Molecular interferometric imaging study of molecular interactions

NASA Astrophysics Data System (ADS)

Zhao, Ming; Wang, Xuefeng; Nolte, David

2008-02-01

Molecular Interferometric Imaging (MI2) is a sensitive detection platform for direct optical detection of immobilized biomolecules. It is based on inline common-path interferometry combined with far-field optical imaging. The substrate is a simple thermal oxide on a silicon surface with a thickness at or near the quadrature condition that produces a π/2 phase shift between the normal-incident wave reflected from the top oxide surface and the bottom silicon surface. The presence of immobilized or bound biomolecules on the surface produces a relative phase shift that is converted to a far-field intensity shift and is imaged by a reflective microscope onto a CCD camera. Shearing interferometry is used to remove the spatial 1/f noise from the illumination to achieve shot-noise-limited detection of surface dipole density profiles. The lateral resolution of this technique is diffraction limited at 0.4 micron, and the best longitudinal resolution is 10 picometers. The minimum detectable mass at the metrology limit is 2 attogram, which is 8 antibody molecules of size 150 kDa. The corresponding scaling mass sensitivity is 5 fg/mm compared with 1 pg/mm for typical SPR sensitivity. We have applied MI2 to immunoassay applications, and real-time binding kinetics has been measured for antibody-antigen reactions. The simplicity of the substrate and optical read-out make MI2 a promising analytical assay tool for high-throughput screening and diagnostics.

Dopamine and executive function: Increased spontaneous eye blink rates correlate with better set-shifting and inhibition, but poorer updating.

PubMed

Zhang, Ting; Mou, Di; Wang, Cuicui; Tan, Fengping; Jiang, Yan; Lijun, Zheng; Li, Hong

2015-06-01

The central dopamine system (DA) has a significant role in the executive function (EF). The spontaneous eye blink rate (EBR) is an effective clinical and non-invasive measure, which is strongly related to the activity of the central dopaminergic system. Previous studies show significant relationships between the two main dimensions of EF (i.e., shifting and inhibition) and the central DA system as measured by EBR. However, most of these studies involve only one EF task for shifting or inhibition; whether or not these relationships are replicated by other EF tasks remains unclear. Besides, the relationship between EBR and another important EF dimension-updating-also remains unknown. The present study examined the correlation between EBR and several EF tasks that captured all the three EF dimensions: shifting, inhibition, and updating. A total of 61 healthy participants were subjected to EBR testing and EF tasks. Results showed that EBR had a different relationship with each of the three tested EF dimensions. An increase in EBR levels was related to an increase in accuracy in shifting and inhibition tasks, a decrease in shifting and inhibition cost, and a decrease in accuracy in updating tasks. These results imply that the role of the central DA system in shifting and inhibition differs from its role in updating. Copyright © 2015 Elsevier B.V. All rights reserved.

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

EL-Shamy, E. F., E-mail: emadel-shamy@hotmail.com; Department of Physics, College of Science, King Khalid University, P.O. 9004, Abha; Al-Asbali, A. M., E-mail: aliaa-ma@hotmail.com

A theoretical investigation is carried out to study the propagation and the head-on collision of dust-acoustic (DA) shock waves in a strongly coupled dusty plasma consisting of negative dust fluid, Maxwellian distributed electrons and ions. Applying the extended Poincaré–Lighthill–Kuo method, a couple of Korteweg–deVries–Burgers equations for describing DA shock waves are derived. This study is a first attempt to deduce the analytical phase shifts of DA shock waves after collision. The impacts of physical parameters such as the kinematic viscosity, the unperturbed electron-to-dust density ratio, parameter determining the effect of polarization force, the ion-to-electron temperature ratio, and the effective dustmore » temperature-to-ion temperature ratio on the structure and the collision of DA shock waves are examined. In addition, the results reveal the increase of the strength and the steepness of DA shock waves as the above mentioned parameters increase, which in turn leads to the increase of the phase shifts of DA shock waves after collision. The present model may be useful to describe the structure and the collision of DA shock waves in space and laboratory dusty plasmas.« less

Thermal Degradation of Synthetic Cathinones: Implications for Forensic Toxicology.

PubMed

Kerrigan, Sarah; Savage, Megan; Cavazos, Cassandra; Bella, Paige

2016-01-01

The synthetic cathinones represent an important class of designer drugs. The widespread attention and publicity associated with these psychostimulants have resulted in numerous legislative actions at state and federal levels throughout the USA. These amphetamine-like compounds are characterized by a β-keto functional group. Although the synthetic cathinones share many properties of their phenethylamine counterparts, the presence of the ketone moiety is responsible for a number of unique and distinct differences in terms of their chemical characteristics and properties. Thermal degradation of methcathinone was first reported several decades ago but has received limited attention. In this study, we identified in situ thermal degradation products for 18 cathinones during gas chromatography-mass spectrometry (GC-MS) analysis. Oxidative degradation arises from the loss of two hydrogens, yielding a characteristic 2 Da mass shift. Degradation products were characterized by prominent iminium base peaks with mass-to-charge ratios 2 Da lower than the parent drug, and in the case of the pyrrolidine-containing cathinones, prominent molecular ions arising from the 2,3-enamine. Chromatographic and mass spectroscopic data are described for 4-ethylmethcathinone, 4-methylethcathinone, buphedrone, butylone, ethcathinone, ethylone, flephedrone, 3,4-methylenedioxy-α-pyrrolidinobutiophenone, 3,4-methylenedioxypyrovalerone, mephedrone, methcathinone, methedrone, methylone, 4-methyl-α-pyrrolidinobutiophenone, naphyrone, pentedrone, pentylone and pyrovalerone. Degradation was minimized by lowering injection temperatures, residence time in the inlet and eliminating active sites during chromatographic analysis. Chromatographic and mass spectral data for the cathinone degradation products are presented and discussed within the context of forensic toxicological analysis, selection of appropriate instrumental methods and implications for the interpretation of results. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Dopamine depletion shifts behavior from activity based reinforcers to more sedentary ones and adenosine receptor antagonism reverses that shift: Relation to ventral striatum DARPP32 phosphorylation patterns.

PubMed

López-Cruz, Laura; San Miguel, Noemí; Carratalá-Ros, Carla; Monferrer, Lidón; Salamone, John D; Correa, Mercè

2018-02-02

The mesolimbic dopamine (DA) system plays a critical role in behavioral activation and effort-based decision-making. DA depletion produces anergia (shifts to low effort options) in animals tested on effort-based decision-making tasks. Caffeine, the most consumed stimulant in the world, acts as an adenosine A 1 /A 2A receptor antagonist, and in striatal areas DA D 1 and D 2 receptors are co-localized with adenosine A 1 and A 2A receptors respectively. In the present work, we evaluated the effect of caffeine on anergia induced by the VMAT-2 inhibitor tetrabenazine (TBZ), which depletes DA. Anergia was evaluated in a three-chamber T-maze task in which animals can chose between running on a wheel (RW) vs. sedentary activities such as consuming sucrose or sniffing a neutral odor. TBZ-caffeine interactions in ventral striatum were evaluated using DARPP-32 phosphorylation patterns as an intracellular marker of DA-adenosine receptor interaction. In the T-maze, control mice spent more time running and much less consuming sucrose or sniffing. TBZ (4.0 mg/kg) reduced ventral striatal DA tissue levels as measured by HPLC, and also shifted preferences in the T-maze, reducing selection of the reinforcer that involved vigorous activity (RW), but increasing consumption of a reinforcer that required little effort (sucrose), at doses that had no effect on independent measures of appetite or locomotion in a RW. Caffeine at doses that had no effect on their own reversed the effects of TBZ on T-maze performance, and also suppressed TBZ-induced pDARPP-32(Thr34) expression as measured by western blot, suggesting a role for D 2 -A 2A interactions. These results support the idea that DA depletion produces anergia, but does not affect the primary motivational effects of sucrose. Caffeine, possibly by acting on A 2A receptors in ventral striatum, reversed the DA depletion effects. It is possible that caffeine, like selective adenosine A2A antagonists, could have some therapeutic benefit for treating effort-related symptoms. Copyright © 2018. Published by Elsevier Ltd.

Metaproteomics of Microbiota in Naturally Fermented Soybean Paste, Da-jiang.

PubMed

Zhang, Ping; Zhang, Pengfei; Xie, Mengxi; An, Feiyu; Qiu, Boshu; Wu, Rina

2018-05-01

Da-jiang is a typical traditional fermented soybean product in China. At present, the proteins in da-jiang are needed to be explored. The composition and species of microbial proteins in traditional fermented da-jiang were analyzed by metaproteomics based on sodium dodecyl sulfonate-polyacrylamide gel electrophoresis (SDS-PAGE) and liquid chromatography-mass spectrometry/mass spectrometry (LC-MS/MS). The results showed that the number and variety of microbial proteins in the traditional fermented da-jiang from different regions were different. The production site influences the fermentation in da-jiang. Then we analyzed the functions of the microbial proteins identified in da-jiang, and found that they were mainly involved in the process of protein synthesis, glycometabolism and nucleic acid synthesis. In addtion, we compared the proteins composition in different da-jiang. There are 51 common proteins of naturally fermented da-jiang, and 25 common microbial sources. The main commonly microbial sources of fungal proteins are Saccharomyces cerevisiae and Schizosaccharomyces; the main commonly microbial sources of bacterial proteins are Enterococcus faecalis, Leuconostoc mesenteroides, Acinetobacter baumannii, and Bacillus subtilis. These common microbes play the predominant role in da-jiang fermentation. The present results help us to understand the fermentation of da-jiang and improve the quality and safety of final products in the future. The study illustrated metaproteome of microbiota in traditional fermented soybean paste, da-jiang, by sodium dodecyl sulfonate-polyacrylamide gel electrophoresis (SDS-PAGE) and liquid chromatography-mass spectrometry/mass spectrometry (LC-MS/MS). A method of extracting metaproteome from microbiota in da-jiang was attempted. The findings help to understand the fermentation of da-jiang and improve the quality and safety of da-jiang in fermented industry. © 2018 Institute of Food Technologists®.

Further insights into cortactin conformational regulation

PubMed Central

Evans, Jason V; Kelley, Laura C; Hayes, Karen E; Ammer, Amanda Gatesman; Martin, Karen H

2011-01-01

The actin regulatory protein cortactin is involved in multiple signaling pathways impinging on the cortical actin cytoskeleton. Cortactin is phosphorylated by ERK1/2 and Src family tyrosine kinases, resulting in neuronal Wiskott Aldrich Syndrome protein (N-WASp) activation and enhanced actin related protein (Arp)2/3-mediated actin nucleation. Cortactin migrates as an 80/85 kDa doublet when analyzed by SDS-PAGE. Phosphorylation by ERK1/2 is associated with conversion of the 80 kDa to the 85 kDa form, postulated to occur by inducing a conformational alteration that releases the carboxyl-terminal SH3 domain from autoinhibition. Our recent analysis of the 80–85 kDa cortactin “shift” in tumor cells indicates that while ERK1/2 phosphorylation is associated with the 85 kDa shift, this phosphorylation event is not required for the shift to occur, nor does ERK1/2 phosphorylation appreciably alter global cortactin confirmation. These data indicate that additional factors besides ERK1/2 phosphorylation contribute to generating and/or maintaining the activated 85 kDa cortactin form in stimulated cells. PMID:21866257

The White-Dwarf Mass-Radius Relation from 40 Eridani B and Other Nearby Visual Binaries

NASA Astrophysics Data System (ADS)

Bond, Howard E.; Bergeron, P.; Bedard, A.

2018-01-01

The bright, nearby DA-type white dwarf (WD) 40 Eridani B is orbited by the M dwarf 40 Eri C, allowing determination of the WD's mass. Until recently, however, the mass depended on orbital elements determined four decades ago, and that mass was so low that it created several astrophysical puzzles. Using new astrometric measurements, the binary-star group at the U.S. Naval Observatory has revised the dynamical mass upward, to 0.573 ± 0.018 M⊙. We have used model-atmosphere analysis to update other parameters of the WD, including effective temperature, surface gravity, radius, and luminosity. We then comparethese results with WD interior models.Within the observational uncertainties, theoretical cooling tracks for CO-core WDs of its measured mass are consistent with the position of 40 Eri B in the H-R diagram; equivalently, the theoretical mass-radius relation (MRR) is consistent with the star's location in the mass-radius plane. This consistency is, however, achieved only if we assume a "thin'' outer hydrogen layer, with qH = MH/MWD ∼ 10–10.We discuss other evidence that a significant fraction of DA WDs have such thin H layers, in spite of expectation from canonical stellar-evolution theory of "thick'' H layers with qH ∼ 10–4 . The cooling age of 40 Eri B is ~122 Myr, and its total age is ~1.8 Gyr. We present the MRRs for 40 Eri B and three other nearby WDs in visual binaries with precise mass determinations, and show that the agreement of current theory with observation is excellent in all cases.However, astrophysical puzzles remain. The eccentricity of the BC orbit has remained high (0.43), even though the progenitor of B ought to have interacted tidally with C when it was an AGB star. This puzzle exists also for the Sirius and Procyon systems. If thin hydrogen layers are common among WDs, the mass scale will need to be shifted downwards by a few hundredths of a solar mass.

