of nonspinning and magic-angle spinning experiments using a range of magnetic field strengths from 9.4 to 21.1 T. Furthermore, we propose a (17)O chemical shift fingerprint region for bound water molecules in biological solids that is well outside the previously determined ranges for carbonyl, carboxylic, and hydroxyl oxygens, thereby offering the ability to resolve multiple (17)O environments using rapid one-dimensional NMR techniques. Finally, we compare our experimental data against quantum chemical calculations using GIPAW and hybrid-DFT, finding intriguing discrepancies between the electric field gradients calculated from structures determined by X-ray and neutron diffraction.

Evidence for weakly bound electrons in non-irradiated alkane crystals: The electrons as a probe of structural differences in crystals

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

Pietrow, M., E-mail: mrk@kft.umcs.lublin.pl; Misiak, L. E.; Gagoś, M.

2015-02-14

It is generally assumed that weakly bound (trapped) electrons in organic solids come only from radiolytical (or photochemical) processes like ionization caused by an excited positron entering the sample. This paper presents evidence for the presence of these electrons in non-irradiated samples of docosane. This can be due to the triboelectrification process. We argue that these electrons can be located (trapped) either in interlamellar gaps or in spaces made by non-planar conformers. Electrons from the former ones are bound more weakly than electrons from the latter ones. The origin of Vis absorption for the samples is explained. These spectra canmore » be used as a probe indicating differences in the solid structures of hydrocarbons.« less

A model for the generation of two-dimensional surf beat

USGS Publications Warehouse

List, Jeffrey H.

1992-01-01

A finite difference model predicting group-forced long waves in the nearshore is constructed with two interacting parts: an incident wave model providing time-varying radiation stress gradients across the nearshore, and a long-wave model which solves the equations of motion for the forcing imposed by the incident waves. Both shallow water group-bound long waves and long waves generated by a time-varying breakpoint are simulated. Model-generated time series are used to calculate the cross correlation between wave groups and long waves through the surf zone. The cross-correlation signal first observed by Tucker (1950) is well predicted. For the first time, this signal is decomposed into the contributions from the two mechanisms of leaky mode forcing. Results show that the cross-correlation signal can be explained by bound long waves which are amplified, though strongly modified, through the surf zone before reflection from the shoreline. The breakpoint-forced long waves are added to the bound long waves at a phase of pi/2 and are a secondary contribution owing to their relatively small size.

Evidence for a positron bound state on the surface of a topological insulator

NASA Astrophysics Data System (ADS)

Shastry, K.; Weiss, A. H.; Barbiellini, B.; Assaf, B. A.; Lim, Z. H.; Joglekar, P. V.; Heiman, D.

2015-06-01

We describe experiments aimed at probing the sticking of positrons to the surfaces of topological insulators using the Positron Annihilation induced Auger Electron Spectrometer (PAES). A magnetically guided beam was used to deposit positrons at the surface of Bi2Te2Se sample at energy of ∼2eV. Peaks observed in the energy spectra and intensities of electrons emitted as a result of positron annihilation showed peaks at energies corresponding to Auger peaks in Bi, Teand Se providing clear evidence of Auger emission associated with the annihilation of positrons in a surface bound state. Theoretical estimates of the binding energy of this state are compared with estimates obtained by measuring the incident beam energy threshold for secondary electron emission and the temperature dependence positronium(Ps) emission. The experiments provide strong evidence for the existence of a positron bound state at the surface of Bi2Te2Se and indicate the practicality of using positron annihilation to selectively probe the critically important top most layer of topological insulator system.

A membrane lipid imbalance plays a role in the phenotypic expression of cystic fibrosis in cftr−/− mice

PubMed Central

Freedman, Steven D.; Katz, Mark H.; Parker, Eliza M.; Laposata, Michael; Urman, Mark Y.; Alvarez, Juan G.

1999-01-01

A deficiency in essential fatty acid metabolism has been reported in plasma from patients with cystic fibrosis (CF). However, its etiology and role in the expression of disease is unknown. The objective of this study was to determine whether alterations in fatty acid metabolism are specific to CF-regulated organs and whether they play a role in the expression of disease. A membrane lipid imbalance was found in ileum, pancreas, and lung from cftr−/− mice characterized by an increase in phospholipid-bound arachidonic acid and a decrease in phospholipid-bound docosahexaenoic acid (DHA). This lipid imbalance was observed in organs pathologically affected by CF including lung, pancreas, and ileum and was not secondary to impaired intestinal absorption or hepatic biosynthesis of DHA. As proof of concept, oral administration of DHA to cftr−/− mice corrected this lipid imbalance and reversed the observed pathological manifestations. These results strongly suggest that certain phenotypic manifestations of CF may result from remediable alterations in phospholipid-bound arachidonic acid and DHA levels. PMID:10570187

Structure of fructose bisphosphate aldolase from Encephalitozoon cuniculi

PubMed Central

Gardberg, Anna; Sankaran, Banumathi; Davies, Doug; Bhandari, Janhavi; Staker, Bart; Stewart, Lance

2011-01-01

Fructose bisphosphate aldolose (FBPA) enzymes have been found in a broad range of eukaryotic and prokaryotic organisms. FBPA catalyses the cleavage of fructose 1,6-bisphosphate into glyceraldehyde 3-phosphate and dihydroxy­acetone phosphate. The SSGCID has reported several FBPA structures from pathogenic sources. Bioinformatic analysis of the genome of the eukaryotic microsporidian parasite Encephalitozoon cuniculi revealed an FBPA homolog. The structures of this enzyme in the presence of the native substrate FBP and also with the partial substrate analog phosphate are reported. The purified enzyme crystallized in 90 mM Bis-Tris propane pH 6.5, 18% PEG 3350, 18 mM NaKHPO4, 10 mM urea for the phosphate-bound form and 100 mM Bis-Tris propane pH 6.5, 20% PEG 3350, 20 mM fructose 1,6-­bisphosphate for the FBP-bound form. In both cases protein was present at 25 mg ml−1 and the sitting-drop vapour-diffusion method was used. For the FBP-bound form, a data set to 2.37 Å resolution was collected from a single crystal at 100 K. The crystal belonged to the orthorhombic space group C2221, with unit-cell parameters a = 121.46, b = 135.82, c = 61.54 Å. The structure was refined to a final free R factor of 20.8%. For the phosphate-bound form, a data set was collected to 2.00 Å resolution. The space group was also C2221 and the unit-cell parameters were a = 121.96, b = 137.61, c = 62.23 Å. The structure shares the typical barrel tertiary structure reported for previous FBPA structures and exhibits the same Schiff base in the active site. The quaternary structure is dimeric. This work provides a direct experimental result for the substrate-binding conformation of the product state of E. cuniculi FBPA. PMID:21904050

Structure of fructose bisphosphate aldolase from Encephalitozoon cuniculi.

PubMed

Gardberg, Anna; Sankaran, Banumathi; Davies, Doug; Bhandari, Janhavi; Staker, Bart; Stewart, Lance

2011-09-01

Fructose bisphosphate aldolose (FBPA) enzymes have been found in a broad range of eukaryotic and prokaryotic organisms. FBPA catalyses the cleavage of fructose 1,6-bisphosphate into glyceraldehyde 3-phosphate and dihydroxyacetone phosphate. The SSGCID has reported several FBPA structures from pathogenic sources. Bioinformatic analysis of the genome of the eukaryotic microsporidian parasite Encephalitozoon cuniculi revealed an FBPA homolog. The structures of this enzyme in the presence of the native substrate FBP and also with the partial substrate analog phosphate are reported. The purified enzyme crystallized in 90 mM Bis-Tris propane pH 6.5, 18% PEG 3350, 18 mM NaKHPO(4), 10 mM urea for the phosphate-bound form and 100 mM Bis-Tris propane pH 6.5, 20% PEG 3350, 20 mM fructose 1,6-bisphosphate for the FBP-bound form. In both cases protein was present at 25 mg ml(-1) and the sitting-drop vapour-diffusion method was used. For the FBP-bound form, a data set to 2.37 Å resolution was collected from a single crystal at 100 K. The crystal belonged to the orthorhombic space group C222(1), with unit-cell parameters a=121.46, b=135.82, c=61.54 Å. The structure was refined to a final free R factor of 20.8%. For the phosphate-bound form, a data set was collected to 2.00 Å resolution. The space group was also C222(1) and the unit-cell parameters were a=121.96, b=137.61, c=62.23 Å. The structure shares the typical barrel tertiary structure reported for previous FBPA structures and exhibits the same Schiff base in the active site. The quaternary structure is dimeric. This work provides a direct experimental result for the substrate-binding conformation of the product state of E. cuniculi FBPA.

Bounded Community: Designing and Facilitating Learning Communities in Formal Courses

ERIC Educational Resources Information Center

Wilson, Brent G.; Ludwig-Hardman, Stacey; Thornam, Christine L.; Dunlap, Joanna C.

2004-01-01

Learning communities can emerge spontaneously when people find common learning goals and pursue projects and tasks together in pursuit of those goals. "Bounded" learning communities (BLCs) are groups that form within a structured teaching or training setting, typically a course. Unlike spontaneous communities, BLCs develop in direct response to…

Impacts of structuring the electronic health record: Results of a systematic literature review from the perspective of secondary use of patient data.

PubMed

Vuokko, Riikka; Mäkelä-Bengs, Päivi; Hyppönen, Hannele; Lindqvist, Minna; Doupi, Persephone

2017-01-01

To explore the impacts that structuring of electronic health records (EHRs) has had from the perspective of secondary use of patient data as reflected in currently published literature. This paper presents the results of a systematic literature review aimed at answering the following questions; (1) what are the common methods of structuring patient data to serve secondary use purposes; (2) what are the common methods of evaluating patient data structuring in the secondary use context, and (3) what impacts or outcomes of EHR structuring have been reported from the secondary use perspective. The reported study forms part of a wider systematic literature review on the impacts of EHR structuring methods and evaluations of their impact. The review was based on a 12-step systematic review protocol adapted from the Cochrane methodology. Original articles included in the study were divided into three groups for analysis and reporting based on their use focus: nursing documentation, medical use and secondary use (presented in this paper). The analysis from the perspective of secondary use of data includes 85 original articles from 1975 to 2010 retrieved from 15 bibliographic databases. The implementation of structured EHRs can be roughly divided into applications for documenting patient data at the point of care and application for retrieval of patient data (post hoc structuring). Two thirds of the secondary use articles concern EHR structuring methods which were still under development or in the testing phase. of structuring patient data such as codes, terminologies, reference information models, forms or templates and documentation standards were usually applied in combination. Most of the identified benefits of utilizing structured EHR data for secondary use purposes concentrated on information content and quality or on technical quality and reliability, particularly in the case of Natural Language Processing (NLP) studies. A few individual articles evaluated impacts on care processes, productivity and costs, patient safety, care quality or other health impacts. In most articles these endpoints were usually discussed as goals of secondary use and less as evidence-supported impacts, resulting from the use of structured EHR data for secondary purposes. Further studies and more sound evaluation methods are needed for evidence on how EHRs are utilized for secondary purposes, and how structured documentation methods can serve different users' needs, e.g. administration, statistics and research and development, in parallel to medical use purposes. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Supramolecular Structures with Blood Plasma Proteins, Sugars and Nanosilica

NASA Astrophysics Data System (ADS)

Turov, V. V.; Gun'ko, V. M.; Galagan, N. P.; Rugal, A. A.; Barvinchenko, V. M.; Gorbyk, P. P.