Tonic dopamine induces persistent changes in the transient potassium current through translational regulation

PubMed Central

Rodgers, EW; Krenz, W-D; Baro, DJ

2012-01-01

Neuromodulatory effects can vary with their mode of transmission. Phasic release produces local and transient increases in dopamine (DA) up to micromolar concentrations. Additionally, since DA is released from open synapses and reuptake mechanisms are not nearby, tonic nanomolar DA exists in the extracellular space. Do phasic and tonic transmissions similarly regulate voltage dependent ionic conductances in a given neuron? It was previously shown that DA could immediately alter the transient potassium current (IA) of identified neurons in the stomatogastric ganglion (STG) of the spiny lobster, Panulirus interruptus. Here we show that DA can also persistently alter IA, and that DA’s immediate and persistent effects oppose one another. The lateral pyloric neuron (LP) exclusively expresses type 1 DA receptors (D1Rs). Micromolar DA produces immediate depolarizing shifts in the voltage dependence of LP IA, whereas tonic nanomolar DA produces a persistent increase in LP IA maximal conductance (Gmax) through a translation dependent mechanism involving target of rapamycin (TOR). The pyloric dilator neuron (PD) exclusively expresses type 2 DA receptors (D2Rs). Micromolar DA produces an immediate hyperpolarizing shift in PD IA voltage dependence of activation, whereas tonic DA persistently decreases PD IA Gmax through a translation dependent mechanism not involving TOR. The persistent effects on IA Gmax do not depend on LP or PD activity. These data suggest a role for tonic modulators in the regulation of voltage gated ion channel number; and furthermore, that dopaminergic systems may be organized to limit the amount of change they can impose on a circuit. PMID:21917788

Detection of Labile Low-Molecular-Mass Transition Metal Complexes in Mitochondria

PubMed Central

McCormick, Sean P.; Moore, Michael J.; Lindahl, Paul A.

2015-01-01

Liquid chromatography was used with an on-line inductively coupled plasma mass spectrometer to detect low-molecular-mass (LMM) transition metal complexes in mitochondria isolated from fermenting yeast cells, human Jurkat cells, and mouse brain and liver. These complexes constituted 20 – 40% of total mitochondrial Mn, Fe, Zn, and Cu ions. The major LMM Mn complex in yeast mitochondria had a mass of ca. 1100 Da and a concentration of ~ 2 μM. Mammalian mitochondria contained a second Mn species with a mass of ca. 2000 Da at a comparable concentration. The major Fe complex in mitochondria isolated from exponentially growing yeast cells had a mass of ca. 580 Da; the concentration of Fe580 in mitochondria was ca. 100 μM. When mitochondria were isolated from fermenting cells in post-exponential phase, the mass of the dominant LMM Fe complex was ca. 1100 Da. Upon incubation, the intensity of Fe1100 declined and Fe580 increased, suggesting that the two are interrelated. Mammalian mitochondria contained Fe580 and 2 other Fe species (Fe2000 and Fe1100) at concentrations of ca. 50 μM each. The dominant LMM Zn species in mitochondria had a mass of ca. 1200 Da and a concentration of ca. 110 μM. Mammalian mitochondria contained a second major LMM Zn species at 1500 Da. The dominant LMM Cu species in yeast mitochondria had a mass of ca. 5000 Da and a concentration in yeast mitochondria of ca. 16 μM; Cu5000 was not observed in mammalian mitochondria. The dominant Co species in mitochondria, Co1200, had a concentration of 20 nM and was probably a cobalamin. Mammalian but not yeast mitochondria contained a LMM Mo species, Mo730, at ca. 1 μM concentration. Increasing Mn, Fe, Cu, and Zn concentrations 10 fold in the medium increased the concentration of the same element in the corresponding isolated mitochondria. Treatment with metal chelators confirmed that these LMM species were labile. The dominant S species at 1100 Da was not free GSH or GSSG. PMID:26018429

Chemical Mass Shifts in a Digital Linear Ion Trap as Analytical Identity of o-, m-, and p-Xylene.

PubMed

Sun, Lulu; Xue, Bing; Huang, Zhengxu; Cheng, Ping; Ma, Li; Ding, Li; Zhou, Zhen

2018-07-01

Chemical mass shifts between isomeric ions of o-, m-, and p-xylene were measured using a digital linear ion trap, and the directions and values of the shifts were found to be correlated to the collision cross sections of the isomers. Both forward and reverse scans were used and the chemical shifts for each pair of isomers in scans of opposite directions were in opposite signs. Using different voltage settings (namely the voltage dividing ratio-VDR) of the ion trap allows adding high order field components in the quadrupole field and results in larger chemical mass shifts. The differential chemical mass shift which combined the shifts from forward and reverse scans doubled the amount of chemical shift, e.g., 0.077 Th between o- and p-xylene, enough for identification of the type of isomer without using an additional ion mobility spectrometer. The feature of equal and opposite chemical mass shifts also allowed to null out the chemical mass shift by calculating the mean m/z value between the two opposite scans and remove or reduce the mass error caused by chemical mass shift. Graphical Abstract ᅟ.

Chemical Mass Shifts in a Digital Linear Ion Trap as Analytical Identity of o-, m-, and p-Xylene

NASA Astrophysics Data System (ADS)

Sun, Lulu; Xue, Bing; Huang, Zhengxu; Cheng, Ping; Ma, Li; Ding, Li; Zhou, Zhen

2018-04-01

Chemical mass shifts between isomeric ions of o-, m-, and p-xylene were measured using a digital linear ion trap, and the directions and values of the shifts were found to be correlated to the collision cross sections of the isomers. Both forward and reverse scans were used and the chemical shifts for each pair of isomers in scans of opposite directions were in opposite signs. Using different voltage settings (namely the voltage dividing ratio-VDR) of the ion trap allows adding high order field components in the quadrupole field and results in larger chemical mass shifts. The differential chemical mass shift which combined the shifts from forward and reverse scans doubled the amount of chemical shift, e.g., 0.077 Th between o- and p-xylene, enough for identification of the type of isomer without using an additional ion mobility spectrometer. The feature of equal and opposite chemical mass shifts also allowed to null out the chemical mass shift by calculating the mean m/z value between the two opposite scans and remove or reduce the mass error caused by chemical mass shift. [Figure not available: see fulltext.

HLA-DR allele reading register shifting is associated with immunity induced by SERA peptide analogues.

PubMed

Salazar, Luz Mary; Bermúdez, Adriana; Patarroyo, Manuel E

2008-07-18

SERA protein is a leading candidate molecule to be included in an antimalarial vaccine. Conserved high activity binding peptides (HABP) binding to red blood cells (RBC) have been identified in this protein. One of them (6762) localising in the 18-kDa C-terminal fragment was used to induce protective immunity with negative result. Critical RBC binding residues (assessed by glycine-analogue scanning) were replaced by others having the same mass, volume and surface but different polarity, rendering some of them immunogenic as assessed by antibody production against the parasite or its proteins and protection-inducing against challenge with a highly infectious Aotus monkey-adapted Plasmodium falciparum strain. A shift in binding to purified HLA-DR allelic molecules from the same haplotype and in their reading register was found, suggesting that modified molecules had adopted a different (1)H NMR 3D structure allowing a better fit into the MHCII-pept-TCR complex, thereby representing a new mechanism for inducing immune protection.

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

Ward, G.E.

When intact Arbacia punctulata spermatozoa are exposed to solubilized egg jelly, the electrophoretic mobility of an abundant sperm flagellar membrane protein changes from an apparent molecular mass of 160 kDa to 150 kDa. A. punctulata spermatozoa can be labeled in vivo with /sup 32/P-labeled cells it was demonstrated that the mobility shift of the 160-kDa protein is due to dephosphorylation. The peptide resact (Cys-Val-Thr-Gly-Ala-Pro-Gly-Cys-Val-Gly-Gly-Gly-Arg-Leu-NH/sub 2/) is the component of egg jelly which is responsible for inducing the dephosphorylation. The 160/150-kdal sperm membrane protein has been purified to homogeneity by affinity chromatography on concanavalin A-agarose, and identified as sperm guanylate cyclase.more » The enzymatic activity of the guanylate cyclase is tightly coupled to its phosphorylation state. Resact has been shown to act as a potent chemoattractant for A. punctulata spermatozoa. The chemotactic response is concentration-dependent, is abolished by pretreatment of the spermatozoa with resact, and shows an absolute requirement for external calcium. This work represents the first demonstration of animal sperm chemotaxis in response to a precisely-defined molecule of egg origin. The results established a new, biologically meaningful function for resact, and may implicate sperm guanylate cyclase and cGMP in flagellar function and the chemotactic response.« less

Covalent modification and time-dependent inhibition of human CYP2E1 by the meta-isomer of acetaminophen.

PubMed

Harrelson, John P; Stamper, Brendan D; Chapman, John D; Goodlett, David R; Nelson, Sidney D

2012-08-01

The hypothesis that N-acetyl-m-aminophenol (AMAP), the meta isomer of acetaminophen, will covalently bind to and inhibit human CYP2E1 in a time- and NADPH-dependent manner was investigated. Liquid chromatography/electrospray ionization-mass spectrometry analysis indicated that AMAP metabolites (i.e., AMAP*) selectively and covalently modified CYP2E1 apoprotein in a ratio of 1.4:1 (AMAP*/CYP2E1) in a reconstituted system. The deconvoluted spectra of CYP2E1 apoprotein from incubations containing NADPH and AMAP displayed mass shifts of 167.2 ± 7.1 and 334.4 ± 6.5 Da, suggesting the addition of one and two hydroxylated AMAP metabolites to CYP2E1, respectively. Mass shifts in cytochrome P450 reductase, cytochrome b(5), and heme from these samples were not observed. CYP2E1 inhibition by AMAP increased with time in the presence of NADPH; a reversible inhibition component was also observed. The results support a bioactivation process that involves formation of a hydroquinone metabolite that undergoes further oxidation to a quinone, which reacts with CYP2E1 nucleophilic residues. The data are consistent with evidence from previous studies that identified hydroxylated AMAP glutathione conjugates collected from mice and indicate that cysteine residues are the most likely sites for adduct formation. This study reports the first direct evidence of AMAP-derived hydroquinone metabolites bound to human CYP2E1.

Nuclear localization signal-dependent and -independent movements of Drosophila melanogaster dUTPase isoforms during nuclear cleavage

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

Muha, Villo; Zagyva, Imre; Venkei, Zsolt

2009-04-03

Two dUTPase isoforms (23 kDa and 21 kDa) are present in the fruitfly with the sole difference of an N-terminal extension. In Drosophila embryo, both isoforms are detected inside the nucleus. Here, we investigated the function of the N-terminal segment using eYFP-dUTPase constructs. In Schneider 2 cells, only the 23 kDa construct showed nuclear localization arguing that it may contain a nuclear localization signal (NLS). Sequence comparisons identified a lysine-rich nonapeptide with similarity to the human c-myc NLS. In Drosophila embryos during nuclear cleavages, the 23 kDa isoform showed the expected localization shifts. Contrariwise, although the 21 kDa isoform wasmore » excluded from the nuclei during interphase, it was shifted to the nucleus during prophase and forthcoming mitotic steps. The observed dynamic localization character showed strict timing to the nuclear cleavage phases and explained how both isoforms can be present within the nuclear microenvironment, although at different stages of cell cycle.« less

The GTP-bound and Sumoylated Form of the rab17 Small Molecular Weight GTPase Selectively Binds Syntaxin 2 in Polarized Hepatic WIF-B Cells*

PubMed Central

Striz, Anneliese C.; Tuma, Pamela L.

2016-01-01

A major focus for our laboratory is identifying the molecules and mechanisms that regulate polarized apical protein sorting in hepatocytes, the major epithelial cells of the liver. These trafficking pathways are regulated, in part, by small molecular weight rab GTPases. We chose to investigate rab17, whose expression is restricted to polarized epithelial cells, is enriched in liver, and has been implicated in regulating basolateral to apical transcytosis. To initiate our studies, we generated three recombinant adenoviruses expressing wild type, constitutively active (GTP bound), or dominant-negative (GDP bound) rab17. Immunoblotting revealed rab17 immunoreactive species at 25 kDa (the predicted rab17 molecular mass) and 40 kDa. We determined that mono-sumoylation of the 25-kDa rab17 is responsible for the shift in molecular mass, and that rab17 prenylation is required for sumoylation. We further determined that sumoylation selectively promotes interactions with syntaxin 2 (but not syntaxins 3 or 4) and that these interactions are nucleotide dependent. Furthermore, a K68R-mutated rab17 led to the redistribution of syntaxin 2 and 5′ nucleotidase from the apical membrane to subapical puncta, whereas multidrug resistance protein 2 distributions were not changed. Together these data are consistent with the proposed role of rab17 in vesicle fusion with the apical plasma membrane and further implicate sumoylation as an important mediator of protein-protein interactions. The selectivity in syntaxin binding and apical protein redistribution further suggests that rab17 and syntaxin 2 mediate fusion of transcytotic vesicles at the apical surface. PMID:26957544

A comparative study of the extracellular glucosyl- and fructosyltransferases from cariogenic and non-cariogenic Streptococcus mutans strains of two different serotypes.

PubMed

Asem, K; Cornish-Bowden, A J; Cole, J A

1986-01-01

Extracellular proteins from continuous cultures of serotype c and g Streptococcus mutans strains were separated by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulphate. Gels stained with raffinose after electrophoresis revealed that although serotype c strains secrete two fructosyltransferases of molecular mass 68 kDa and 79 kDa, no fructosyltransferase was secreted by the serotype g strain K1. A sucrose activity stain was used to detect two glucosyltransferases (GTF) of molecular mass 162 kDa (bifunctional 1,6-alpha-D-glucan 3-alpha- and 6-alpha GTF or 'dextransucrase') and 153 kDa (a 1,3-alpha-D-glucan 3-alpha-GTF) in samples from cariogenic serotype c strains. Neither the 153 kDa protein nor the corresponding GTF activity was secreted by the non-cariogenic mutant C 67-25. The molecular masses of the corresponding 1,3-alpha and 1,6-alpha-GTF proteins from the serotype g strain K1 were 164 kDa and 158 kDa, respectively. All of the GTF proteins were degraded to discrete bands of lower molecular mass on storage at 4 degrees C even after extensive purification. The results provide an explanation for several outstanding controversies in the GTF literature.

Molecular Cloning, Expression and Characterization of Pectin Methylesterase (CtPME) from Clostridium thermocellum.

PubMed

Rajulapati, Vikky; Goyal, Arun

2017-05-01

Many phytopathogenic micro-organisms such as bacteria and fungi produce pectin methylesterases (PME) during plant invasion. Plants and insects also produce PME to degrade plant cell wall. In the present study, a thermostable pectin methylesterase (CtPME) from Clostridium thermocellum belonging to family 8 carbohydrate esterase (CE8) was cloned, expressed and purified. The amino acid sequence of CtPME exhibited similarity with pectin methylesterase from Erwinia chrysanthemi with 38% identity. The gene encoding CtPME was cloned into pET28a(+) vector and expressed using Escherichia coli BL21(DE3) cells. The recombinant CtPME expressed as a soluble protein and exhibited a single band of molecular mass approximately 35.2 kDa on SDS-PAGE gels. The molecular mass, 35.5 kDa of the enzyme, was also confirmed by MALDI-TOF MS analysis. Notably, highest protein concentration (11.4 mg/mL) of CtPME was achieved in auto-induction medium, as compared with LB medium (1.5 mg/mL). CtPME showed maximum activity (18.1 U/mg) against citrus pectin with >85% methyl esterification. The optimum pH and temperature for activity of CtPME were 8.5 and 50 °C, respectively. The enzyme was stable in pH range 8.0-9.0 and thermostable between 45 and 70 °C. CtPME activity was increased by 40% by 5 mM Ca 2+ or Mg 2+ ions. Protein melting curve of CtPME gave a peak at 80 °C. The peak was shifted to 85 °C in the presence of 5 mM Ca 2+ ions, and the addition of 5 mM EDTA shifted back the melting peak to 80 °C. CtPME can be potentially used in food and textile industry applications.

Preventing Fusion Mass Shift Avoids Postoperative Distal Curve Adding-on in Adolescent Idiopathic Scoliosis.

PubMed

Shigematsu, Hideki; Cheung, Jason Pui Yin; Bruzzone, Mauro; Matsumori, Hiroaki; Mak, Kin-Cheung; Samartzis, Dino; Luk, Keith Dip Kei

2017-05-01

Surgery for adolescent idiopathic scoliosis (AIS) is only complete after achieving fusion to maintain the correction obtained intraoperatively. The instrumented or fused segments can be referred to as the "fusion mass". In patients with AIS, the ideal fusion mass strategy has been established based on fulcrum-bending radiographs for main thoracic curves. Ideally, the fusion mass should achieve parallel endplates of the upper and lower instrumented vertebra and correct any "shift" for truncal balance. Distal adding-on is an important element to consider in AIS surgery. This phenomenon represents a progressive increase in the number of vertebrae included distally in the primary curvature and it should be avoided as it is associated with unsatisfactory cosmesis and an increased risk of revision surgery. However, it remains unknown whether any fusion mass shift, or shift in the fusion mass or instrumented segments, affects global spinal balance and distal adding-on after curve correction surgery in patients with AIS. (1) To investigate the relationship among postoperative fusion mass shift, global balance, and distal adding-on phenomenon in patients with AIS; and (2) to identify a cutoff value of fusion mass shift that will lead to distal adding-on. This was a retrospective study of patients with AIS from a single institution. Between 2006 and 2011 we performed 69 selective thoracic fusions for patients with main thoracic AIS. All patients were evaluated preoperatively and at 2 years postoperatively. The Cobb angle between the cranial and caudal endplates of the fusion mass and the coronal shift between them, which was defined as "fusion mass shift", were measured. Patients with a fusion mass Cobb angle greater than 20° were excluded to specifically determine the effect of fusion mass shift on distal adding-on phenomenon. Fusion mass shift was empirically set as 20 mm for analysis. Therefore, of the 69 patients who underwent selective thoracic fusion, only 52 with a fusion mass Cobb angle of 20° or less were recruited for study. We defined patients with a fusion mass shift of 20 mm or less as the balanced group and those with a fusion mass shift greater than 20 mm as the unbalanced group. A receiver operating characteristic (ROC) curve was used to determine the cutoff point of fusion mass shift for adding-on. Of the 52 patients studied, fusion mass shift (> 20 mm) was noted in 11 (21%), and six of those patients had distal adding-on at final followup. Although global spinal balance did not differ significantly between patients with or without fusion mass shift, the occurrence of adding-on phenomenon was significantly higher in the unbalanced group (55% (six of 11 patients), odds ratio [OR], 8.6; 95% CI, 2-39; p < 0.002) than the balanced group (12% [five of 41 patients]). Based on the ROC curve analysis, a fusion mass shift more than 18 mm was observed as the cutoff point for distal adding-on phenomenon (area under the curve, 0.70; 95% CI, 0.5-0.9; likelihood ratio, 5.0; sensitivity, 0.64; specificity, 0.73; positive predictive value, 39% [seven of 18 patients]; negative predictive value, 88% [30 of 34 patients]; OR, 4.8; 95% CI, 1-20; p = 0.02). Our study illustrates the substantial utility of the fulcrum-bending radiograph in determining fusion levels that can avoid fusion mass shift; thereby, underlining its importance in designing personalized surgical strategies for patients with scoliosis. Preoperatively, determining fusion levels by fulcrum-bending radiographs to avoid residual fusion mass shift is imperative. Intraoperatively, any fusion mass shift should be corrected to avoid distal adding-on, reoperation, and elevated healthcare costs. Level II, prognostic study.

Mass defect effects in atomic clocks

NASA Astrophysics Data System (ADS)

Yudin, Valeriy; Taichenachev, Alexey

2018-03-01

We consider some implications of the mass defect on the frequency of atomic transitions. We have found that some well-known frequency shifts (the gravitational shift and motion-induced shifts such as quadratic Doppler and micromotion shifts) can be interpreted as consequences of the mass defect in quantum atomic physics, i.e. without the need for the concept of time dilation used in special and general relativity theories. Moreover, we show that the inclusion of the mass defect leads to previously unknown shifts for clocks based on trapped ions.

Attitude dynamics and control of a spacecraft using shifting mass distribution

NASA Astrophysics Data System (ADS)

Ahn, Young Tae

Spacecraft need specific attitude control methods that depend on the mission type or special tasks. The dynamics and the attitude control of a spacecraft with a shifting mass distribution within the system are examined. The behavior and use of conventional attitude control actuators are widely developed and performing at the present time. However, the advantage of a shifting mass distribution concept can complement spacecraft attitude control, save mass, and extend a satellite's life. This can be adopted in practice by moving mass from one tank to another, similar to what an airplane does to balance weight. Using this shifting mass distribution concept, in conjunction with other attitude control devices, can augment the three-axis attitude control process. Shifting mass involves changing the center-of-mass of the system, and/or changing the moments of inertia of the system, which then ultimately can change the attitude behavior of the system. This dissertation consists of two parts. First, the equations of motion for the shifting mass concept (also known as morphing) are developed. They are tested for their effects on attitude control by showing how shifting the mass changes the spacecraft's attitude behavior. Second, a method for optimal mass redistribution is shown using a combinatorial optimization theory under constraints. It closes with a simple example demonstrating an optimal reconfiguration. The procedure of optimal reconfiguration from one mass distribution to another to accomplish attitude control has been demonstrated for several simple examples. Mass shifting could work as an attitude controller for fine-tuning attitude behavior in small satellites. Various constraints can be applied for different situations, such as no mass shift between two tanks connected by a failed pipe or total amount of shifted mass per pipe being set for the time optimum solution. Euler angle changes influenced by the mass reconfiguration are accomplished while stability conditions are satisfied. In order to increase the accuracy, generally, more than two control systems are installed in a satellite. Combination with another actuator will be examined to fulfill the full attitude control maneuver. Future work can also include more realistic spacecraft design and operational considerations on the behavior of this type of control system.

Counting individual sulfur atoms in a protein by ultrahighresolution Fourier transform ion cyclotron resonance mass spectrometry: Experimental resolution of isotopic fine structure in proteins

PubMed Central

Shi, Stone D.-H.; Hendrickson, Christopher L.; Marshall, Alan G.

1998-01-01

A typical molecular ion mass spectrum consists of a sum of signals from species of various possible isotopic compositions. Only the monoisotopic peak (e.g., all carbons are 12C; all nitrogens are 14N, etc.) has a unique elemental composition. Every other isotope peak at approximately integer multiples of ∼1 Da higher in nominal mass represents a sum of contributions from isotope combinations differing by a few mDa (e.g., two 13C vs. two 15N vs. one 13C and one 15N vs. 34S, vs. 18O, etc., at ∼2 Da higher in mass than the monoisotopic mass). At sufficiently high mass resolving power, each of these nominal-mass peaks resolves into its isotopic fine structure. Here, we report resolution of the isotopic fine structure of proteins up to 15.8 kDa (isotopic 13C,15N doubly depleted tumor suppressor protein, p16), made possible by electrospray ionization followed by ultrahigh-resolution Fourier transform ion cyclotron resonance mass analysis at 9.4 tesla. Further, a resolving power of m/Δm50% ≈8,000,000 has been achieved on bovine ubiquitin (8.6 kDa). These results represent a 10-fold increase in the highest mass at which isotopic fine structure previously had been observed. Finally, because isotopic fine structure reveals elemental composition directly, it can be used to confirm or determine molecular formula. For p16, for example, we were able to determine (5.1 ± 0.3) the correct number (five) of sulfur atoms solely from the abundance ratio of the resolved 34S peak to the monoisotopic peak. PMID:9751700

In-Line Reactions and Ionizations of Vaporized Diphenylchloroarsine and Diphenylcyanoarsine in Atmospheric Pressure Chemical Ionization Mass Spectrometry

NASA Astrophysics Data System (ADS)

Okumura, Akihiko; Takada, Yasuaki; Watanabe, Susumu; Hashimoto, Hiroaki; Ezawa, Naoya; Seto, Yasuo; Takayama, Yasuo; Sekioka, Ryoji; Yamaguchi, Shintaro; Kishi, Shintaro; Satoh, Takafumi; Kondo, Tomohide; Nagashima, Hisayuki; Nagoya, Tomoki

2016-07-01

We propose detecting a fragment ion (Ph2As+) using counter-flow introduction atmospheric pressure chemical ionization ion trap mass spectrometry for sensitive air monitoring of chemical warfare vomiting agents diphenylchloroarsine (DA) and diphenylcyanoarsine (DC). The liquid sample containing of DA, DC, and bis(diphenylarsine)oxide (BDPAO) was heated in a dry air line, and the generated vapor was mixed into the humidified air flowing through the sampling line of a mass spectrometer. Humidity effect on the air monitoring was investigated by varying the humidity of the analyzed air sample. Evidence of the in-line conversion of DA and DC to diphenylarsine hydroxide (DPAH) and then BDPAO was obtained by comparing the chronograms of various ions from the beginning of heating. Multiple-stage mass spectrometry revealed that the protonated molecule (MH+) of DA, DC, DPAH, and BDPAO could produce Ph2As+ through their in-source fragmentation. Among the signals of the ions that were investigated, the Ph2As+ signal was the most intense and increased to reach a plateau with the increased air humidity, whereas the MH+ signal of DA decreased. It was suggested that DA and DC were converted in-line into BDPAO, which was a major source of Ph2As+.