Supramolecular structures with blood plasma proteins (albumin, immunoglobulin and fibrinogen (HPF)), protein/water/silica and protein/water/ silica/sugar (glucose, fructose and saccharose) were studied by NMR, adsorption, IR and UV spectroscopy methods. Hydration parameters, amounts of weakly and strongly bound waters and interfacial energy (γ S) were determined over a wide range of component concentrations. The γ S(C protein,C silica) graphs were used to estimate the energy of protein-protein, protein-surface and particle-particle interactions. It was shown that interfacial energy of self-association (γ as) of protein molecules depends on a type of proteins. A large fraction of water bound to proteins can be displaced by sugars, and the effect of disaccharide (saccharose) was greater than that of monosugars. Changes in the structural parameters of cavities in HPF molecules and complexes with HPF/silica nanoparticles filled by bound water were analysed using NMR-cryoporometry showing that interaction of proteins with silica leads to a significant decrease in the amounts of water bound to both protein and silica surfaces. Bionanocomposites with BSA/nanosilica/sugar can be used to influence states of living cells and tissues after cryopreservation or other treatments. It was shown that interaction of proteins with silica leads to strong decrease in the volume of all types of internal cavities filled by water.

Fragment-based modelling of single stranded RNA bound to RNA recognition motif containing proteins

PubMed Central

de Beauchene, Isaure Chauvot; de Vries, Sjoerd J.; Zacharias, Martin

2016-01-01

Abstract Protein-RNA complexes are important for many biological processes. However, structural modeling of such complexes is hampered by the high flexibility of RNA. Particularly challenging is the docking of single-stranded RNA (ssRNA). We have developed a fragment-based approach to model the structure of ssRNA bound to a protein, based on only the protein structure, the RNA sequence and conserved contacts. The conformational diversity of each RNA fragment is sampled by an exhaustive library of trinucleotides extracted from all known experimental protein–RNA complexes. The method was applied to ssRNA with up to 12 nucleotides which bind to dimers of the RNA recognition motifs (RRMs), a highly abundant eukaryotic RNA-binding domain. The fragment based docking allows a precise de novo atomic modeling of protein-bound ssRNA chains. On a benchmark of seven experimental ssRNA–RRM complexes, near-native models (with a mean heavy-atom deviation of <3 Å from experiment) were generated for six out of seven bound RNA chains, and even more precise models (deviation < 2 Å) were obtained for five out of seven cases, a significant improvement compared to the state of the art. The method is not restricted to RRMs but was also successfully applied to Pumilio RNA binding proteins. PMID:27131381

Fundamental limits in 3D landmark localization.

PubMed

Rohr, Karl

2005-01-01

This work analyses the accuracy of estimating the location of 3D landmarks and characteristic image structures. Based on nonlinear estimation theory we study the minimal stochastic errors of the position estimate caused by noisy data. Given analytic models of the image intensities we derive closed-form expressions for the Cramér-Rao bound for different 3D structures such as 3D edges, 3D ridges, 3D lines, and 3D blobs. It turns out, that the precision of localization depends on the noise level, the size of the region-of-interest, the width of the intensity transitions, as well as on other parameters describing the considered image structure. The derived lower bounds can serve as benchmarks and the performance of existing algorithms can be compared with them. To give an impression of the achievable accuracy numeric examples are presented. Moreover, by experimental investigations we demonstrate that the derived lower bounds can be achieved by fitting parametric intensity models directly to the image data.

Fluoride ion encapsulation by Mg2+ and phosphates in a fluoride riboswitch

PubMed Central

Ren, Aiming; Rajashankar, Kanagalaghatta R.; Patel, Dinshaw J.

2012-01-01

Significant advances in our understanding of RNA architecture, folding and recognition have emerged from structure-function studies on riboswicthes, non-coding RNAs whose sensing domains bind small ligands and whose adjacent expression platforms contain RNA elements involved in the control of gene regulation. We now report on the ligand-bound structure of the Thermotoga petrophila fluoride riboswitch, which adopts a higher-order RNA architecture stabilized by pseudoknot and long-range reversed Watson-Crick and Hoogsteen A•U pair formation. The bound fluoride ion is encapsulated within the junctional architecture, anchored in place through direct coordination to three Mg2+ ions, which in turn are octahedrally coordinated to waters and five inwardly-pointing backbone phosphates. Our structure of the fluoride riboswitch in the bound state defines how RNA can form a binding pocket selective for fluoride, while discriminating against larger halide ions. The T. petrophila fluoride riboswitch most likely functions in gene regulation through a transcription termination mechanism. PMID:22678284

Lattice QCD evidence that the Λ(1405) resonance is an antikaon-nucleon molecule.

PubMed

Hall, Jonathan M M; Kamleh, Waseem; Leinweber, Derek B; Menadue, Benjamin J; Owen, Benjamin J; Thomas, Anthony W; Young, Ross D

2015-04-03

For almost 50 years the structure of the Λ(1405) resonance has been a mystery. Even though it contains a heavy strange quark and has odd parity, its mass is lower than any other excited spin-1/2 baryon. Dalitz and co-workers speculated that it might be a molecular state of an antikaon bound to a nucleon. However, a standard quark-model structure is also admissible. Although the intervening years have seen considerable effort, there has been no convincing resolution. Here we present a new lattice QCD simulation showing that the strange magnetic form factor of the Λ(1405) vanishes, signaling the formation of an antikaon-nucleon molecule. Together with a Hamiltonian effective-field-theory model analysis of the lattice QCD energy levels, this strongly suggests that the structure is dominated by a bound antikaon-nucleon component. This result clarifies that not all states occurring in nature can be described within a simple quark model framework and points to the existence of exotic molecular meson-nucleon bound states.

Structural Insights into the Allosteric Operation of the Lon AAA+ Protease.

PubMed

Lin, Chien-Chu; Su, Shih-Chieh; Su, Ming-Yuan; Liang, Pi-Hui; Feng, Chia-Cheng; Wu, Shih-Hsiung; Chang, Chung-I

2016-05-03

The Lon AAA+ protease (LonA) is an evolutionarily conserved protease that couples the ATPase cycle into motion to drive substrate translocation and degradation. A hallmark feature shared by AAA+ proteases is the stimulation of ATPase activity by substrates. Here we report the structure of LonA bound to three ADPs, revealing the first AAA+ protease assembly where the six protomers are arranged alternately in nucleotide-free and bound states. Nucleotide binding induces large coordinated movements of conserved pore loops from two pairs of three non-adjacent protomers and shuttling of the proteolytic groove between the ATPase site and a previously unknown Arg paddle. Structural and biochemical evidence supports the roles of the substrate-bound proteolytic groove in allosteric stimulation of ATPase activity and the conserved Arg paddle in driving substrate degradation. Altogether, this work provides a molecular framework for understanding how ATP-dependent chemomechanical movements drive allosteric processes for substrate degradation in a major protein-destruction machine. Copyright © 2016 Elsevier Ltd. All rights reserved.

An analytical study of nitrogen oxides and carbon monoxide emissions in hydrocarbon combustion with added nitrogen - Preliminary results

NASA Technical Reports Server (NTRS)

Bittker, D. A.

1980-01-01

The influence of ground-based gas turbine combustor operating conditions and fuel-bound nitrogen (FBN) found in coal-derived liquid fuels on the formation of nitrogen oxides and carbon monoxide is investigated. Analytical predictions of NOx and CO concentrations are obtained for a two-stage, adiabatic, perfectly-stirred reactor operating on a propane-air mixture, with primary equivalence ratios from 0.5 to 1.7, secondary equivalence ratios of 0.5 or 0.7, primary stage residence times from 12 to 20 msec, secondary stage residence times of 1, 2 and 3 msec and fuel nitrogen contents of 0.5, 1.0 and 2.0 wt %. Minimum nitrogen oxide but maximum carbon monoxide formation is obtained at primary zone equivalence ratios between 1.4 and 1.5, with percentage conversion of FBN to NOx decreasing with increased fuel nitrogen content. Additional secondary dilution is observed to reduce final pollutant concentrations, with NOx concentration independent of secondary residence time and CO decreasing with secondary residence time; primary zone residence time is not observed to affect final NOx and CO concentrations significantly. Finally, comparison of computed results with experimental values shows a good semiquantitative agreement.

An experimental study of secondary vortex structure in mixing layers

NASA Technical Reports Server (NTRS)

Bell, J. H.; Mehta, Rabindra D.

1990-01-01

This report covers the first eight months of an experimental research project on the secondary vortex structure in plane mixing layers. The aim of the project is to obtain quantitative data on the behavior of the secondary structure in a turbulent mixing layer at reasonable reynolds numbers (Re(sub delta(sub w)) approx. 50,000). In particular, we hope to resolve the questions of how the scale of the secondary vortex structure changes with the scale of the mixing layer, and whether the structures are fixed in space, or whether they 'meander' in the spanwise direction.

Crystal structures of the CBS and DRTGG domains of the regulatory region of Clostridiumperfringens pyrophosphatase complexed with the inhibitor, AMP, and activator, diadenosine tetraphosphate.