High-Resolution Mass Spectrometers

NASA Astrophysics Data System (ADS)

Marshall, Alan G.; Hendrickson, Christopher L.

2008-07-01

Over the past decade, mass spectrometry has been revolutionized by access to instruments of increasingly high mass-resolving power. For small molecules up to ˜400 Da (e.g., drugs, metabolites, and various natural organic mixtures ranging from foods to petroleum), it is possible to determine elemental compositions (CcHhNnOoSsPp…) of thousands of chemical components simultaneously from accurate mass measurements (the same can be done up to 1000 Da if additional information is included). At higher mass, it becomes possible to identify proteins (including posttranslational modifications) from proteolytic peptides, as well as lipids, glycoconjugates, and other biological components. At even higher mass (˜100,000 Da or higher), it is possible to characterize posttranslational modifications of intact proteins and to map the binding surfaces of large biomolecule complexes. Here we review the principles and techniques of the highest-resolution analytical mass spectrometers (time-of-flight and Fourier transform ion cyclotron resonance and orbitrap mass analyzers) and describe some representative high-resolution applications.

Cloning and characterization of an alternative splicing transcript of the gene coding for human cytidine deaminase.

PubMed

Lisboa, Bianca Cristina Garcia; Machado, Tamara da Rocha; Pimenta, Daniel Carvalho; Han, Sang Won

2007-02-01

Human cytidine deaminase (HCD) catalyzes the deamination of cytidine or deoxycytidine to uridine or deoxyuridine, respectively. The genomic sequence of HCD is formed by 31 kb with 4 exons and several alternative splicing signals, but an alternative form of HCD has yet to be reported. Here we describe the cloning and characterization of a small form of HCD, HSCD, and it is likely to be a product of alternative splicing of HCD. The alignment of DNA sequences shows that the HSCD matches HCD in 2 parts, except for a deletion of 170 bp. Based on the HCD genome organization, exons 1 and 4 should be joined and all sequences of introns and exons 2 and 3 should be deleted by splicing. This alternative splicing shifted the translation of the reading frame from the point of splicing. The estimated molecular mass is 9.8 kDa, and this value was confirmed by Western blot and mass spectroscopy after expressing the gene fused with glutathionine-S-transferase in the pGEX vector. The deletion and shift of the reading frame caused a loss of HCD activity, which was confirmed by enzyme assay and also with NIH3T3 cells modified to express HSCD and challenged against cytosine arabinoside. In this work we describe the identification and characterization of HSCD, which is the product of alternative splicing of the HCD gene.

Isolation of a 5-Kilodalton Actin-Sequestering Peptide from Human Blood Platelets

NASA Astrophysics Data System (ADS)

Safer, Daniel; Golla, Rajasree; Nachmias, Vivianne T.

1990-04-01

Resting human platelets contain ≈0.3 mM unpolymerized actin. When freshly drawn and washed platelets are treated with saponin, 85-90% of the unpolymerized actin diffuses out. Analysis by polyacrylamide gel electrophoresis under nondenaturing conditions shows that the bulk of this unpolymerized actin migrates with a higher mobility than does pure G-actin, profilactin, or actin-gelsolin complex. When muscle G-actin is added to fresh or boiled saponin extract, the added muscle actin is shifted to the high-mobility form. The saponin extract contains an acidic peptide having a molecular mass in the range of 5 kDa, which has been purified to homogeneity by reverse-phase HPLC. This peptide also shifts muscle actin to the high-mobility form. Addition of either boiled saponin extract or the purified peptide to muscle G-actin also strongly and stoichiometrically inhibits salt-induced polymerization, as assayed by falling-ball viscometry and by sedimentation. We conclude that this peptide binds to the bulk of the unpolymerized actin in platelets and prevents it from polymerizing.

Quaternary structure and apical membrane sorting of the mammalian NaSi-1 sulfate transporter in renal cell lines.

PubMed

Regeer, Ralf R; Nicke, Annette; Markovich, Daniel

2007-01-01

NaSi-1 encodes a Na(+)-sulfate cotransporter expressed on the apical membrane of renal proximal tubular cells, which is responsible for body sulfate homeostasis. Limited information is available on NaSi-1 protein structure and the mechanisms controlling its apical membrane sorting. The aims of this study were to biochemically determine the quaternary structure of the rat NaSi-1 protein and to characterize its expression in renal epithelial cell lines. Hexahistidyl-tagged NaSi-1 (NaSi-1-His) proteins expressed in Xenopus oocytes, appeared as two bands of about 60 and 75 kDa. PNGase F treatment shifted both bands to 57 kDa while endoglycosidase H treatment led to a downward shift of the lower molecular mass band only. Mutagenesis of a putative N-glycosylation site (N591S) produced a single band that was not shifted by endoglycosidase H or PNGase F, confirming a single glycosylation site at residue 591. Blue native-PAGE and cross-linking experiments revealed dimeric complexes, suggesting the native form of NaSi-1 to be a dimer. Transient transfection of EGFP/NaSi-1 in renal epithelial cells (OK, LLC-PK1 and MDCK) demonstrated apical membrane sorting, which was insensitive to tunicamycin. Transfection of the EGFP/NaSi-1 N591S glycosylation mutant also showed apical expression, suggesting N591 is not essential for apical sorting. Treatment with cholesterol depleting compounds did not disrupt apical sorting, but brefeldin A led to misrouting to the basolateral membrane, suggesting that NaSi-1 sorting is through the ER to Golgi pathway. Our data demonstrates that NaSi-1 forms a dimeric protein which is glycosylated at N591, whose sorting to the apical membrane in renal epithelial cells is brefeldin A-sensitive and independent of lipid rafts or glycosylation.

Selective detection of thiosulfate-containing peptides using tandem mass spectrometry.

PubMed

Raftery, Mark J

2005-01-01

Incubation of proteins or peptides containing disulfide bonds (S-S) with sodium sulfite (Na(2)SO(3)) cleaves S-S bonds producing approximately equimolar amounts of free thiols (-SH) and thiosulfates (-S-SO(3)H), a process known as sulfitolysis. Proteins and peptides containing thiosulfates were separated by reverse-phase high-performance liquid chromatography (RP-HPLC) and characterized by mass spectrometry (MS) and peptide mapping. The mass of the thiosulfate-containing peptide formed from oxidized insulin B chain was 3478.02 Da, 80 Da greater than the reduced peptide and corresponding precisely to addition of sulfur trioxide (SO(3)). Disulfide bond cleavage was also observed using RP-HPLC and MS after incubation of the intramolecular homodimer of mouse S100A8 (mass 20614 Da). The mass of HPLC-separated A8-SH was 10308 Da, and 10388 Da for A8-S-SO(3)H. Loss of SO(3) from multiply charged precursor ions was generally observed at elevated declustering potentials in the source region or within q(2) at relatively low collision energies (approximately 20 V). The characteristic loss of SO(3) at low collision energies preceded peptide backbone fragmentations at higher collision energies. Accurate mass measurement and charge-state discrimination, using a hybrid quadrupole time-of-flight mass spectrometer, allowed specific detection of thiosulfate-containing peptides. An information-dependent acquisition method, where the switch criterion was loss of m/z 79.9568, specifically identified 11 thiosulfate-containing peptides using nano-LC/MS from a tryptic digest of bovine serum albumin (BSA).

The Cosmic Dust Analyzer for Cassini

NASA Technical Reports Server (NTRS)

Bradley, James G.; Gruen, Eberhard; Srama, Ralf

1996-01-01

The Cosmic Dust Analyzer (CDA) is designed to characterize the dust environment in interplanetary space, in the Jovian and in the Saturnian systems. The instrument consists of two major components, the Dust Analyzer (DA) and the High Rate Detector (HRD). The DA has a large aperture to provide a large cross section for detection in low flux environments. The DA has the capability of determining dust particle mass, velocity, flight direction, charge, and chemical composition. The chemical composition is determined by the Chemical Analyzer system based on a time-of-flight mass spectrometer. The DA is capable of making full measurements up to one impact/second. The HRD contains two smaller PVDF detectors and electronics designed to characterize dust particle masses at impact rates up to 10(exp 4) impacts/second. These high impact rates are expected during Saturn ring, plane crossings.

Chromatography and mass spectrometry of prebiological and biological molecules

NASA Astrophysics Data System (ADS)

Navale, Vivek

The detection and identification of prebiological and biological molecules are of importance for understanding chemical and biological processes occurring within the solar system. Molecular mass measurements, peptide mapping, and disulfide bond analysis of enzymes and recombinant proteins are important in the development of therapeutic drugs for human diseases. Separation of hydrocarbons (C1 to C6) and nitriles was achieved by 14%-cyanopropylphenyl-86%- dimethylpolysiloxane (CPPS-DMPS) stationary phase in a narrow bore metal capillary column. The calculation of modeling numbers enabled the differentiation of the C4 hydrocarbon isomers of 1-butene (cis and trans). The modeled retention time values for benzene, toluene, xylene, acetonitrile, propane, and propene nitriles were in good agreement with the measurements. The separation of C2 hydrocarbons (ethane and ethene) from predominantly N2 matrix was demonstrated for the first time on wall coated narrow bore low temperature glassy carbon column. Identification and accurate mass measurements of pepsin, an enzymatic protein with less number of basic amino acid residues were successfully demonstrated by matrix- assisted laser desorption ionization mass spectrometry (MALDI-MS). The molecular mass of pepsin was found to be 34,787 Da. Several decomposition products of pepsin, in m/z range of 3,500 to 4,700 were identified. Trypsin, an important endopeptidase enzyme had a mass of 46829.7 Da. Lower mass components with m/z 8047.5, 7776.6, 5722, 5446.2 and 5185 Da were also observed in trypsin spectrum. Both chemokine and growth factor recombinant proteins were mass analyzed as 8848.1 ± 3.5 and 16178.52 ± 4.1 Da, respectively. The accuracy of the measurements was in the range of 0.01 to 0.02%. Reduction and alkylation experiments on the chemokine showed the presence of six cysteines and three disulfide bonds. The two cysteines of the growth factor contained the free sulfhydryl groups and the accurate average mass of the growth factor protein was 16175.6 Da. MALDI analysis of trypsin digest of Myeloid progenitor inhibitory factor chemokine verified the disulfide bridging among cysteine residues. Several partially digested trypsin and V8 peptides were detected that verified significant portions of the primary structure of the chemokine. Mass difference amounting to the loss of a single amino acid, serine was also identified. The cyanogen bromide (CNBr) treated chemokine produced three peptides 7051, 6910.1 and 1492 Da. The analysis of Keratinocyte growth factor (KGF) peptide mixtures showed suppression effects during the MALDI ionization process. Several partially digested peptides with mass values 3214, 9980, 10325 and 10497 Da were identified. Direct MALDI-MS analysis of cyanogen bromide treated KGF molecule demonstrated the formation of peptides with mass 7567.3, 4992.6 and 3118.6 Da. The high sensitivity of MALDI-MS provided a rapid method for confirming the fidelity of gene expression in the host system. The present work showed that the combined methods of chromatography and mass spectrometry are efficient means for identification and characterization of prebiological and biological molecules.

Bounding the Higgs boson width through interferometry.

PubMed

Dixon, Lance J; Li, Ye

2013-09-13

We study the change in the diphoton-invariant-mass distribution for Higgs boson decays to two photons, due to interference between the Higgs resonance in gluon fusion and the continuum background amplitude for gg→γγ. Previously, the apparent Higgs mass was found to shift by around 100 MeV in the standard model in the leading-order approximation, which may potentially be experimentally observable. We compute the next-to-leading-order QCD corrections to the apparent mass shift, which reduce it by about 40%. The apparent mass shift may provide a way to measure, or at least bound, the Higgs boson width at the Large Hadron Collider through "interferometry." We investigate how the shift depends on the Higgs width, in a model that maintains constant Higgs boson signal yields. At Higgs widths above 30 MeV, the mass shift is over 200 MeV and increases with the square root of the width. The apparent mass shift could be measured by comparing with the ZZ* channel, where the shift is much smaller. It might be possible to measure the shift more accurately by exploiting its strong dependence on the Higgs transverse momentum.

Detection of over 100 selenium metabolites in selenized yeast by liquid chromatography electrospray time-of-flight mass spectrometry.

PubMed

Gilbert-López, Bienvenida; Dernovics, Mihaly; Moreno-González, David; Molina-Díaz, Antonio; García-Reyes, Juan F

2017-08-15

The characterization of the selenometabolome of Selenized(Se)-yeast, that is the fraction of water soluble low-molecular weight Se-metabolites produced in Se-yeast is of paramount interest to expand the knowledge on the composition of this food supplement. In this work, we have applied liquid chromatography electrospray time-of-flight mass spectrometry (LC-TOFMS) to search for Se-species from the low molecular weight range fraction of the selenized yeast used for food supplements. Prior to LC-TOFMS, sample treatment consisted of ultrasound assisted water extraction followed by size exclusion fractionation assisted with off-line inductively coupled plasma mass spectrometry detection of isotope 82 Se. The fraction corresponding to low-molecular weight species was subjected to LC-TOFMS using electrospray ionization in the positive ion mode. The detection of the suspected selenized species has been based on the information obtained from accurate mass measurements of both the protonated molecules and fragments from in-source CID fragmentation; along with the characteristic isotope pattern exhibited by the presence of Se. The approach enables the detection of 103 selenized species, most of them not previously reported, in the range from ca. 300-650Da. Besides the detection of selenium species, related sulphur derivate metabolites were detected based on the accurate mass shift due to the substitution of sulphur and selenium. Copyright © 2017. Published by Elsevier B.V.

Application of parylene for surface (polymer) enhanced laser desorption/ionization of synthetic polymers.

PubMed

Miksa, Beata J; Sochacki, Marek; Sroka-Bartnicka, Anna; Uznański, Paweł; Nosal, Andrzej; Potrzebowski, Marek J

2013-04-15

Synthetic polymers of molecular masses up to a few kDa can be analyzed without the use of any matrix by direct laser desorption/ionization mass spectrometry (LDI-MS). In this technique, the surface of the sample plate plays a crucial role, and many attempts have been made to understand the influence of the surface on the ease of desorption. Since this technique requires no tedious sample pretreatment, it is a promising method for the rapid characterization of various synthetic polymers. Parylene (poly(p-xylylenes), PPX) was tested as a surface support for studying the molecular masses of biocompatible polymers: poly(ethylene glycol) (PEG), poly(L-lactide) (PLLA), and poly(methyl methacrylate) (PMMA). The average molecular masses of the polymers were: PEG (600.0 Da and 3.5 kDa), PMMA (2.0 kDa), and PLLA (2.8 kDa). LDI mass spectra of polymers deposited on parylene were enhanced by a factor of two over those obtained directly from the gold target plate. Modification of the surface of the target plate by the addition of a PPX layer extended the functionality of LDI-TOF MS, especially for the analysis of low-mass compounds. The LDI analysis using the PPX-coated target plate provided details of polymers including: end-group, composition, monomer unit, and molecular mass distribution. The average molecular weights of four tested polymers on the gold target plate and the PPX support were unchanged, indicating that sample degradation was not occurring despite the high energy of the laser beam. The LDI investigations showed that the PPX support boosted ion yields by a factor of two compared with the gold target plate. Copyright © 2013 John Wiley & Sons, Ltd.

MOLECULAR MECHANISM OF HUMAN NRF2 ACTIVATION AND DEGRADATION: ROLE OF SEQUENTIAL PHOSPHORYLATION BY PROTEIN KINASE CK2

PubMed Central

Pi, Jingbo; Bai, Yushi; Reece, Jeffrey M.; Williams, Jason; Liu, Dianxin; Freeman, Michael L.; Fahl, William E.; Shugar, David; Liu, Jie; Qu, Wei; Collins, Sheila; Waalkes, Michael P.

2007-01-01

Nrf2 is a key transcription factor in the cellular response to oxidative stress. In this study we first identify two phosphorylated forms of endogenous human Nrf2 after chemically-induced oxidative stress and provide evidence that protein kinase CK2-mediated sequential phosphorylation plays potential role in Nrf2 activation and degradation. Human Nrf2 has a predicted molecular mass of 66 kDa. However, immunoblots showed that two bands at 98 and 118 kDa, which are identified as phosphorylated forms, are increased in response to Nrf2 inducers. In addition, human Nrf2 was found to be a substrate for CK2 which mediated two steps of phosphorylation, resulting in two forms of Nrf2 migrating with differing Mr at 98 kDa (Nrf2–98) and 118 kDa (Nrf2–118). Our results support a role in which calmodulin binding regulates CK2 activity, in that cold (25 °C) in Ca2+-free media (cold/Ca2+-free) decreased both cellular calcium levels and CK2-calmodulin binding and induced Nrf2–118 formation, the latter of which was prevented by CK2 specific inhibitors. Gel-shift assays showed that the Nrf2–118 generated under cold/Ca2+-free conditions does not bind to the antioxidant response element, indicating that Nrf2–98 has transcriptional activity. In contrast, Nrf2–118 is more susceptible to degradation. These results provide evidence for phosphorylation by CK2 as a critical controlling factor in Nrf2-mediated cellular antioxidant response. PMID:17512459

Examining Troughs in the Mass Distribution of All Theoretically Possible Tryptic Peptides

PubMed Central

Nefedov, Alexey V.; Mitra, Indranil; Brasier, Allan R.; Sadygov, Rovshan G.

2011-01-01

This work describes the mass distribution of all theoretically possibly tryptic peptides made of 20 amino acids, up to the mass of 3 kDa, with resolution of 0.001 Da. We characterize regions between the peaks of the distribution, including gaps (forbidden zones) and low-populated areas (quiet zones). We show how the gaps shrink over the mass range, and when they completely disappear. We demonstrate that peptide compositions in quiet zones are less diverse than those in the peaks of the distribution, and that by eliminating certain types of unrealistic compositions the gaps in the distribution may be increased. The mass distribution is generated using a parallel implementation of a recursive procedure that enumerates all amino acid compositions. It allows us to enumerate all compositions of tryptic peptides below 3 kDa in 48 minutes using a computer cluster with 12 Intel Xeon X5650 CPUs (72 cores). The results of this work can be used to facilitate protein identification and mass defect labeling in mass spectrometry-based proteomics experiments. PMID:21780838

Mass Spectrometry and Fourier Transform Infrared Spectroscopy for Analysis of Biological Materials

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

Anderson, Timothy J.

Time-of-flight mass spectrometry along with statistical analysis was utilized to study metabolic profiles among rats fed resistant starch (RS) diets. Fischer 344 rats were fed four starch diets consisting of 55% (w/w, dbs) starch. A control starch diet consisting of corn starch was compared against three RS diets. The RS diets were high-amylose corn starch (HA7), HA7 chemically modified with octenyl succinic anhydride, and stearic-acid-complexed HA7 starch. A subgroup received antibiotic treatment to determine if perturbations in the gut microbiome were long lasting. A second subgroup was treated with azoxymethane (AOM), a carcinogen. At the end of the eight weekmore » study, cecal and distal-colon contents samples were collected from the sacrificed rats. Metabolites were extracted from cecal and distal colon samples into acetonitrile. The extracts were then analyzed on an accurate-mass time-of-flight mass spectrometer to obtain their metabolic profile. The data were analyzed using partial least-squares discriminant analysis (PLS-DA). The PLS-DA analysis utilized a training set and verification set to classify samples within diet and treatment groups. PLS-DA could reliably differentiate the diet treatments for both cecal and distal colon samples. The PLS-DA analyses of the antibiotic and no antibiotic treated subgroups were well classified for cecal samples and modestly separated for distal-colon samples. PLS-DA analysis had limited success separating distal colon samples for rats given AOM from those not treated; the cecal samples from AOM had very poor classification. Mass spectrometry profiling coupled with PLS-DA can readily classify metabolite differences among rats given RS diets.« less

High-resolution time-of-flight mass spectrometry fingerprinting of metabolites from cecum and distal colon contents of rats fed resistant starch

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

Anderson, Timothy J.; Jones, Roger W.; Ai, Yongfeng

Time-of-flight mass spectrometry along with statistical analysis was utilized to study metabolic profiles among rats fed resistant starch (RS) diets. Fischer 344 rats were fed four starch diets consisting of 55 % (w/w, dbs) starch. A control starch diet consisting of corn starch was compared against three RS diets. The RS diets were high-amylose corn starch (HA7), HA7 chemically modified with octenyl succinic anhydride, and stearic-acid-complexed HA7 starch. A subgroup received antibiotic treatment to determine if perturbations in the gut microbiome were long lasting. A second subgroup was treated with azoxymethane (AOM), a carcinogen. At the end of the 8-weekmore » study, cecal and distal colon content samples were collected from the sacrificed rats. Metabolites were extracted from cecal and distal colon samples into acetonitrile. The extracts were then analyzed on an accurate-mass time-of-flight mass spectrometer to obtain their metabolic profile. The data were analyzed using partial least-squares discriminant analysis (PLS-DA). The PLS-DA analysis utilized a training set and verification set to classify samples within diet and treatment groups. PLS-DA could reliably differentiate the diet treatments for both cecal and distal colon samples. The PLS-DA analyses of the antibiotic and no antibiotic-treated subgroups were well classified for cecal samples and modestly separated for distal colon samples. PLS-DA analysis had limited success separating distal colon samples for rats given AOM from those not treated; the cecal samples from AOM had very poor classification. Mass spectrometry profiling coupled with PLS-DA can readily classify metabolite differences among rats given RS diets.« less

The interaction between ketamine and some crown ethers in common organic solvents studied by NMR: The effect of donating atoms and ligand structure

NASA Astrophysics Data System (ADS)

Chekin, Fereshteh; Bordbar, Maryam; Fathollahi, Yaghoub; Alizadeh, Naader

2006-02-01

1H NMR spectroscopy was used to investigate the stoichiometry and stability of the drug ketamine cation complexes with some crown ethers, such as 15-crown-5 (15C5), aza-15-crown-5 (A15C5), 18-crown-6 (18C6), aza-18-crown-6 (A18C6), diaza-18-crown-6 (DA18C6), dibenzyl-diaza-18-crown-6 (DBzDA18C6) and cryptant [2,2,2] (C222) in acetonitrile (AN), dimethylsulfoxide (DMSO) and methanol (MeOH) at 27 °C. In order to evaluate the formation constants of the ketamine cation complexes, the CH 3 protons chemical shift (on the nitrogen atom of ketamine) was measured as function of ligand/ketamine mole ratio. The formation constant of resulting complexes were calculated by the computer fitting of chemical shift versus mole ratio data to appropriate equations. A significant chemical shift variation was not observed for 15C5 and 18C6. The stoichiometry of the mono aza and diaza ligands are 1:1 and 1:2 (ligand/ketamine), respectively. In all of the solvents studied, DA18C6 formed more stable complexes than other ligands. The solvent effect on the stability of these complexes is discussed.

Dopamine cells respond to predicted events during classical conditioning: evidence for eligibility traces in the reward-learning network.

PubMed

Pan, Wei-Xing; Schmidt, Robert; Wickens, Jeffery R; Hyland, Brian I

2005-06-29

Behavioral conditioning of cue-reward pairing results in a shift of midbrain dopamine (DA) cell activity from responding to the reward to responding to the predictive cue. However, the precise time course and mechanism underlying this shift remain unclear. Here, we report a combined single-unit recording and temporal difference (TD) modeling approach to this question. The data from recordings in conscious rats showed that DA cells retain responses to predicted reward after responses to conditioned cues have developed, at least early in training. This contrasts with previous TD models that predict a gradual stepwise shift in latency with responses to rewards lost before responses develop to the conditioned cue. By exploring the TD parameter space, we demonstrate that the persistent reward responses of DA cells during conditioning are only accurately replicated by a TD model with long-lasting eligibility traces (nonzero values for the parameter lambda) and low learning rate (alpha). These physiological constraints for TD parameters suggest that eligibility traces and low per-trial rates of plastic modification may be essential features of neural circuits for reward learning in the brain. Such properties enable rapid but stable initiation of learning when the number of stimulus-reward pairings is limited, conferring significant adaptive advantages in real-world environments.

Cocaine inhibition of nicotinic acetylcholine receptors influences dopamine release

PubMed Central

Acevedo-Rodriguez, Alexandra; Zhang, Lifen; Zhou, Fuwen; Gong, Suzhen; Gu, Howard; De Biasi, Mariella; Zhou, Fu-Ming; Dani, John A.

2014-01-01

Nicotinic acetylcholine receptors (nAChRs) potently regulate dopamine (DA) release in the striatum and alter cocaine's ability to reinforce behaviors. Since cocaine is a weak nAChR inhibitor, we hypothesized that cocaine may alter DA release by inhibiting the nAChRs in DA terminals in the striatum and thus contribute to cocaine's reinforcing properties primarily associated with the inhibition of DA transporters. We found that biologically relevant concentrations of cocaine can mildly inhibit nAChR-mediated currents in midbrain DA neurons and consequently alter DA release in the dorsal and ventral striatum. At very high concentrations, cocaine also inhibits voltage-gated Na channels in DA neurons. Furthermore, our results show that partial inhibition of nAChRs by cocaine reduces evoked DA release. This diminution of DA release via nAChR inhibition more strongly influences release evoked at low or tonic stimulation frequencies than at higher (phasic) stimulation frequencies, particularly in the dorsolateral striatum. This cocaine-induced shift favoring phasic DA release may contribute to the enhanced saliency and motivational value of cocaine-associated memories and behaviors. PMID:25237305

On the Mechanism for a Gravity Effect Using Type 2 Superconductors

NASA Technical Reports Server (NTRS)

Robertson, Glen A.