PubMed

Tuominen, H; Salminen, A; Oksanen, E; Jämsen, J; Heikkilä, O; Lehtiö, L; Magretova, N N; Goldman, A; Baykov, A A; Lahti, R

2010-05-07

Nucleotide-binding cystathionine beta-synthase (CBS) domains serve as regulatory units in numerous proteins distributed in all kingdoms of life. However, the underlying regulatory mechanisms remain to be established. Recently, we described a subfamily of CBS domain-containing pyrophosphatases (PPases) within family II PPases. Here, we express a novel CBS-PPase from Clostridium perfringens (CPE2055) and show that the enzyme is inhibited by AMP and activated by a novel effector, diadenosine 5',5-P1,P4-tetraphosphate (AP(4)A). The structures of the AMP and AP(4)A complexes of the regulatory region of C. perfringens PPase (cpCBS), comprising a pair of CBS domains interlinked by a DRTGG domain, were determined at 2.3 A resolution using X-ray crystallography. The structures obtained are the first structures of a DRTGG domain as part of a larger protein structure. The AMP complex contains two AMP molecules per cpCBS dimer, each bound to a single monomer, whereas in the activator-bound complex, one AP(4)A molecule bridges two monomers. In the nucleotide-bound structures, activator binding induces significant opening of the CBS domain interface, compared with the inhibitor complex. These results provide structural insight into the mechanism of CBS-PPase regulation by nucleotides. Copyright 2010 Elsevier Ltd. All rights reserved.

Computation-Guided Backbone Grafting of a Discontinuous Motif onto a Protein Scaffold

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

Azoitei, Mihai L.; Correia, Bruno E.; Ban, Yih-En Andrew

2012-02-07

The manipulation of protein backbone structure to control interaction and function is a challenge for protein engineering. We integrated computational design with experimental selection for grafting the backbone and side chains of a two-segment HIV gp120 epitope, targeted by the cross-neutralizing antibody b12, onto an unrelated scaffold protein. The final scaffolds bound b12 with high specificity and with affinity similar to that of gp120, and crystallographic analysis of a scaffold bound to b12 revealed high structural mimicry of the gp120-b12 complex structure. The method can be generalized to design other functional proteins through backbone grafting.

Single-Sex Schools, Student Achievement, and Course Selection: Evidence from Rule-Based Student Assignments in Trinidad and Tobago. NBER Working Paper No. 16817

ERIC Educational Resources Information Center

Jackson, C. Kirabo

2011-01-01

Existing studies on single-sex schooling suffer from biases due to student selection to schools and single-sex schools being better in unmeasured ways. In Trinidad and Tobago students are assigned to secondary schools based on an algorithm allowing one to address self-selection bias and cleanly estimate an upper-bound single-sex school effect. The…

Antibodies with 'Original Antigenic Sin' Properties Are Valuable Components of Secondary Immune Responses to Influenza Viruses.

PubMed

Linderman, Susanne L; Hensley, Scott E

2016-08-01

Human antibodies (Abs) elicited by influenza viruses often bind with a high affinity to past influenza virus strains, but paradoxically, do not bind to the viral strain actually eliciting the response. This phenomena is called 'original antigenic sin' (OAS) since this can occur at the expense of generating new de novo Abs. Here, we characterized the specificity and functionality of Abs elicited in mice that were sequentially exposed to two antigenically distinct H1N1 influenza virus strains. Many Abs elicited under these conditions had an OAS phenotype, in that they bound strongly to the viral strain used for the first exposure and very weakly to the viral strain used for the second exposure. We found that OAS and non-OAS Abs target the same general region of the influenza hemagglutinin protein and that B cells expressing these two types of Abs can be clonally-related. Surprisingly, although OAS Abs bound with very low affinities, some were able to effectively protect against an antigenically drifted viral strain following passive transfer in vivo. Taken together, our data indicate that OAS Abs share some level of cross-reactivity between priming and recall viral strains and that B cells producing these Abs can be protective when recalled into secondary immune responses.

Convergence of an iterative procedure for large-scale static analysis of structural components

NASA Technical Reports Server (NTRS)

Austin, F.; Ojalvo, I. U.

1976-01-01

The paper proves convergence of an iterative procedure for calculating the deflections of built-up component structures which can be represented as consisting of a dominant, relatively stiff primary structure and a less stiff secondary structure, which may be composed of one or more substructures that are not connected to one another but are all connected to the primary structure. The iteration consists in estimating the deformation of the primary structure in the absence of the secondary structure on the assumption that all mechanical loads are applied directly to the primary structure. The j-th iterate primary structure deflections at the interface are imposed on the secondary structure, and the boundary loads required to produce these deflections are computed. The cycle is completed by applying the interface reaction to the primary structure and computing its updated deflections. It is shown that the mathematical condition for convergence of this procedure is that the maximum eigenvalue of the equation relating primary-structure deflection to imposed secondary-structure deflection be less than unity, which is shown to correspond with the physical requirement that the secondary structure be more flexible at the interface boundary.

Personal goals as predictors of intended classroom goals: comparing elementary and secondary school pre-service teachers.

PubMed

Daniels, Lia M; Frenzel, Anne C; Stupnisky, Robert H; Stewart, Tara L; Perry, Raymond P

2013-09-01

The literature documents fewer classroom mastery goal structures in secondary school compared to elementary. However, little is known about how personal achievement goals may influence classroom goal structures. This is especially true at the level of pre-service teachers. Our objective was to investigate if pre-service teachers' personal goals predicted their intended classroom goal structures. Participants were 125 elementary and 175 secondary school pre-service teachers from two Western Canadian universities. Structural equation modelling was used to examine if the structural relationships and latent means of personal and intended classroom goal structures differed for elementary and secondary school pre-service teachers. The results revealed that personal goals predicted the goal structures that pre-service teachers intended to establish; however, the relationships and means differed between elementary and secondary school pre-service teachers. Specifically, personal mastery-approach goals positively predicted classroom mastery goals much more strongly at the elementary than the secondary level. Furthermore, elementary pre-service teachers had significantly higher latent mean scores on personal mastery-approach goals than their secondary counterparts. It seems possible that the currently documented differences between classroom goal structures noted for elementary compared to secondary school may be based on the personal goals endorsed as pre-service teachers. The results are further discussed in terms of alignment with research on practising teachers' personal and classroom goals and implications for teacher education. © 2012 The British Psychological Society.

Photoswitching a Molecular Catalyst to Regulate CO 2 Hydrogenation

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

Priyadarshani, Nilusha; Ginovska-Pangovska, Bojana; Bays, J. Timothy

2015-07-24

Inspired by nature’s ability to regulate catalysis using physiological stimuli, azobenzene was incorporated into Rh(bis)diphosphine CO 2 hydrogenation catalysts to photoinitiate structural changes to modulate the resulting catalytic activity. The rhodium bound diphosphine ligands (P(Ph 2)-CH 2-N(R) CH 2-P(Ph2)) contain the terminal amine of a non-natural amino acid, either β-alanine (β-Ala) or γ-aminobutyric acid (GABA). For both β-ala and GABA containing complexes, the carboxylic acids of the amino acids were coupled to the amines of diaminoazobenzene, creating a rhodium bound trans-spanning tetraphosphine complex. The photo-induced cis-trans isomerization of the azobenzene-containing complexes imposes structural changes on these complexes, as evidenced bymore » 1H NMR. We found that the CO 2 hydrogenation activity for the β-ala bound Rh complex is 40% faster with azobenzene in the cis configuration (16 s -1) than in the trans conformation (11 s -1), while the γ-aminobutyric acid containing Rh complex has the same rate (~17 s -1) in either the cis or the trans configuration at 27 °C. The corresponding complexes without the attached azobenzene were also prepared, characterized, and catalytically tested for comparison, and have rates of 30 s -1. Computational studies were undertaken to evaluate the difference in rate between the cis and trans isomers for the β-Ala bound Rh complex, and revealed major structural changes between all cis and trans structures, but only minor structural changes that would be unique to the β-Ala bound Rh complex. We postulate that the slower rate of the azobenzene-containing β-Ala bound Rh complex is due to subtle changes in the bite angle arising from steric strain due to the trans-spanning azobenzene, altering hydricity and consequently rate. This work was supported by the US Department of Energy, Office of Science, Office of Basic Energy Sciences, Division of Chemical Sciences, Geosciences & Biosciences. Pacific Northwest National Laboratory (PNNL) is a multiprogram national laboratory operated for the DOE by Battelle. A portion of this research was performed using EMSL, a national scientific user facility sponsored by the Department of Energy's Office of Biological and Environmental Research and located at Pacific Northwest National Laboratory.« less

Metastable ripple phase of fully hydrated dipalmitoylphosphatidylcholine as studied by small angle x-ray scattering

PubMed Central

Yao, Haruhiko; Matuoka, Sinzi; Tenchov, Boris; Hatta, Ichiro

1991-01-01

Fully hydrated dipalmitoylphosphatidylcholine (DPPC) undergoes liquid crystalline to metastable Pβ, phase transition in cooling. A small angle x-ray scattering study has been performed for obtaining further evidence about the structure of this phase. From a high-resolution observation of x-ray diffraction profiles, a distinct multipeak pattern has become obvious. Among them the (01) reflection in the secondary ripple structure is identified clearly. There are peaks assigned straightforwardly to (10) and (20) reflections in the primary ripple structure and peaks assigned to (10) and (20) reflections in the secondary ripple structure. Therefore the multipeak pattern is due to superposition of the reflections cause by the primary and secondary ripple structures. The lattice parameters are estimated as follows: for the primary ripple structure a = 7.09 nm, b = 13.64 nm, and γ = 95°, and for the secondary ripple structure a = 8.2 nm, b = 26.6 nm, and γ = 90°. The lattice parameters thus obtained for the secondary ripple structure are not conclusive, however. The hydrocarbon chains in the primary ripple structure have been reported as being tilted against the bilayer plane and, on the other hand, the hydrocarbon chains in the secondary ripple structure are likely to be perpendicular to the bilayer plane. This fact seems to be related to a sequential mechanism of phase transitions. On heating from the Lβ, phase where the hydrocarbon chains are tilted the primary ripple structure having tilted hydrocarbon chains takes place and on cooling from the Lα phase where the hydrocarbon chains are not tilted the secondary ripple structure with untilted chains tends to be stabilized. It appears that the truly metastable ripple phase is expressed by the second ripple structure although in the course of the actual cooling transition both the secondary and primary ripple structures form and coexist. PMID:19431787

Double-Wall Carbon Nanotube Hybrid Mode-Locker in Tm-doped Fibre Laser: A Novel Mechanism for Robust Bound-State Solitons Generation

NASA Astrophysics Data System (ADS)

Chernysheva, Maria; Bednyakova, Anastasia; Al Araimi, Mohammed; Howe, Richard C. T.; Hu, Guohua; Hasan, Tawfique; Gambetta, Alessio; Galzerano, Gianluca; Rümmeli, Mark; Rozhin, Aleksey

2017-03-01

The complex nonlinear dynamics of mode-locked fibre lasers, including a broad variety of dissipative structures and self-organization effects, have drawn significant research interest. Around the 2 μm band, conventional saturable absorbers (SAs) possess small modulation depth and slow relaxation time and, therefore, are incapable of ensuring complex inter-pulse dynamics and bound-state soliton generation. We present observation of multi-soliton complex generation in mode-locked thulium (Tm)-doped fibre laser, using double-wall carbon nanotubes (DWNT-SA) and nonlinear polarisation evolution (NPE). The rigid structure of DWNTs ensures high modulation depth (64%), fast relaxation (1.25 ps) and high thermal damage threshold. This enables formation of 560-fs soliton pulses; two-soliton bound-state with 560 fs pulse duration and 1.37 ps separation; and singlet+doublet soliton structures with 1.8 ps duration and 6 ps separation. Numerical simulations based on the vectorial nonlinear Schr¨odinger equation demonstrate a transition from single-pulse to two-soliton bound-states generation. The results imply that DWNTs are an excellent SA for the formation of steady single- and multi-soliton structures around 2 μm region, which could not be supported by single-wall carbon nanotubes (SWNTs). The combination of the potential bandwidth resource around 2 μm with the soliton molecule concept for encoding two bits of data per clock period opens exciting opportunities for data-carrying capacity enhancement.