1999-01-01

In this paper, we formulate a percent mass change equation based on Woodward's transient mass shift and the Cavendish balance equations applied to superconductor Josephson junctions, A correction to the transient mass shift equation is presented due to the emission of the mass energy from the superconductor. The percentage of mass change predicted by the equation was estimated against the maximum percent mass change reported by Podkletnov in his gravity shielding experiments. An experiment is then discussed, which could shed light on the transient mass shift near superconductor and verify the corrected gravitational potential.

Demonstration by Mass Spectrometry that Purified Native Treponema pallidum Rare Outer Membrane Protein 1 (Tromp1) Has a Cleaved Signal Peptide

PubMed Central

Blanco, David R.; Whitelegge, Julian P.; Miller, James N.; Lovett, Michael A.

1999-01-01

Purified native Tromp1 was subjected to mass spectrometric analysis in order to determine conclusively whether this protein possesses a cleaved or uncleaved signal peptide. The molecular masses of Tromp1, three Treponema pallidum lipoproteins, and a bovine serum albumin (BSA) control were determined by matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry. The molecular masses of all of the T. pallidum lipoproteins and BSA were within 0.7% of their respective calculated masses. The molecular mass of Tromp1 was 31,510 Da, which is consistent with a signal-less form of Tromp1, given a calculated mass of unprocessed Tromp1 of 33,571 Da, a difference of 2,061 Da (a 6.5% difference). Purified native Tromp1 was also subjected to MALDI-TOF analysis in comparison to recombinant Tromp1 following cyanogen bromide cleavage, which further confirmed the identity of Tromp1 and showed that native Tromp1 was not degraded at the carboxy terminus. These studies confirm that Tromp1 is processed and does not contain an uncleaved signal peptide as previously reported. PMID:10438785

Anomalous center of mass shift: gravitational dipole moment.

NASA Astrophysics Data System (ADS)

Jeong, Eue Jin

1997-02-01

The anomalous, energy dependent shift of the center of mass of an idealized, perfectly rigid, uniformly rotating hemispherical shell which is caused by the relativistic mass increase effect is investigated in detail. It is shown that a classical object on impact which has the harmonic binding force between the adjacent constituent particles has the similar effect of the energy dependent, anomalous shift of the center of mass. From these observations, the general mode of the linear acceleration is suggested to be caused by the anomalous center of mass shift whether it's due to classical or relativistic origin. The effect of the energy dependent center of mass shift perpendicular to the plane of rotation of a rotating hemisphere appears as the non zero gravitational dipole moment in general relativity. Controlled experiment for the measurement of the gravitational dipole field and its possible links to the cylindrical type line formation of a worm hole in the extreme case are suggested. The jets from the black hole accretion disc and the observed anomalous red shift from far away galaxies are considered to be the consequences of the two different aspects of the dipole gravity.

Identification of New Drug Targets in Multi-Drug Resistant Bacterial Infections

DTIC Science & Technology

2014-10-01

adenylation domain of ABBFA_003406. We used mass spectrometry to test if threonine once activated by the adenylating activity of ABBFA_003406 becomes...loaded with threonine , forming a thioester linkage. The ABBFA_003404 carrier protein is produced with a histidine-tag, used for purification, that is...increase in mass of 341 Da. Further addition of the threonine moiety results in an increase in mass of an additional 73 Da. 13 W81XWH-11-2-0218

Two new 4-Cys conotoxins (framework 14) of the vermivorous snail Conus austini from the Gulf of Mexico with activity in the central nervous system of mice

PubMed Central

Zugasti-Cruz, Alejandro; Falcón, Andrés; Heimer de la Cotera, Edgar P.; Olivera, Baldomero M.; Aguilar, Manuel B.

2008-01-01

As part of continuing studies of the venom components present in Conus austini (syn.: Conus cancellatus), a vermivorous cone snail collected in the western Gulf of Mexico, Mexico, two major peptides, as14a and as14b, were purified and characterized. Their amino acid sequences were determined by automatic Edman sequencing after reduction and alkylation. Their molecular masses, established by matrix-assisted laser desorption ionization time-of-flight mass spectrometry, confirmed the chemical analyses and indicated that as14a and as14b have free C-termini. Each peptide contains four Cys residues arranged in a pattern (C-C-C-C, framework 14). The primary structure of as14a is GGVGRCIYNCMNSGGGLNFIQCKTMCY (experimental monoisotopic mass 2,883.92 Da; calculated monoisotopic mass 2,884.20 Da), whereas that of as14b is RWDVDQCIYYCLNGVVGYSYTECQTMCT (experimental monoisotopic mass 3,308.63 Da; calculated monoisotopic mass 3,308.34 Da). Both purified peptides elicited scratching and grooming activity in mice, and as14b also caused body and rear limb extension and tail curling immediately upon injection. The high sequence similarity of peptide as14a with peptide vil14a from the vermivorous C. villepinii suggests that the former might block K+ channels. PMID:18206266

Keratin capped silver nanoparticles - synthesis and characterization of a nanomaterial with desirable handling properties

USDA-ARS?s Scientific Manuscript database

Silver nanoparticles (NPs) were produced with keratin stabilizer and the NPs exhibited unimodal Gaussian distribution with average diameter of 3.5nm +/- 0.7 nm. The molecular mass of keratin stabilizer was 6-8 kDa. The mass of keratin capped NPs was >250 kDa to indicate the formation of crosslinked...

Chemical Visualization of Sweat Pores in Fingerprints Using GO-Enhanced TOF-SIMS.

PubMed

Cai, Lesi; Xia, Meng-Chan; Wang, Zhaoying; Zhao, Ya-Bin; Li, Zhanping; Zhang, Sichun; Zhang, Xinrong

2017-08-15

Time-of-flight secondary ion mass spectrometry (TOF-SIMS) has been used in imaging of small molecules (<500 Da) in fingerprints, such as gunshot residues and illicit drugs. However, identifying and mapping relatively high mass molecules are quite difficult owing to insufficient ion yield of their molecular ions. In this report, graphene oxide (GO)-enhanced TOF-SIMS was used to detect and image relatively high mass molecules such as poison, alkaloids (>600 Da) and controlled drugs, and antibiotics (>700 Da) in fingerprints. Detail features of fingerprints such as the number and distribution of sweat pores in a ridge and even the delicate morphology of one pore were clearly revealed in SIMS images of relatively high mass molecules. The detail features combining with identified chemical composition were sufficient to establish a human identity and link the suspect to a crime scene. The wide detectable mass range and high spatial resolution make GO-enhanced TOF-SIMS a promising tool in accurate and fast analysis of fingerprints, especially in fragmental fingerprint analysis.

High molecular weight glutenin subunits in some durum wheat cultivars investigated by means of mass spectrometric techniques.

PubMed

Muccilli, Vera; Lo Bianco, Marisol; Cunsolo, Vincenzo; Saletti, Rosaria; Gallo, Giulia; Foti, Salvatore

2011-11-23

The primary structures of high molecular weight glutenin subunits (HMW-GS) of 5 Triticum durum Desf. cultivars (Simeto, Svevo, Duilio, Bronte, and Sant'Agata), largely cultivated in the south of Italy, and of 13 populations of the old spring Sicilian durum wheat landrace Timilia (Triticum durum Desf.) (accession nos. 1, 2, 3, 4, 7, 8, 9, 13, 14, 15, SG1, SG2, and SG3) were investigated using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) and reversed-phase high performance liquid chromatography/nanoelectrospray ionization mass spectrometry (RP-HPLC/nESI-MS/MS). M(r) of the intact proteins determined by MALDI mass spectrometry showed that all the 13 populations of Timilia contained the same two HMW-GS with 75.2 kDa and 86.4 kDa, whereas the other durum wheat cultivars showed the presence of the expected HMW-GS 1By8 and 1Bx7 at 75.1 kDa and 83.1 kDa, respectively. By MALDI mass spectrometry of the tryptic digestion peptides of the isolated HMW-GS of Timilia, the 1Bx and 1By subunits were identified as the NCBInr Acc. No AAQ93629, and AAQ93633, respectively. Sequence verification for HMW-GS 1Bx and 1By both in Simeto and Timilia was obtained by MALDI mass mapping and HPLC/nESI-MSMS of the tryptic peptides. The Bx subunit of Timila presents a sequence similarity of 96% with respect to Simeto, with differences in the insertion of 3 peptides of 5, 9, and 15 amino acids, for a total insertion of 29 amino acids and 25 amino acid substitutions. These differences in the amino acidic sequence account for the determined Δm of 3294 Da between the M(r) of the 1Bx subunits in Timilia and Simeto. Sequence alignment between the two By subunits shows 10 amino acid substitutions and is consistent with the Δm of 148 Da found in the MALDI mass spectra of the intact subunits.

Quadrupole Ion Mass Spectrometer for Masses of 2 to 50 Da

NASA Technical Reports Server (NTRS)

Helms, William; Griffin, Timothy P.; Ottens, Andrew; Harrison, Willard

2005-01-01

A customized quadrupole ion-trap mass spectrometer (QITMS) has been built to satisfy a need for a compact, rugged instrument for measuring small concentrations of hydrogen, helium, oxygen, and argon in a nitrogen atmosphere. This QITMS can also be used to perform quantitative analyses of other gases within its molecular-mass range, which is 2 to 50 daltons (Da). (More precisely, it can be used to perform quantitative analysis of gases that, when ionized, are characterized by m/Z ratios between 2 and 50, where m is the mass of an ion in daltons and Z is the number of fundamental electric charges on the ion.

High-Resolution NMR Reveals Secondary Structure and Folding of Amino Acid Transporter from Outer Chloroplast Membrane

PubMed Central

Zook, James D.; Molugu, Trivikram R.; Jacobsen, Neil E.; Lin, Guangxin; Soll, Jürgen; Cherry, Brian R.; Brown, Michael F.; Fromme, Petra

2013-01-01

Solving high-resolution structures for membrane proteins continues to be a daunting challenge in the structural biology community. In this study we report our high-resolution NMR results for a transmembrane protein, outer envelope protein of molar mass 16 kDa (OEP16), an amino acid transporter from the outer membrane of chloroplasts. Three-dimensional, high-resolution NMR experiments on the 13C, 15N, 2H-triply-labeled protein were used to assign protein backbone resonances and to obtain secondary structure information. The results yield over 95% assignment of N, HN, CO, Cα, and Cβ chemical shifts, which is essential for obtaining a high resolution structure from NMR data. Chemical shift analysis from the assignment data reveals experimental evidence for the first time on the location of the secondary structure elements on a per residue basis. In addition T 1Z and T2 relaxation experiments were performed in order to better understand the protein dynamics. Arginine titration experiments yield an insight into the amino acid residues responsible for protein transporter function. The results provide the necessary basis for high-resolution structural determination of this important plant membrane protein. PMID:24205117

A Bayesian Markov-chain-based heteroscedastic regression model for the analysis of 18O-labeled mass spectra.

PubMed

Zhu, Qi; Burzykowski, Tomasz

2011-03-01

To reduce the influence of the between-spectra variability on the results of peptide quantification, one can consider the (18)O-labeling approach. Ideally, with such labeling technique, a mass shift of 4 Da of the isotopic distributions of peptides from the labeled sample is induced, which allows one to distinguish the two samples and to quantify the relative abundance of the peptides. It is worth noting, however, that the presence of small quantities of (16)O and (17)O atoms during the labeling step can cause incomplete labeling. In practice, ignoring incomplete labeling may result in the biased estimation of the relative abundance of the peptide in the compared samples. A Markov model was developed to address this issue (Zhu, Valkenborg, Burzykowski. J. Proteome Res. 9, 2669-2677, 2010). The model assumed that the peak intensities were normally distributed with heteroscedasticity using a power-of-the-mean variance funtion. Such a dependence has been observed in practice. Alternatively, we formulate the model within the Bayesian framework. This opens the possibility to further extend the model by the inclusion of random effects that can be used to capture the biological/technical variability of the peptide abundance. The operational characteristics of the model were investigated by applications to real-life mass-spectrometry data sets and a simulation study. © American Society for Mass Spectrometry, 2011

Different protein profile in amniotic fluid with nervous system malformations by surface-enhanced laser desorption-ionization/time-of-flight mass spectrometry (SELDI-TOF-MS) technology.

PubMed

Ma, Zhe; Liu, Cun; Deng, Biping; Dong, Shaogang; Tao, Guowei; Zhan, Xinfeng; Wang, Chuner; Liu, Shaoping; Qu, Xun

2010-12-01

To detect the distinct proteins in amniotic fluid (AF) between nervous system malformations fetuses and normal fetuses. Surface-enhanced laser desorption-ionization/time-of-flight mass spectrometry was used to characterize AF peptides in AF between nervous system malformations fetuses and normal fetuses. WCX2 protein chips were used to characterize AF peptides in AF. Protein chips were examined in a PBSIIC protein reader, the protein profiling was collected by ProteinChip software version 3.1 (Ciphergen Biosystems, Fremont, CA, USA) and analyzed by Biomarker Wizard software (Ciphergen Biosystems). Nine distinct proteins were identified in AF between nervous system malformations fetuses and normal fetuses. Compared with the control group, three proteins with m/z 4967.5 Da, 5258.0 Da, and 11,717.0 Da were down-regulated, and six proteins with m/z 2540.4 Da, 3107.1 Da, 3396.8 Da, 4590.965 Da, 5589.2 Da and 6429.4 Da up-regulated in nervous system malformations fetuses. The results suggest that there are distinct proteins in protein profiling of AF between nervous system malformations fetuses and normal fetuses. © 2010 The Authors. Journal of Obstetrics and Gynaecology Research © 2010 Japan Society of Obstetrics and Gynecology.

Morphine disinhibits glutamatergic input to VTA dopamine neurons and promotes dopamine neuron excitation.

PubMed

Chen, Ming; Zhao, Yanfang; Yang, Hualan; Luan, Wenjie; Song, Jiaojiao; Cui, Dongyang; Dong, Yi; Lai, Bin; Ma, Lan; Zheng, Ping

2015-07-24

One reported mechanism for morphine activation of dopamine (DA) neurons of the ventral tegmental area (VTA) is the disinhibition model of VTA-DA neurons. Morphine inhibits GABA inhibitory neurons, which shifts the balance between inhibitory and excitatory input to VTA-DA neurons in favor of excitation and then leads to VTA-DA neuron excitation. However, it is not known whether morphine has an additional strengthening effect on excitatory input. Our results suggest that glutamatergic input to VTA-DA neurons is inhibited by GABAergic interneurons via GABAB receptors and that morphine promotes presynaptic glutamate release by removing this inhibition. We also studied the contribution of the morphine-induced disinhibitory effect on the presynaptic glutamate release to the overall excitatory effect of morphine on VTA-DA neurons and related behavior. Our results suggest that the disinhibitory action of morphine on presynaptic glutamate release might be the main mechanism for morphine-induced increase in VTA-DA neuron firing and related behaviors.

Effects of porcine 25 kDa amelogenin and its proteolytic derivatives on bone sialoprotein expression.

PubMed

Nakayama, Y; Yang, L; Mezawa, M; Araki, S; Li, Z; Wang, Z; Sasaki, Y; Takai, H; Nakao, S; Fukae, M; Ogata, Y

2010-10-01

Amelogenins are hydrophobic proteins that are the major component of developing enamel. Enamel matrix derivative has been used for periodontal regeneration. Bone sialoprotein is an early phenotypic marker of osteoblast differentiation. In this study, we examined the ability of porcine amelogenins to regulate bone sialoprotein transcription. To determine the molecular basis of the transcriptional regulation of the bone sialoprotein gene by amelogenins, we conducted northern hybridization, transient transfection analyses and gel mobility shift assays using the osteoblast-like ROS 17/2.8 cells. Amelogenins (100 ng/mL) up-regulated bone sialoprotein mRNA at 3 h, with maximal mRNA expression occurring at 12 h (25 and 20 kDa) and 6 h (13 and 6 kDa). Amelogenins (100 ng/mL, 12 h) increased luciferase activities in pLUC3 (nucleotides -116 to +60), and 6 kDa amelogenin up-regulated pLUC4 (nucleotides -425 to +60) activity. The tyrosine kinase inhibitor inhibited amelogenin-induced luciferase activities, whereas the protein kinase A inhibitor abolished 25 kDa amelogenin-induced bone sialoprotein transcription. The effects of amelogenins were abrogated by 2-bp mutations in the fibroblast growth factor 2 response element (FRE). Gel-shift assays with radiolabeled FRE, homeodomain-protein binding site (HOX) and transforming growth factor-beta1 activation element (TAE) double-strand oligonucleotides revealed increased binding of nuclear proteins from amelogenin-stimulated ROS 17/2.8 cells at 3 h (25 and 13 kDa) and 6 h (20 and 6 kDa). These results demonstrate that porcine 25 kDa amelogenin and its proteolytic derivatives stimulate bone sialoprotein transcription by targeting FRE, HOX and TAE in the bone sialoprotein gene promoter, and that full-length amelogenin and amelogenin cleavage products are able to regulate bone sialoprotein transcription via different signaling pathways. (c) 2010 John Wiley & Sons A/S.

In Vitro Interactions between 17β-Estradiol and DNA Result in Formation of the Hormone-DNA Complexes

PubMed Central

Heger, Zbynek; Guran, Roman; Zitka, Ondrej; Beklova, Miroslava; Adam, Vojtech; Kizek, Rene

2014-01-01

Beyond the role of 17β-estradiol (E2) in reproduction and during the menstrual cycle, it has been shown to modulate numerous physiological processes such as cell proliferation, apoptosis, inflammation and ion transport in many tissues. The pathways in which estrogens affect an organism have been partially described, although many questions still exist regarding estrogens’ interaction with biomacromolecules. Hence, the present study showed the interaction of four oligonucleotides (17, 20, 24 and/or 38-mer) with E2. The strength of these interactions was evaluated using optical methods, showing that the interaction is influenced by three major factors, namely: oligonucleotide length, E2 concentration and interaction time. In addition, the denaturation phenomenon of DNA revealed that the binding of E2 leads to destabilization of hydrogen bonds between the nitrogenous bases of DNA strands resulting in a decrease of their melting temperatures (Tm). To obtain a more detailed insight into these interactions, MALDI-TOF mass spectrometry was employed. This study revealed that E2 with DNA forms non-covalent physical complexes, observed as the mass shifts for app. 270 Da (Mr of E2) to higher molecular masses. Taken together, our results indicate that E2 can affect biomacromolecules, as circulating oligonucleotides, which can trigger mutations, leading to various unwanted effects. PMID:25089777

Formation of low charge state ions of synthetic polymers using quaternary ammonium compounds.

PubMed

Nasioudis, Andreas; Joyce, William F; van Velde, Jan W; Heeren, Ron M A; van den Brink, Oscar F

2010-07-01

Factors such as high polymer dispersity and variation in elemental composition (of copolymers) often complicate the electrospray ionization mass spectrometry (ESI-MS) analysis of synthetic polymers with high molar mass. In the experiments described in this study, quaternary ammonium compounds were observed to facilitate the production of low charge state pseudomolecular ions when added to the spray solution for ESI-MS. This approach was then used for the ESI time-of-flight mass spectrometry (TOF-MS) analysis of synthetic polymers. Hexadecyltrimethylammonium chloride permitted the successful analysis of poly(ethylene glycol) of 2-40 kDa, poly(propylene glycol) and poly(tetramethylene glycol) oligomers. Increasing the quaternary ammonium compounds' concentration results in the production of low charge state pseudomolecular ions. A comparison of structurally different quaternary ammonium compounds showed that the best performance is expected from large molecules with specific charge localization, which leaves the charge available for interactions. The applicability of the method for the MS analysis of other polymeric systems was also studied. In the case of poly(tetramethylene glycol), the method not only shifted the distributions to higher m/z values but also allowed the detection of high molecular weight material that was not observed without addition of the modifier to the spray solution.

Simultaneous characterization of prenylated flavonoids and isoflavonoids in Psoralea corylifolia L. by liquid chromatography with diode-array detection and quadrupole time-of-flight mass spectrometry.

PubMed

Xu, Mei-Juan; Wu, Bin; Ding, Tao; Chu, Ji-Hong; Li, Chang-Yin; Zhang, Jun; Wu, Ting; Wu, Jian; Liu, Shi-Jia; Liu, Shen-Lin; Ju, Wen-Zheng; Li, Ping

2012-10-15

Prenylated flavonoids and isoflavonoids are widely distributed throughout the plant kingdom, with many biological effects. Psoralea corylifolia, which contains many kinds of prenylated components, has been widely used as a medicinal plant in Asia and India for thousands of years. The goal of this study was to characterize the components in P. corylifolia using a liquid chromatography with diode-array detection and quadrupole time-of-flight mass spectrometry (LC-DAD/Q-TOF-MS) method, and to elucidate the fragmentation behavior of the different prenyl substituent groups and their appropriate characteristic pathways in positive ion mode. The calculated accurate masses of the protonated molecules, the fragment ions, the retention behavior, and the data from UV spectra were used for identification of the components in P. corylifolia. A total of 45 compounds, including 43 prenylated components, were identified or tentatively identified in P. corylifolia. Different diagnostic fragment ions and neutral losses were observed in different prenyl substructures: neutral loss of 56 Da (C(4)H(8)) and a fragment ion at m/z 69 (C(5)H(9)(+)) were generated by a prenyl chain; neutral losses of 42 Da (C(3)H(6)), 54 Da (C(4)H(6)), 15 Da (CH(3•)) and 16 Da (CH(4)) were observed in a ring-closed prenyl group; neutral losses of 72 Da (C(4)H(8)O), 60 Da (C(2)H(4)O(2)), 58 Da (C(3)H(6)O) and 18 Da (H(2)O) were detected in a 2,2-dimethyl-3,4-dihydroxydihydropyran ring; neutral losses of 72 Da (C(4)H(8)O), 60 Da (C(3)H(8)O) and 18 Da (H(2)O) were yielded from a 2,2-dimethyl-3-hydroxydihydropyran ring, a 2-(1-hydroxy-1-methylethyl)dihydrofuran ring or a 1-hydroxy-3-methylbut-3-enyl chain. This method can be applied for analysis of prenylated components in P. corylifolia and other herbal medicines. Copyright © 2012 John Wiley & Sons, Ltd.

Mass and Charge Measurements on Heavy Ions

PubMed Central

Sugai, Toshiki

2017-01-01

The relationship between mass and charge has been a crucial topic in mass spectrometry (MS) because the mass itself is typically evaluated based on the m/z ratio. Despite the fact that this measurement is indirect, a precise mass can be obtained from the m/z value with a high m/z resolution up to 105 for samples in the low mass and low charge region under 10,000 Da and 20 e, respectively. However, the target of MS has recently been expanded to the very heavy region of Mega or Giga Da, which includes large particles and biocomplexes, with very large and widely distributed charge from kilo to Mega range. In this region, it is necessary to evaluate charge and mass simultaneously. Recent studies for simultaneous mass and charge observation and related phenomena are discussed in this review. PMID:29302406

Biochemical characterization of Aspergillus oryzae native tannase and the recombinant enzyme expressed in Pichia pastoris.

PubMed

Mizuno, Toshiyuki; Shiono, Yoshihito; Koseki, Takuya

2014-10-01

In this study, the biochemical properties of the recombinant tannase from Aspegillus oryzae were compared with those of the native enzyme. Extracellular native tannase was purified from a commercial enzyme source. Recombinant tannase highly expressed in Pichia pastoris was prepared as an active extracellular protein. Purified native and recombinant tannases produced smeared bands with apparent molecular masses of 45-80 kDa and 45-75 kDa, respectively, by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. After N-deglycosylation, the native enzyme yielded molecular masses of 33 kDa and 30 kDa, whereas the recombinant enzyme yielded molecular masses of 34 kDa and 30 kDa. Purified native and recombinant tannases had an optimum pH of 4.0-5.0 and 5.0, respectively, and were stable up to 40°C. After N-deglycosylation, both enzymes exhibited reduced thermostability. Catalytic efficiencies of both purified enzymes were greater with natural substrates, such as (-)-catechin, (-)-epicatechin, and (-)-epigallocatechin gallates, than those with synthetic substrates, such as methyl, ethyl, and propyl gallates. However, there were no activities against the methyl esters of ferulic, p-coumaric, caffeic, and sinapic acids, which indicate feruloyl esterase activity, or the ethyl, propyl, and butyl esters of 4-hydroxybenzoic acid, which indicate paraben hydrolase activity. Copyright © 2014 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

NADH:ubiquinone oxidoreductase from bovine heart mitochondria. cDNA sequences of the import precursors of the nuclear-encoded 39 kDa and 42 kDa subunits.