Algorithm-Dependent Generalization Bounds for Multi-Task Learning.

PubMed

Liu, Tongliang; Tao, Dacheng; Song, Mingli; Maybank, Stephen J

2017-02-01

Often, tasks are collected for multi-task learning (MTL) because they share similar feature structures. Based on this observation, in this paper, we present novel algorithm-dependent generalization bounds for MTL by exploiting the notion of algorithmic stability. We focus on the performance of one particular task and the average performance over multiple tasks by analyzing the generalization ability of a common parameter that is shared in MTL. When focusing on one particular task, with the help of a mild assumption on the feature structures, we interpret the function of the other tasks as a regularizer that produces a specific inductive bias. The algorithm for learning the common parameter, as well as the predictor, is thereby uniformly stable with respect to the domain of the particular task and has a generalization bound with a fast convergence rate of order O(1/n), where n is the sample size of the particular task. When focusing on the average performance over multiple tasks, we prove that a similar inductive bias exists under certain conditions on the feature structures. Thus, the corresponding algorithm for learning the common parameter is also uniformly stable with respect to the domains of the multiple tasks, and its generalization bound is of the order O(1/T), where T is the number of tasks. These theoretical analyses naturally show that the similarity of feature structures in MTL will lead to specific regularizations for predicting, which enables the learning algorithms to generalize fast and correctly from a few examples.

CHARMM Force-Fields with Modified Polyphosphate Parameters Allow Stable Simulation of the ATP-Bound Structure of Ca(2+)-ATPase.

PubMed

Komuro, Yasuaki; Re, Suyong; Kobayashi, Chigusa; Muneyuki, Eiro; Sugita, Yuji

2014-09-09

Adenosine triphosphate (ATP) is an indispensable energy source in cells. In a wide variety of biological phenomena like glycolysis, muscle contraction/relaxation, and active ion transport, chemical energy released from ATP hydrolysis is converted to mechanical forces to bring about large-scale conformational changes in proteins. Investigation of structure-function relationships in these proteins by molecular dynamics (MD) simulations requires modeling of ATP in solution and ATP bound to proteins with accurate force-field parameters. In this study, we derived new force-field parameters for the triphosphate moiety of ATP based on the high-precision quantum calculations of methyl triphosphate. We tested our new parameters on membrane-embedded sarcoplasmic reticulum Ca(2+)-ATPase and four soluble proteins. The ATP-bound structure of Ca(2+)-ATPase remains stable during MD simulations, contrary to the outcome in shorter simulations using original parameters. Similar results were obtained with the four ATP-bound soluble proteins. The new force-field parameters were also tested by investigating the range of conformations sampled during replica-exchange MD simulations of ATP in explicit water. Modified parameters allowed a much wider range of conformational sampling compared with the bias toward extended forms with original parameters. A diverse range of structures agrees with the broad distribution of ATP conformations in proteins deposited in the Protein Data Bank. These simulations suggest that the modified parameters will be useful in studies of ATP in solution and of the many ATP-utilizing proteins.

Stem-Loop RNA Hairpins in Giant Viruses: Invading rRNA-Like Repeats and a Template Free RNA

PubMed Central

Seligmann, Hervé; Raoult, Didier

2018-01-01

We examine the hypothesis that de novo template-free RNAs still form spontaneously, as they did at the origins of life, invade modern genomes, contribute new genetic material. Previously, analyses of RNA secondary structures suggested that some RNAs resembling ancestral (t)RNAs formed recently de novo, other parasitic sequences cluster with rRNAs. Here positive control analyses of additional RNA secondary structures confirm ancestral and de novo statuses of RNA grouped according to secondary structure. Viroids with branched stems resemble de novo RNAs, rod-shaped viroids resemble rRNA secondary structures, independently of GC contents. 5′ UTR leading regions of West Nile and Dengue flavivirid viruses resemble de novo and rRNA structures, respectively. An RNA homologous with Megavirus, Dengue and West Nile genomes, copperhead snake microsatellites and levant cotton repeats, not templated by Mimivirus' genome, persists throughout Mimivirus' infection. Its secondary structure clusters with candidate de novo RNAs. The saltatory phyletic distribution and secondary structure of Mimivirus' peculiar RNA suggest occasional template-free polymerization of this sequence, rather than noncanonical transcriptions (swinger polymerization, posttranscriptional editing). PMID:29449833

Entropy-Driven Folding of an RNA Helical Junction: An Isothermal Titration Calorimetric Analysis of the Hammerhead Ribozyme†

PubMed Central

Mikulecky, Peter J.; Takach, Jennifer C.; Feig, Andrew L.

2008-01-01

Helical junctions are extremely common motifs in naturally occurring RNAs, but little is known about the thermodynamics that drive their folding. Studies of junction folding face several challenges: non-two-state folding behavior, superposition of secondary and tertiary structural energetics, and drastically opposing enthalpic and entropic contributions to folding. Here we describe a thermodynamic dissection of the folding of the hammerhead ribozyme, a three-way RNA helical junction, by using isothermal titration calorimetry of bimolecular RNA constructs. By using this method, we show that tertiary folding of the hammerhead core occurs with a highly unfavorable enthalpy change, and is therefore entropically driven. Furthermore, the enthalpies and heat capacities of core folding are the same whether supported by monovalent or divalent ions. These properties appear to be general to the core sequence of bimolecular hammerhead constructs. We present a model for the ion-induced folding of the hammerhead core that is similar to those advanced for the folding of much larger RNAs, involving ion-induced collapse to a structured, non-native state accompanied by rearrangement of core residues to produce the native fold. In agreement with previous enzymological and structural studies, our thermodynamic data suggest that the hammerhead structure is stabilized in vitro predominantly by diffusely bound ions. Our approach addresses several significant challenges that accompany the study of junction folding, and should prove useful in defining the thermodynamic determinants of stability in these important RNA motifs. PMID:15134461

Short-distance probes for protein backbone structure based on energy transfer between bimane and transition metal ions

PubMed Central

Taraska, Justin W.; Puljung, Michael C.; Zagotta, William N.

2009-01-01

The structure and dynamics of proteins underlies the workings of virtually every biological process. Existing biophysical methods are inadequate to measure protein structure at atomic resolution, on a rapid time scale, with limited amounts of protein, and in the context of a cell or membrane. FRET can measure distances between two probes, but depends on the orientation of the probes and typically works only over long distances comparable with the size of many proteins. Also, common probes used for FRET can be large and have long, flexible attachment linkers that position dyes far from the protein backbone. Here, we improve and extend a fluorescence method called transition metal ion FRET that uses energy transfer to transition metal ions as a reporter of short-range distances in proteins with little orientation dependence. This method uses a very small cysteine-reactive dye monobromobimane, with virtually no linker, and various transition metal ions bound close to the peptide backbone as the acceptor. We show that, unlike larger fluorophores and longer linkers, this donor–acceptor pair accurately reports short-range distances and changes in backbone distances. We further extend the method by using cysteine-reactive metal chelators, which allow the technique to be used in protein regions of unknown secondary structure or when native metal ion binding sites are present. This improved method overcomes several of the key limitations of classical FRET for intramolecular distance measurements. PMID:19805285

Structural basis of Na(+)-independent and cooperative substrate/product antiport in CaiT.

PubMed

Schulze, Sabrina; Köster, Stefan; Geldmacher, Ulrike; Terwisscha van Scheltinga, Anke C; Kühlbrandt, Werner

2010-09-09

Transport of solutes across biological membranes is performed by specialized secondary transport proteins in the lipid bilayer, and is essential for life. Here we report the structures of the sodium-independent carnitine/butyrobetaine antiporter CaiT from Proteus mirabilis (PmCaiT) at 2.3-A and from Escherichia coli (EcCaiT) at 3.5-A resolution. CaiT belongs to the family of betaine/carnitine/choline transporters (BCCT), which are mostly Na(+) or H(+) dependent, whereas EcCaiT is Na(+) and H(+) independent. The three-dimensional architecture of CaiT resembles that of the Na(+)-dependent transporters LeuT and BetP, but in CaiT a methionine sulphur takes the place of the Na(+) ion to coordinate the substrate in the central transport site, accounting for Na(+)-independent transport. Both CaiT structures show the fully open, inward-facing conformation, and thus complete the set of functional states that describe the alternating access mechanism. EcCaiT contains two bound butyrobetaine substrate molecules, one in the central transport site, the other in an extracellular binding pocket. In the structure of PmCaiT, a tryptophan side chain occupies the transport site, and access to the extracellular site is blocked. Binding of both substrates to CaiT reconstituted into proteoliposomes is cooperative, with Hill coefficients up to 1.7, indicating that the extracellular site is regulatory. We propose a mechanism whereby the occupied regulatory site increases the binding affinity of the transport site and initiates substrate translocation.

Solution structure of the C-terminal X domain of the measles virus phosphoprotein and interaction with the intrinsically disordered C-terminal domain of the nucleoprotein.

PubMed

Gely, Stéphane; Lowry, David F; Bernard, Cédric; Jensen, Malene R; Blackledge, Martin; Costanzo, Stéphanie; Bourhis, Jean-Marie; Darbon, Hervé; Daughdrill, Gary; Longhi, Sonia

2010-01-01

In this report, the solution structure of the nucleocapsid-binding domain of the measles virus phosphoprotein (XD, aa 459-507) is described. A dynamic description of the interaction between XD and the disordered C-terminal domain of the nucleocapsid protein, (N(TAIL), aa 401-525), is also presented. XD is an all alpha protein consisting of a three-helix bundle with an up-down-up arrangement of the helices. The solution structure of XD is very similar to the crystal structures of both the free and bound form of XD. One exception is the presence of a highly dynamic loop encompassing XD residues 489-491, which is involved in the embedding of the alpha-helical XD-binding region of N(TAIL). Secondary chemical shift values for full-length N(TAIL) were used to define the precise boundaries of a transient helical segment that coincides with the XD-binding domain, thus shedding light on the pre-recognition state of N(TAIL). Titration experiments with unlabeled XD showed that the transient alpha-helical conformation of N(TAIL) is stabilized upon binding. Lineshape analysis of NMR resonances revealed that residues 483-506 of N(TAIL) are in intermediate exchange with XD, while the 475-482 and 507-525 regions are in fast exchange. The N(TAIL) resonance behavior in the titration experiments is consistent with a complex binding model with more than two states.

Substrate sequence selectivity of APOBEC3A implicates intra-DNA interactions.

PubMed

Silvas, Tania V; Hou, Shurong; Myint, Wazo; Nalivaika, Ellen; Somasundaran, Mohan; Kelch, Brian A; Matsuo, Hiroshi; Kurt Yilmaz, Nese; Schiffer, Celia A

2018-05-14

The APOBEC3 (A3) family of human cytidine deaminases is renowned for providing a first line of defense against many exogenous and endogenous retroviruses. However, the ability of these proteins to deaminate deoxycytidines in ssDNA makes A3s a double-edged sword. When overexpressed, A3s can mutate endogenous genomic DNA resulting in a variety of cancers. Although the sequence context for mutating DNA varies among A3s, the mechanism for substrate sequence specificity is not well understood. To characterize substrate specificity of A3A, a systematic approach was used to quantify the affinity for substrate as a function of sequence context, length, secondary structure, and solution pH. We identified the A3A ssDNA binding motif as (T/C)TC(A/G), which correlated with enzymatic activity. We also validated that A3A binds RNA in a sequence specific manner. A3A bound tighter to substrate binding motif within a hairpin loop compared to linear oligonucleotide, suggesting A3A affinity is modulated by substrate structure. Based on these findings and previously published A3A-ssDNA co-crystal structures, we propose a new model with intra-DNA interactions for the molecular mechanism underlying A3A sequence preference. Overall, the sequence and structural preferences identified for A3A leads to a new paradigm for identifying A3A's involvement in mutation of endogenous or exogenous DNA.

Protein structure modeling and refinement by global optimization in CASP12.

PubMed

Hong, Seung Hwan; Joung, InSuk; Flores-Canales, Jose C; Manavalan, Balachandran; Cheng, Qianyi; Heo, Seungryong; Kim, Jong Yun; Lee, Sun Young; Nam, Mikyung; Joo, Keehyoung; Lee, In-Ho; Lee, Sung Jong; Lee, Jooyoung

2018-03-01

For protein structure modeling in the CASP12 experiment, we have developed a new protocol based on our previous CASP11 approach. The global optimization method of conformational space annealing (CSA) was applied to 3 stages of modeling: multiple sequence-structure alignment, three-dimensional (3D) chain building, and side-chain re-modeling. For better template selection and model selection, we updated our model quality assessment (QA) method with the newly developed SVMQA (support vector machine for quality assessment). For 3D chain building, we updated our energy function by including restraints generated from predicted residue-residue contacts. New energy terms for the predicted secondary structure and predicted solvent accessible surface area were also introduced. For difficult targets, we proposed a new method, LEEab, where the template term played a less significant role than it did in LEE, complemented by increased contributions from other terms such as the predicted contact term. For TBM (template-based modeling) targets, LEE performed better than LEEab, but for FM targets, LEEab was better. For model refinement, we modified our CASP11 molecular dynamics (MD) based protocol by using explicit solvents and tuning down restraint weights. Refinement results from MD simulations that used a new augmented statistical energy term in the force field were quite promising. Finally, when using inaccurate information (such as the predicted contacts), it was important to use the Lorentzian function for which the maximal penalty arising from wrong information is always bounded. © 2017 Wiley Periodicals, Inc.

Chemical structure, comparison antioxidant capacity and separation antioxidant of hen, duck and quail egg white protein hydrolysate

NASA Astrophysics Data System (ADS)

Fatah, A.; Meihu, M.; Ning, Q.; Setiani, B. E.; Bintoro, V. P.

2018-01-01

Amino acid linkages as proteins are nutritional substance which important for diet intake. Purification protein procesing undergo heating procedure process followed by additional of proteolytic enzymes or acid had been resulting in protein hydrolysates. A protein hydrolysate describe as many free amino acids bound together through a complex mixture of peptides. Egg white protein hydrolysates is one of subject interested to study for human health or industry product. The objectives of the research are to determine and identification the antioxidant derived from egg white hydrolysate protein. Identification of chemical structure of albumen and albumen protein hydrolysate was examine using IR Spectrophotometry. While comparison of antioxidant capacity and antioxidant separation egg albumen was also investigate using FTIR method (Fourier Transform Infrared Spectroscopy). Hen, duck and quail albumen egg white and on hydrolisate form were used as research materials. The results were showing that different time and enzyme of hydrolysis were not influence at secondary structure of hydrolysate albumen protein. Phytochemical content such as alcohol and hydroxyl compound which have potential as functional group of antioxidant were detected in all of the samples. Their results of radical scavenging activities samples hydrolyzed by pepsin were respectively 89.40%, 50.25% and 85.13%. Whereas the radical scavenging activities of hydrolysates hydrolyzed by papain were 72.85%, 61% and 76.45% respectively.

Molecular mechanism of DNA association with single-stranded DNA binding protein

PubMed Central

Maffeo, Christopher

2017-01-01

Abstract During DNA replication, the single-stranded DNA binding protein (SSB) wraps single-stranded DNA (ssDNA) with high affinity to protect it from degradation and prevent secondary structure formation. Although SSB binds ssDNA tightly, it can be repositioned along ssDNA to follow the advancement of the replication fork. Using all-atom molecular dynamics simulations, we characterized the molecular mechanism of ssDNA association with SSB. Placed in solution, ssDNA–SSB assemblies were observed to change their structure spontaneously; such structural changes were suppressed in the crystallographic environment. Repeat simulations of the SSB–ssDNA complex under mechanical tension revealed a multitude of possible pathways for ssDNA to come off SSB punctuated by prolonged arrests at reproducible sites at the SSB surface. Ensemble simulations of spontaneous association of short ssDNA fragments with SSB detailed a three-dimensional map of local affinity to DNA; the equilibrium amount of ssDNA bound to SSB was found to depend on the electrolyte concentration but not on the presence of the acidic tips of the SSB tails. Spontaneous formation of ssDNA bulges and their diffusive motion along SSB surface was directly observed in multiple 10-µs-long simulations. Such reptation-like motion was confined by DNA binding to high-affinity spots, suggesting a two-step mechanism for SSB diffusion. PMID:29059392

Trifluoperazine Regulation of Calmodulin Binding to Fas: A Computational Study

PubMed Central

Pan, Di; Yan, Qi; Chen, Yabing; McDonald, Jay M; Song, Yuhua

2011-01-01

Death-inducing signaling complex (DISC) formation is a critical step in Fas-mediated signaling for apoptosis. Previous experiments have demonstrated that the calmodulin (CaM) antagonist, trifluoperazine (TFP) regulates CaM-Fas binding and affects Fas-mediated DISC formation. In this study, we investigated the anti-cooperative characteristics of TFP binding to CaM and the effect of TFP on the CaM-Fas interaction from both structural and thermodynamic perspectives using combined molecular dynamics simulations and binding free energy analyses. We studied the interactions of different numbers of TFP molecules with CaM and explored the effects of the resulting conformational changes in CaM on CaM-Fas binding. Results from these analyses showed that the number of TFP molecules bound to CaM directly influenced α-helix formation and hydrogen bond occupancy within the α-helices of CaM, contributing to the conformational and motion changes in CaM. These changes affected CaM binding to Fas, resulting in secondary structural changes in Fas and conformational and motion changes of Fas in CaM-Fas complexes, potentially perturbing the recruitment of Fas-associated death domain (FADD) for DISC formation. The computational results from this study reveal the structural and molecular mechanisms that underlie the role of the CaM antagonist, TFP, in regulation of CaM-Fas binding and Fas-mediated DISC formation in a concentration-dependent manner. PMID:21656570

The effect of Berberine on the secondary structure of human serum albumin

NASA Astrophysics Data System (ADS)

Li, Ying; He, WenYing; Tian, Jianniao; Tang, Jianghong; Hu, Zhide; Chen, Xingguo

2005-05-01

The presence of several high affinity binding sites on human serum albumin (HSA) makes it a possible target for many drugs. This study is designed to examine the effect of Berberine (an ancient Chinese drug used for antimicrobial, antiplasmodial, antidiarrheal and cardiovascular) on the solution structure of HSA using fluorescence, Fourier transform infrared (FT-IR), circular dichroism (CD) spectroscopic methods. The fluorescence spectroscopic results show that the fluorescence intensity of HSA was significantly decreased in the presence of Berberine. The Scatchard's plots indicated that the binding of Berberine to HSA at 296, 303, 318 K is characterized by one binding site with the binding constant is 4.071(±0.128)×10 4, 3.741(±0.089)×10 4, 3.454(±0.110)×10 4 M -1, respectively. The protein conformation is altered (FT-IR and CD data) with reductions of α-helices from 54 to 47% for free HSA to 45-32% and with increases of turn structure5% for free HSA to 18% in the presence of Berberine. The binding process was exothermic, enthalpy driven and spontaneous, as indicated by the thermodynamic analyses, Berberine bound to HSA was mainly based on hydrophobic interaction and electrostatic interaction cannot be excluded from the binding. Furthermore, the displace experiments indicate that Berberine can bind to the subdomain IIA, that is, high affinity site (site II).

A Review on Structures and Functions of Bcl-2 Family Proteins from Homo sapiens.

PubMed

Sivakumar, Dakshinamurthy; Sivaraman, Thirunavukkarasu

2016-01-01

Cancer cells evade apoptosis, which is regulated by proteins of Bcl-2 family in the intrinsic pathways. Numerous experimental three-dimensional (3D) structures of the apoptotic proteins and the proteins bound with small chemical molecules/peptides/proteins have been reported in the literature. In this review article, the 3D structures of the Bcl-2 family proteins from Homo sapiens and as well complex structures of the anti-apoptotic proteins bound with small molecular inhibitors reported in the literature to date have been comprehensively listed out and described in detail. Moreover, the molecular mechanisms by which the Bcl-2 family proteins modulate the apoptotic processes and strategies for designing antagonists to anti-apoptotic proteins have been concisely discussed.

CsrA Participates in a PNPase Autoregulatory Mechanism by Selectively Repressing Translation of pnp Transcripts That Have Been Previously Processed by RNase III and PNPase

PubMed Central

Park, Hongmarn; Yakhnin, Helen; Connolly, Michael; Romeo, Tony

2015-01-01

ABSTRACT Csr is a conserved global regulatory system that represses or activates gene expression posttranscriptionally. CsrA of Escherichia coli is a homodimeric RNA binding protein that regulates transcription elongation, translation initiation, and mRNA stability by binding to the 5′ untranslated leader or initial coding sequence of target transcripts. pnp mRNA, encoding the 3′ to 5′ exoribonuclease polynucleotide phosphorylase (PNPase), was previously identified as a CsrA target by transcriptome sequencing (RNA-seq). Previous studies also showed that RNase III and PNPase participate in a pnp autoregulatory mechanism in which RNase III cleavage of the untranslated leader, followed by PNPase degradation of the resulting 5′ fragment, leads to pnp repression by an undefined translational repression mechanism. Here we demonstrate that CsrA binds to two sites in pnp leader RNA but only after the transcript is fully processed by RNase III and PNPase. In the absence of processing, both of the binding sites are sequestered in an RNA secondary structure, which prevents CsrA binding. The CsrA dimer bridges the upstream high-affinity site to the downstream site that overlaps the pnp Shine-Dalgarno sequence such that bound CsrA causes strong repression of pnp translation. CsrA-mediated translational repression also leads to a small increase in the pnp mRNA decay rate. Although CsrA has been shown to regulate translation and mRNA stability of numerous genes in a variety of organisms, this is the first example in which prior mRNA processing is required for CsrA-mediated regulation. IMPORTANCE CsrA protein represses translation of numerous mRNA targets, typically by binding to multiple sites in the untranslated leader region preceding the coding sequence. We found that CsrA represses translation of pnp by binding to two sites in the pnp leader transcript but only after it is processed by RNase III and PNPase. Processing by these two ribonucleases alters the mRNA secondary structure such that it becomes accessible to the ribosome for translation as well as to CsrA. As one of the CsrA binding sites overlaps the pnp ribosome binding site, bound CsrA prevents ribosome binding. This is the first example in which regulation by CsrA requires prior mRNA processing and should link pnp expression to conditions affecting CsrA activity. PMID:26438818

A structure-based catalytic mechanism for the xanthine oxidase family of molybdenum enzymes.

PubMed Central

Huber, R; Hof, P; Duarte, R O; Moura, J J; Moura, I; Liu, M Y; LeGall, J; Hille, R; Archer, M; Romão, M J

1996-01-01

The crystal structure of the xanthine oxidase-related molybdenum-iron protein aldehyde oxido-reductase from the sulfate reducing anaerobic Gram-negative bacterium Desulfovibrio gigas (Mop) was analyzed in its desulfo-, sulfo-, oxidized, reduced, and alcohol-bound forms at 1.8-A resolution. In the sulfo-form the molybdenum molybdopterin cytosine dinucleotide cofactor has a dithiolene-bound fac-[Mo, = O, = S, ---(OH2)] substructure. Bound inhibitory isopropanol in the inner compartment of the substrate binding tunnel is a model for the Michaelis complex of the reaction with aldehydes (H-C = O,-R). The reaction is proposed to proceed by transfer of the molybdenum-bound water molecule as OH- after proton transfer to Glu-869 to the carbonyl carbon of the substrate in concert with hydride transfer to the sulfido group to generate [MoIV, = O, -SH, ---(O-C = O, -R)). Dissociation of the carboxylic acid product may be facilitated by transient binding of Glu-869 to the molybdenum. The metal-bound water is replenished from a chain of internal water molecules. A second alcohol binding site in the spacious outer compartment may cause the strong substrate inhibition observed. This compartment is the putative binding site of large inhibitors of xanthine oxidase. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 Fig. 6 PMID:8799115

RNAstructure: software for RNA secondary structure prediction and analysis.

PubMed

Reuter, Jessica S; Mathews, David H

2010-03-15

To understand an RNA sequence's mechanism of action, the structure must be known. Furthermore, target RNA structure is an important consideration in the design of small interfering RNAs and antisense DNA oligonucleotides. RNA secondary structure prediction, using thermodynamics, can be used to develop hypotheses about the structure of an RNA sequence. RNAstructure is a software package for RNA secondary structure prediction and analysis. It uses thermodynamics and utilizes the most recent set of nearest neighbor parameters from the Turner group. It includes methods for secondary structure prediction (using several algorithms), prediction of base pair probabilities, bimolecular structure prediction, and prediction of a structure common to two sequences. This contribution describes new extensions to the package, including a library of C++ classes for incorporation into other programs, a user-friendly graphical user interface written in JAVA, and new Unix-style text interfaces. The original graphical user interface for Microsoft Windows is still maintained. The extensions to RNAstructure serve to make RNA secondary structure prediction user-friendly. The package is available for download from the Mathews lab homepage at http://rna.urmc.rochester.edu/RNAstructure.html.

Study on the Effect of Secondary Banded Structure on the Fatigue Property of Non-Quenched and Tempered Micro Alloyed Steel

NASA Astrophysics Data System (ADS)

Yajie, Cheng; Qingliang, Liao; Yue, Zhang

Due to composition segregation and cooling speed, streamline or banded structure were often obtained in the thermal forming parts along the direction of parts forming. Generally speaking, banded structure doesn't decrease the longitudinal mechanical properties, so the secondary banded structure can't get enough attention. The effect of secondary banded structure on the fatigue properties of micro alloyed DG20Mn and 35CrMo steel was investigated using the axial tensile fatigue test of stress ratio of 0.1. The result shows that secondary banded structure was obtained in the center of the steel parts, because of the composition segregation and the lower cooling rate in center part of steel. Secondary banded structure has no significant effect on axial tensile properties of both DG20Mn and 35CrMo, but decreases the axial tensile fatigue performance of DG20Mn steel. This study suggests that under the high cyclic tensile stress, multi-source damage cracks in steel initiated by large strain of pearlite of secondary banded structure, which is larger than damage strain, is the major factor of the decrease of fatigue life of steel.

Internal structure of acceptor-bound excitons in wide-band-gap wurtzite semiconductors

NASA Astrophysics Data System (ADS)

Gil, Bernard; Bigenwald, Pierre; Paskov, Plamen P.; Monemar, Bo

2010-02-01

We describe the internal structure of acceptor-bound excitons in wurtzite semiconductors. Our approach consists in first constructing, in the context of angular momentum algebra, the wave functions of the two-hole system that fulfill Pauli’s exclusion’s principle. Second, we construct the acceptor-bound exciton states by adding the electron states in a similar manner that two-hole states are constructed. We discuss the optical selection rules for the acceptor-bound exciton recombination. Finally, we compare our theory with experimental data for CdS and GaN. In the specific case of CdS for which much experimental information is available, we demonstrate that, compared with cubic semiconductors, the sign of the short-range hole-exchange interaction is reversed and more than one order of magnitude larger. The whole set of data is interpreted in the context of a large value of the short-range hole-exchange interaction Ξ0=3.4±0.2meV . This value dictates the splitting between the ground-state line I1 and the other transitions. The values we find for the electron-hole spin-exchange interaction and of the crystal-field splitting of the two-hole state are, respectively, -0.4±0.1 and 0.2±0.1meV . In the case of GaN, the experimental data for the acceptor-bound excitons in the case of Mg and Zn acceptors, show more than one bound-exciton line. We discuss a possible assignment of these states.

Structure of the Balmer jump. The isolated hydrogen atom

NASA Astrophysics Data System (ADS)

Calvo, F.; Belluzzi, L.; Steiner, O.

2018-06-01

Context. The spectrum of the hydrogen atom was explained by Bohr more than one century ago. We revisit here some of the aspects of the underlying quantum structure, with a modern formalism, focusing on the limit of the Balmer series. Aims: We investigate the behaviour of the absorption coefficient of the isolated hydrogen atom in the neighbourhood of the Balmer limit. Methods: We analytically computed the total cross-section arising from bound-bound and bound-free transitions in the isolated hydrogen atom at the Balmer limit, and established a simplified semi-analytical model for the surroundings of that limit. We worked within the framework of the formalism of Landi Degl'Innocenti & Landolfi (2004, Astrophys. Space Sci. Lib., 307), which permits an almost straight-forward generalization of our results to other atoms and molecules, and which is perfectly suitable for including polarization phenomena in the problem. Results: We analytically show that there is no discontinuity at the Balmer limit, even though the concept of a "Balmer jump" is still meaningful. Furthermore, we give a possible definition of the location of the Balmer jump, and we check that this location is dependent on the broadening mechanisms. At the Balmer limit, we compute the cross-section in a fully analytical way. Conclusions: The Balmer jump is produced by a rapid drop of the total Balmer cross-section, yet this variation is smooth and continuous when both bound-bound and bound-free processes are taken into account, and its shape and location is dependent on the broadening mechanisms.

DNA Secondary Structure at Chromosomal Fragile Sites in Human Disease

PubMed Central

Thys, Ryan G; Lehman, Christine E; Pierce, Levi C. T; Wang, Yuh-Hwa

2015-01-01

DNA has the ability to form a variety of secondary structures that can interfere with normal cellular processes, and many of these structures have been associated with neurological diseases and cancer. Secondary structure-forming sequences are often found at chromosomal fragile sites, which are hotspots for sister chromatid exchange, chromosomal translocations, and deletions. Structures formed at fragile sites can lead to instability by disrupting normal cellular processes such as DNA replication and transcription. The instability caused by disruption of replication and transcription can lead to DNA breakage, resulting in gene rearrangements and deletions that cause disease. In this review, we discuss the role of DNA secondary structure at fragile sites in human disease. PMID:25937814

Structural dynamics of Casein Kinase I (CKI) from malarial parasite Plasmodium falciparum (Isolate 3D7): Insights from theoretical modelling and molecular simulations.

PubMed

Dehury, Budheswar; Behera, Santosh Kumar; Mahapatra, Namita

2017-01-01

The protein kinases (PKs), belonging to serine/threonine kinase (STKs), are important drug targets for a wide spectrum of diseases in human. Among protein kinases, the Casein Kinases (CKs) are vastly expanded in various organisms, where, the malarial parasite Plasmodium falciparum possesses a single member i.e., PfCKI, which can phosphorylate various proteins in parasite extracts in vitro condition. But, the structure-function relationship of PfCKI and dynamics of ATP binding is yet to be understood. Henceforth, an attempt was made to study the dynamics, stability, and ATP binding mechanisms of PfCKI through computational modelling, docking, molecular dynamics (MD) simulations, and MM/PBSA binding free energy estimation. Bi-lobed catalytic domain of PfCKI shares a high degree of secondary structure topology with CKI domains of rice, human, and mouse indicating co-evolution of these kinases. Molecular docking study revealed that ATP binds to the active site where the glycine-rich ATP-binding motif (G16-X-G18-X-X-G21) along with few conserved residues plays a crucial role maintaining stability of the complex. Structural superposition of PfCKI with close structural homologs depicted that the location and length of important loops are different, indicating the dynamic properties of these loops among CKIs, which is consistent with principal component analysis (PCA). PCA displayed that the overall global motion of ATP-bound form is comparatively higher than that of apo form. The present study provides insights into the structural features of PfCKI, which could contribute towards further understanding of related protein structures, dynamics of catalysis and phosphorylation mechanism in these important STKs from malarial parasite in near future. Copyright © 2016 Elsevier Inc. All rights reserved.

Identification of structural domains in proteins by a graph heuristic.

PubMed

Wernisch, L; Hunting, M; Wodak, S J

1999-05-15

A novel automatic procedure for identifying domains from protein atomic coordinates is presented. The procedure, termed STRUDL (STRUctural Domain Limits), does not take into account information on secondary structures and handles any number of domains made up of contiguous or non-contiguous chain segments. The core algorithm uses the Kernighan-Lin graph heuristic to partition the protein into residue sets which display minimum interactions between them. These interactions are deduced from the weighted Voronoi diagram. The generated partitions are accepted or rejected on the basis of optimized criteria, representing basic expected physical properties of structural domains. The graph heuristic approach is shown to be very effective, it approximates closely the exact solution provided by a branch and bound algorithm for a number of test proteins. In addition, the overall performance of STRUDL is assessed on a set of 787 representative proteins from the Protein Data Bank by comparison to domain definitions in the CATH protein classification. The domains assigned by STRUDL agree with the CATH assignments in at least 81% of the tested proteins. This result is comparable to that obtained previously using PUU (Holm and Sander, Proteins 1994;9:256-268), the only other available algorithm designed to identify domains with any number of non-contiguous chain segments. A detailed discussion of the structures for which our assignments differ from those in CATH brings to light some clear inconsistencies between the concept of structural domains based on minimizing inter-domain interactions and that of delimiting structural motifs that represent acceptable folding topologies or architectures. Considering both concepts as complementary and combining them in a layered approach might be the way forward.

Dynamics of a New 5D Hyperchaotic System of Lorenz Type

NASA Astrophysics Data System (ADS)

Zhang, Fuchen; Chen, Rui; Wang, Xingyuan; Chen, Xiusu; Mu, Chunlai; Liao, Xiaofeng

Ultimate boundedness of chaotic dynamical systems is one of the fundamental concepts in dynamical systems, which plays an important role in investigating the stability of the equilibrium, estimating the Lyapunov dimension of attractors and the Hausdorff dimension of attractors, the existence of periodic solutions, chaos control, chaos synchronization. However, it is often difficult to obtain the bounds of the hyperchaotic systems due to the complex algebraic structure of the hyperchaotic systems. This paper has investigated the boundedness of solutions of a nonlinear hyperchaotic system. We have obtained the global exponential attractive set and the ultimate bound set for this system. To obtain the ellipsoidal ultimate bound, the ultimate bound of the proposed system is theoretically estimated using Lagrange multiplier method, Lyapunov stability theory and optimization theory. To show the ultimate bound region, numerical simulations are provided.

Do All X-ray Structures of Protein-Ligand Complexes Represent Functional States? EPOR, a Case Study.

PubMed

Corbett, Michael S P; Mark, Alan E; Poger, David

2017-02-28

Based on differences between the x-ray crystal structures of ligand-bound and unbound forms, the activation of the erythropoietin receptor (EPOR) was initially proposed to involve a cross-action scissorlike motion. However, the validity of the motions involved in the scissorlike model has been recently challenged. Here, atomistic molecular dynamics simulations are used to examine the structure of the extracellular domain of the EPOR dimer in the presence and absence of erythropoietin and a series of agonistic or antagonistic mimetic peptides free in solution. The simulations suggest that in the absence of crystal packing effects, the EPOR chains in the different dimers adopt very similar conformations with no clear distinction between the agonist and antagonist-bound complexes. This questions whether the available x-ray crystal structures of EPOR truly represent active or inactive conformations. The study demonstrates the difficulty in using such structures to infer a mechanism of action, especially in the case of membrane receptors where just part of the structure has been considered in addition to potential confounding effects that arise from the comparison of structures in a crystal as opposed to a membrane environment. The work highlights the danger of assigning functional significance to small differences between structures of proteins bound to different ligands in a crystal environment without consideration of the effects of the crystal lattice and thermal motion. Copyright © 2017 Biophysical Society. Published by Elsevier Inc. All rights reserved.

Cloning, characterisation and comparative analysis of a starch synthase IV gene in wheat: functional and evolutionary implications

PubMed Central

Leterrier, Marina; Holappa, Lynn D; Broglie, Karen E; Beckles, Diane M

2008-01-01

Background Starch is of great importance to humans as a food and biomaterial, and the amount and structure of starch made in plants is determined in part by starch synthase (SS) activity. Five SS isoforms, SSI, II, III, IV and Granule Bound SSI, have been identified, each with a unique catalytic role in starch synthesis. The basic mode of action of SSs is known; however our knowledge of several aspects of SS enzymology at the structural and mechanistic level is incomplete. To gain a better understanding of the differences in SS sequences that underscore their specificity, the previously uncharacterised SSIVb from wheat was cloned and extensive bioinformatics analyses of this and other SSs sequences were done. Results The wheat SSIV cDNA is most similar to rice SSIVb with which it shows synteny and shares a similar exon-intron arrangement. The wheat SSIVb gene was preferentially expressed in leaf and was not regulated by a circadian clock. Phylogenetic analysis showed that in plants, SSIV is closely related to SSIII, while SSI, SSII and Granule Bound SSI clustered together and distinctions between the two groups can be made at the genetic level and included chromosomal location and intron conservation. Further, identified differences at the amino acid level in their glycosyltransferase domains, predicted secondary structures, global conformations and conserved residues might be indicative of intragroup functional associations. Conclusion Based on bioinformatics analysis of the catalytic region of 36 SSs and 3 glycogen synthases (GSs), it is suggested that the valine residue in the highly conserved K-X-G-G-L motif in SSIII and SSIV may be a determining feature of primer specificity of these SSs as compared to GBSSI, SSI and SSII. In GBSSI, the Ile485 residue may partially explain that enzyme's unique catalytic features. The flexible 380s Loop in the starch catalytic domain may be important in defining the specificity of action for each different SS and the G-X-G in motif VI could define SSIV and SSIII action particularly. PMID:18826586

Fine tuning of coenzyme specificity in family 2 aldo-keto reductases revealed by crystal structures of the Lys-274 → Arg mutant of Candida tenuis xylose reductase (AKR2B5) bound to NAD + and NADP +

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

Leitgeb, Stefan; Petschacher, Barbara; Wilson, David K.

2005-01-11

Aldo-keto reductases of family 2 employ single site replacement Lys → Arg to switch their cosubstrate preference from NADPH to NADH. X-ray crystal structures of Lys-274 → Arg mutant of Candida tenuis xylose reductase (AKR2B5) bound to NAD + and NADP + were determined at a resolution of 2.4 and 2.3 Å, respectively. Due to steric conflicts in the NADP +-bound form, the arginine side chain must rotate away from the position of the original lysine side chain, thereby disrupting a network of direct and water-mediated interactions between Glu-227, Lys-274 and the cofactor 2'-phosphate and 3'-hydroxy groups. Because anchoring contactsmore » of its Glu-227 are lost, the coenzyme-enfolding loop that becomes ordered upon binding of NAD(P) + in the wild-type remains partly disordered in the NADP +-bound mutant. The results delineate a catalytic reaction profile for the mutant in comparison to wild-type.« less

Resolving the Spatial Structures of Bound Hole States in Black Phosphorus.

PubMed

Qiu, Zhizhan; Fang, Hanyan; Carvalho, Alexandra; Rodin, A S; Liu, Yanpeng; Tan, Sherman J R; Telychko, Mykola; Lv, Pin; Su, Jie; Wang, Yewu; Castro Neto, A H; Lu, Jiong

2017-11-08

Understanding the local electronic properties of individual defects and dopants in black phosphorus (BP) is of great importance for both fundamental research and technological applications. Here, we employ low-temperature scanning tunnelling microscope (LT-STM) to probe the local electronic structures of single acceptors in BP. We demonstrate that the charge state of individual acceptors can be reversibly switched by controlling the tip-induced band bending. In addition, acceptor-related resonance features in the tunnelling spectra can be attributed to the formation of Rydberg-like bound hole states. The spatial mapping of the quantum bound states shows two distinct shapes evolving from an extended ellipse shape for the 1s ground state to a dumbbell shape for the 2p x excited state. The wave functions of bound hole states can be well-described using the hydrogen-like model with anisotropic effective mass, corroborated by our theoretical calculations. Our findings not only provide new insight into the many-body interactions around single dopants in this anisotropic two-dimensional material but also pave the way to the design of novel quantum devices.

Internalization of exogenous ADP-ribosylation factor 6 (Arf6) proteins into cells.

PubMed

Afroze, Syeda H; Uddin, M Nasir; Cao, Xiaobo; Asea, Alexzander; Gizachew, Dawit

2011-08-01

Endogenous Arf6 is a myristoylated protein mainly involved in endosomal membrane traffic and structural organization at the plasma membrane. It has been shown that Arf6 mediates cancer cell invasion and shedding of plasma membrane microvesicles derived from tumor cells. In this article, we determined that Arf6 proteins both in the GDP and GTPγS bound forms can enter cells when simply added in the cell culture medium without requiring the myristoyl group. The GTPγS bound can enter cells at a faster rate than the GDP-bound Arf6. Despite the role of the endogenous Arf6 in endocytosis and membrane trafficking, the internalization of exogenous Arf6 may involve non-endocytic processes. As protein therapeutics is becoming important in medicine, we examined the effect of the uptake of Arf6 proteins on cellular functions and determined that exogenous Arf6 inhibits proliferation, invasion, and migration of cells. Future studies of the internalization of Arf6 mutants will reveal key residues that play a role in the internalization of Arf6 and its interaction and possible structural conformations bound to the plasma membrane.

Protein secondary structure prediction using modular reciprocal bidirectional recurrent neural networks.

PubMed

Babaei, Sepideh; Geranmayeh, Amir; Seyyedsalehi, Seyyed Ali

2010-12-01

The supervised learning of recurrent neural networks well-suited for prediction of protein secondary structures from the underlying amino acids sequence is studied. Modular reciprocal recurrent neural networks (MRR-NN) are proposed to model the strong correlations between adjacent secondary structure elements. Besides, a multilayer bidirectional recurrent neural network (MBR-NN) is introduced to capture the long-range intramolecular interactions between amino acids in formation of the secondary structure. The final modular prediction system is devised based on the interactive integration of the MRR-NN and the MBR-NN structures to arbitrarily engage the neighboring effects of the secondary structure types concurrent with memorizing the sequential dependencies of amino acids along the protein chain. The advanced combined network augments the percentage accuracy (Q₃) to 79.36% and boosts the segment overlap (SOV) up to 70.09% when tested on the PSIPRED dataset in three-fold cross-validation. Copyright © 2010 Elsevier Ireland Ltd. All rights reserved.

New insights from cluster analysis methods for RNA secondary structure prediction

PubMed Central

Rogers, Emily; Heitsch, Christine

2016-01-01

A widening gap exists between the best practices for RNA secondary structure prediction developed by computational researchers and the methods used in practice by experimentalists. Minimum free energy (MFE) predictions, although broadly used, are outperformed by methods which sample from the Boltzmann distribution and data mine the results. In particular, moving beyond the single structure prediction paradigm yields substantial gains in accuracy. Furthermore, the largest improvements in accuracy and precision come from viewing secondary structures not at the base pair level but at lower granularity/higher abstraction. This suggests that random errors affecting precision and systematic ones affecting accuracy are both reduced by this “fuzzier” view of secondary structures. Thus experimentalists who are willing to adopt a more rigorous, multilayered approach to secondary structure prediction by iterating through these levels of granularity will be much better able to capture fundamental aspects of RNA base pairing. PMID:26971529

Assessing the Effect of Stellar Companions from High-resolution Imaging of Kepler Objects of Interest

NASA Astrophysics Data System (ADS)

Hirsch, Lea A.; Ciardi, David R.; Howard, Andrew W.; Everett, Mark E.; Furlan, Elise; Saylors, Mindy; Horch, Elliott P.; Howell, Steve B.; Teske, Johanna; Marcy, Geoffrey W.

2017-03-01

We report on 176 close (<2″) stellar companions detected with high-resolution imaging near 170 hosts of Kepler Objects of Interest (KOIs). These Kepler targets were prioritized for imaging follow-up based on the presence of small planets, so most of the KOIs in these systems (176 out of 204) have nominal radii <6 {R}\\oplus . Each KOI in our sample was observed in at least two filters with adaptive optics, speckle imaging, lucky imaging, or the Hubble Space Telescope. Multi-filter photometry provides color information on the companions, allowing us to constrain their stellar properties and assess the probability that the companions are physically bound. We find that 60%-80% of companions within 1″ are bound, and the bound fraction is >90% for companions within 0.″5 the bound fraction decreases with increasing angular separation. This picture is consistent with simulations of the binary and background stellar populations in the Kepler field. We also reassess the planet radii in these systems, converting the observed differential magnitudes to a contamination in the Kepler bandpass and calculating the planet radius correction factor, X R = R p (true)/R p (single). Under the assumption that planets in bound binaries are equally likely to orbit the primary or secondary, we find a mean radius correction factor for planets in stellar multiples of X R = 1.65. If stellar multiplicity in the Kepler field is similar to the solar neighborhood, then nearly half of all Kepler planets may have radii underestimated by an average of 65%, unless vetted using high-resolution imaging or spectroscopy.

Airborne particle-bound brominated flame retardants: Levels, size distribution and indoor-outdoor exchange.

PubMed

Zhu, Yue-Shan; Yang, Wan-Dong; Li, Xiu-Wen; Ni, Hong-Gang; Zeng, Hui

2018-02-01

The quality of indoor environments has a significant impact on public health. Usually, an indoor environment is treated as a static box, in which physicochemical reactions of indoor air contaminants are negligible. This results in conservative estimates for primary indoor air pollutant concentrations, while also ignoring secondary pollutants. Thus, understanding the relationship between indoor and outdoor particles and particle-bound pollutants is of great significance. For this reason, we collected simultaneous indoor and outdoor measurements of the size distribution of airborne brominated flame retardant (BFR) congeners. The time-dependent concentrations of indoor particles and particle-bound BFRs were then estimated with the mass balance model, accounting for the outdoor concentration, indoor source strength, infiltration, penetration, deposition and indoor resuspension. Based on qualitative observation, the size distributions of ΣPBDE and ΣHBCD were characterized by bimodal peaks. According to our results, particle-bound BDE209 and γ-HBCD underwent degradation. Regardless of the surface adsorption capability of particles and the physicochemical properties of the target compounds, the concentration of BFRs in particles of different size fractions seemed to be governed by the particle distribution. Based on our estimations, for airborne particles and particle-bound BFRs, a window-open ventilated room only takes a quarter of the time to reach an equilibrium between the concentration of pollutants inside and outside compared to a closed room. Unfortunately, indoor pollutants and outdoor pollutants always exist simultaneously, which poses a window-open-or-closed dilemma to achieve proper ventilation. Copyright © 2017 Elsevier Ltd. All rights reserved.

Bounds on isocurvature perturbations from cosmic microwave background and large scale structure data.

PubMed

Crotty, Patrick; García-Bellido, Juan; Lesgourgues, Julien; Riazuelo, Alain

2003-10-24

We obtain very stringent bounds on the possible cold dark matter, baryon, and neutrino isocurvature contributions to the primordial fluctuations in the Universe, using recent cosmic microwave background and large scale structure data. Neglecting the possible effects of spatial curvature, tensor perturbations, and reionization, we perform a Bayesian likelihood analysis with nine free parameters, and find that the amplitude of the isocurvature component cannot be larger than about 31% for the cold dark matter mode, 91% for the baryon mode, 76% for the neutrino density mode, and 60% for the neutrino velocity mode, at 2sigma, for uncorrelated models. For correlated adiabatic and isocurvature components, the fraction could be slightly larger. However, the cross-correlation coefficient is strongly constrained, and maximally correlated/anticorrelated models are disfavored. This puts strong bounds on the curvaton model.

Cd(2+) Triggered the FRET "ON": A New Molecular Switch for the Ratiometric Detection of Cd(2+) with Live-Cell Imaging and Bound X-ray Structure.

PubMed

Aich, Krishnendu; Goswami, Shyamaprosad; Das, Sangita; Mukhopadhyay, Chitrangada Das; Quah, Ching Kheng; Fun, Hoong-Kun

2015-08-03

On the basis of the Förster resonance energy transfer mechanism between rhodamine and quinoline-benzothiazole conjugated dyad, a new colorimetric as well as fluorescence ratiometric probe was synthesized for the selective detection of Cd(2+). The complex formation of the probe with Cd(2+) was confirmed through Cd(2+)-bound single-crystal structure. Capability of the probe as imaging agent to detect the cellular uptake of Cd(2+) was demonstrated here using living RAW cells.

Low pressure-induced secondary structure transitions of regenerated silk fibroin in its wet film studied by time-resolved infrared spectroscopy.

PubMed

He, Zhipeng; Liu, Zhao; Zhou, Xiaofeng; Huang, He

2018-06-01

The secondary structure transitions of regenerated silk fibroin (RSF) under different external perturbations have been studied extensively, except for pressure. In this work, time-resolved infrared spectroscopy with the attenuated total reflectance (ATR) accessory was employed to follow the secondary structure transitions of RSF in its wet film under low pressure. It has been found that pressure alone is favorable only to the formation of β-sheet structure. Under constant pressure there is an optimum amount of D 2 O in the wet film (D 2 O : film = 2:1) so as to provide the optimal condition for the reorganization of the secondary structure and to have the largest formation of β-sheet structure. Under constant amount of D 2 O and constant pressure, the secondary structure transitions of RSF in its wet film can be divided into three stages along with time. In the first stage, random coil, α-helix, and β-turn were quickly transformed into β-sheet. In the second stage, random coil and β-turn were relatively slowly transformed into β-sheet and α-helix, and the content of α-helix was recovered to the value prior to the application of pressure. In the third and final stage, no measurable changes can be found for each secondary structure. This study may be helpful to understand the secondary structure changes of silk fibroin in silkworm's glands under hydrostatic pressure. © 2018 Wiley Periodicals, Inc.

Structures of a Nonribosomal Peptide Synthetase Module Bound to MbtH-like Proteins Support a Highly Dynamic Domain Architecture

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

Miller, Bradley R.; Drake, Eric J.; Shi, Ce

Nonribosomal peptide synthetases (NRPSs) produce a wide variety of peptide natural products. During synthesis, the multidomain NRPSs act as an assembly line, passing the growing product from one module to the next. Each module generally consists of an integrated peptidyl carrier protein, an amino acid-loading adenylation domain, and a condensation domain that catalyzes peptide bond formation. Some adenylation domains interact with small partner proteins called MbtH-like proteins (MLPs) that enhance solubility or activity. A structure of an MLP bound to an adenylation domain has been previously reported using a truncated adenylation domain, precluding any insight that might be derived frommore » understanding the influence of the MLP on the intact adenylation domain or on the dynamics of the entire NRPS module. Here, we present the structures of the full-length NRPS EntF bound to the MLPs from Escherichia coli and Pseudomonas aeruginosa. These new structures, along with biochemical and bioinformatics support, further elaborate the residues that define the MLP-adenylation domain interface. Additionally, the structures highlight the dynamic behavior of NRPS modules, including the module core formed by the adenylation and condensation domains as well as the orientation of the mobile thioesterase domain.« less