PubMed Central

Fearnley, I M; Finel, M; Skehel, J M; Walker, J E

1991-01-01

The 39 kDa and 42 kDa subunits of NADH:ubiquinone oxidoreductase from bovine heart mitochondria are nuclear-coded components of the hydrophobic protein fraction of the enzyme. Their amino acid sequences have been deduced from the sequences of overlapping cDNA clones. These clones were amplified from total bovine heart cDNA by means of the polymerase chain reaction, with the use of complex mixtures of oligonucleotide primers based upon fragments of protein sequence determined at the N-terminals of the proteins and at internal sites. The protein sequences of the 39 kDa and 42 kDa subunits are 345 and 320 amino acid residues long respectively, and their calculated molecular masses are 39,115 Da and 36,693 Da. Both proteins are predominantly hydrophilic, but each contains one or two hydrophobic segments that could possibly be folded into transmembrane alpha-helices. The bovine 39 kDa protein sequence is related to that of a 40 kDa subunit from complex I from Neurospora crassa mitochondria; otherwise, it is not related significantly to any known sequence, including redox proteins and two polypeptides involved in import of proteins into mitochondria, known as the mitochondrial processing peptidase and the processing-enhancing protein. Therefore the functions of the 39 kDa and 42 kDa subunits of complex I are unknown. The mitochondrial gene product, ND4, a hydrophobic component of complex I with an apparent molecular mass of about 39 kDa, has been identified in preparations of the enzyme. This subunit stains faintly with Coomassie Blue dye, and in many gel systems it is not resolved from the nuclearcoded 36 kDa subunit. Images Fig. 1. PMID:1832859

Native and subunit molecular mass and quarternary structure of the hemoglobin from the primitive branchiopod crustacean Triops cancriformis.

PubMed

Rousselot, Morgane; Jaenicke, Elmar; Lamkemeyer, Tobias; Harris, J Robin; Pirow, Ralph

2006-09-01

Many branchiopod crustaceans are endowed with extracellular, high-molecular-weight hemoglobins whose exact structural characteristics have remained a matter of conjecture. By using a broad spectrum of techniques, we provide precise and coherent information on the hemoglobin of one of the phylogenetically 'oldest' extant branchiopods, the tadpole shrimp Triops cancriformis. The hemoglobin dissociated under reducing conditions into two subunits, designated TcHbA and TcHbB, with masses of 35,775+/-4 and 36,055+/-4 Da, respectively, determined by ESI-MS. Nonreducing conditions showed only two disulfide-bridged dimers, a homodimer of TcHbA, designated D1 (71,548+/-5 Da), and the heterodimer D2 (71,828+/-5 Da). Carbamidomethylation of free SH groups revealed the presence of three cysteines per subunit and indicated one intrasubunit and one intersubunit disulfide bridge. Ultracentrifugation and light-scattering experiments under nondenaturating conditions yielded mass estimates that suggested an uneven number of 17 subunits forming the native hemoglobin. This unrealistic number resulted from the presence of two size classes (16-mer and 18-mer), which were recognized by native PAGE and Ferguson plot analysis. ESI-MS revealed three hemoglobin isoforms with masses of 588.1 kDa, 662.0 kDa, and 665.0 kDa. The 16-mer and the smaller 18-mer species are supposed to be composed of TcHbA only, given the dominance of this subunit type in SDS/PAGE. Transmission electron microscopy of negatively stained specimens showed a population of compact molecules with geometrical extensions of 14, 16 and 9 nm. The proposed stoichiometric model of quarternary structure provides the missing link to achieve a mechanistic understanding of the structure-function relationships among the multimeric arthropodan hemoglobins.

CN-GELFrEE - Clear Native Gel-eluted Liquid Fraction Entrapment Electrophoresis

PubMed Central

Skinner, Owen S.; Do Vale, Luis H. F.; Catherman, Adam D.; Havugimana, Pierre C.; Valle de Sousa, Marcelo; Domont, Gilberto B.; Kelleher, Neil L.; Compton, Philip D.

2016-01-01

Protein complexes perform an array of crucial cellular functions. Elucidating their non-covalent interactions and dynamics is paramount for understanding the role of complexes in biological systems. While the direct characterization of biomolecular assemblies has become increasingly important in recent years, native fractionation techniques that are compatible with downstream analysis techniques, including mass spectrometry, are necessary to further expand these studies. Nevertheless, the field lacks a high-throughput, wide-range, high-recovery separation method for native protein assemblies. Here, we present clear native gel-eluted liquid fraction entrapment electrophoresis (CN-GELFrEE), which is a novel separation modality for non-covalent protein assemblies. CN-GELFrEE separation performance was demonstrated by fractionating complexes extracted from mouse heart. Fractions were collected over 2 hr and displayed discrete bands ranging from ~30 to 500 kDa. A consistent pattern of increasing molecular weight bandwidths was observed, each ranging ~100 kDa. Further, subsequent reanalysis of native fractions via SDS-PAGE showed molecular-weight shifts consistent with the denaturation of protein complexes. Therefore, CN-GELFrEE was proved to offer the ability to perform high-resolution and high-recovery native separations on protein complexes from a large molecular weight range, providing fractions that are compatible with downstream protein analyses. PMID:26967310

Matrix-assisted laser desorption-ionization time-of-flight mass spectrometry in the subunit stoichiometry study of high-mass non-covalent complexes

NASA Astrophysics Data System (ADS)

Moniatte, M.; Lesieur, C.; Vecsey-Semjen, B.; Buckley, J. T.; Pattus, F.; van der Goot, F. G.; van Dorsselaer, A.

1997-12-01

This study explores the potential of MALDI-TOF MS for the mass measurement of large non-covalent protein complexes. The following non-covalent complexes have been investigated: aerolysin from Aeromonas hydrophila (335 kDa) and [alpha]-haemolysin from Staphylococcus aureus (233 kDa) which are both cytolytic toxins, three enzymes known to be homotetramers in solution: bovine liver catalase (235 kDa), rabbit muscle pyruvate kinase (232 kDa), yeast alcohol dehydrogenase (147 kDa) and finally a lectin, concanavalin A (102 kDa). Three different matrix preparations were systematically tested under various conditions: ferulic acid dissolved in THF, 2,6-dihydroxyacetophenone in 20 mM aqueous ammonium citrate and a two-step sample preparation with sinapinic acid. It was possible to find a suitable combination of matrix and preparation type which allowed the molecularity of all complexes tested to be deduced from the MALDI mass spectrum. Trimeric and tetrameric intermediates accumulating during the formation of the active heptameric aerolysin complex were also identified, this allowing a formation mechanism to be proposed. The observation of large specific non-covalent complexes has been found to be dependent on the choice of matrix, the type of sample preparation used, the solvent evaporation speed, the pH of the resulting matrix-sample mixture and the number of shots acquired on a given area. From this set of experiments, some useful guidelines for the observation of large complexes by MALDI could therefore be deduced. Fast evaporation of the solvent is particularly necessary in the case of pH sensitive complexes. An ESMS study on the same non-covalent complexes indicated that, rather surprisingly, reliable results could be obtained by MALDI-TOF MS on several very large complexes (above 200 kDa) for which ESMS yielded no clear spectra.

Electrospray ionisation mass spectrometry facilitates detection of fibrinogen (Bbeta 14 Arg --> Cys) mutation in a family with thrombosis.

PubMed

Brennan, S O; Hammonds, B; Spearing, R; George, P M

1997-12-01

We report the first direct detection of a fibrinogen mutation by electrospray ionisation mass spectrometry. The propositus, from a family with a history of thrombosis, came to attention after a pulmonary embolism subsequent to a spontaneous abortion. Prolonged thrombin (41 s) and reptilase times (26 s) together with an impairment of fibrinopeptide B release suggested a mutation at the thrombin cleavage site of the Bbeta chain. Direct mass analysis of purified fibrin chains from a thrombin induced clot showed that 50% of the Bbeta chains remained uncleaved. The measured mass of the mono sialo isoform of this uncleaved chain was 54150 Da, compared to a value of 54198 Da for normal Bbeta chains. This decrease of 48 Da in the intact protein is indicative of either a Bbeta 14 Arg to Cys, or Arg to Leu substitution. Heterozygosity for the Bbeta 14 Arg --> Cys mutation was verified by PCR amplification and DNA sequence analysis.

Onsite Gaseous Centrifuge Enrichment Plant UF6 Cylinder Destructive Analysis

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

Anheier, Norman C.; Cannon, Bret D.; Qiao, Hong

2012-07-17

The IAEA safeguards approach for gaseous centrifuge enrichment plants (GCEPs) includes measurements of gross, partial, and bias defects in a statistical sampling plan. These safeguard methods consist principally of mass and enrichment nondestructive assay (NDA) verification. Destructive assay (DA) samples are collected from a limited number of cylinders for high precision offsite mass spectrometer analysis. DA is typically used to quantify bias defects in the GCEP material balance. Under current safeguards measures, the operator collects a DA sample from a sample tap following homogenization. The sample is collected in a small UF6 sample bottle, then sealed and shipped under IAEAmore » chain of custody to an offsite analytical laboratory. Current practice is expensive and resource intensive. We propose a new and novel approach for performing onsite gaseous UF6 DA analysis that provides rapid and accurate assessment of enrichment bias defects. DA samples are collected using a custom sampling device attached to a conventional sample tap. A few micrograms of gaseous UF6 is chemically adsorbed onto a sampling coupon in a matter of minutes. The collected DA sample is then analyzed onsite using Laser Ablation Absorption Ratio Spectrometry-Destructive Assay (LAARS-DA). DA results are determined in a matter of minutes at sufficient accuracy to support reliable bias defect conclusions, while greatly reducing DA sample volume, analysis time, and cost.« less

Formation of multiply charged ions from large molecules using massive-cluster impact.

PubMed

Mahoney, J F; Cornett, D S; Lee, T D

1994-05-01

Massive-cluster impact is demonstrated to be an effective ionization technique for the mass analysis of proteins as large as 17 kDa. The design of the cluster source permits coupling to both magnetic-sector and quadrupole mass spectrometers. Mass spectra are characterized by the almost total absence of chemical background and a predominance of multiply charged ions formed from 100% glycerol matrix. The number of charge states produced by the technique is observed to range from +3 to +9 for chicken egg lysozyme (14,310 Da). The lower m/z values provided by higher charge states increase the effective mass range of analyses performed with conventional ionization by fast-atom bombardment or liquid secondary ion mass spectrometry.

Analysis of new growth promoting black market products.

PubMed

Krug, Oliver; Thomas, Andreas; Malerød-Fjeld, Helle; Dehnes, Yvette; Laussmann, Tim; Feldmann, Ingo; Sickmann, Albert; Thevis, Mario

2018-05-19

Detecting agents allegedly or evidently promoting growth such as human growth hormone (GH) or growth hormone releasing peptides (GHRP) in doping controls has represented a pressing issue for sports drug testing laboratories. While GH is a recombinant protein with a molecular weight of 22 kDa, the GHRPs are short (3-6 amino acids long) peptides with GH releasing properties. The endogenously produced GH (22 kDa isoform) consists of 191 amino acids and has a monoisotopic molecular mass of 22,124 Da. Within this study, a slightly modified form of GH was discovered consisting of 192 amino acids carrying an additional alanine at the N-terminus, leading to a monoisotopic mass of 22,195 Da. This was confirmed by top-down and bottom-up experiments using liquid chromatography coupled to high resolution/high accuracy mass spectrometry. Additionally, three analogues of GHRPs were identified as Gly-GHRP-6, Gly-GHRP-2 and Gly-Ipamorelin, representing the corresponding GHRP extended by a N-terminal glycine residue. The structure of these peptides was characterised by means of high resolution (tandem) mass spectrometry, and for Gly-Ipamorelin and Gly-GHRP-2 their identity was additionally confirmed by custom synthesis. Further, established in-vitro experiments provided preliminary information considering the potential metabolism after administration. Copyright © 2018. Published by Elsevier Ltd.

Isolation and Purification of Complex II from Proteus Mirabilis Strain ATCC 29245

PubMed Central

Shabbiri, Khadija; Ahmad, Waqar; Syed, Quratulain; Adnan, Ahmad

2010-01-01

A respiratory complex was isolated from plasma membrane of pathogenic Proteus mirabilis strain ATCC 29245. It was identified as complex II consisting of succinate:quinone oxidoreductase (EC 1.3.5.1) containing single heme b. The complex II was purified by ion-exchange chromatography and gel filtration. The molecular weight of purified complex was 116.5 kDa and it was composed of three subunits with molecular weights of 19 kDa, 29 kDa and 68.5 kDa. The complex II contained 9.5 nmoles of cytochrome b per mg protein. Heme staining indicated that the 19 kDa subunit was cytochrome b. Its reduced form showed absorptions peaks at 557.0, 524.8 and 424.4 nm. The α-band was shifted from 557.0 nm to 556.8 nm in pyridine ferrohemochrome spectrum. The succinate: quinone oxidoreductase activity was found to be high in this microorganism. PMID:24031557

High-Molecular-Mass Multi-c-Heme Cytochromes from Methylococcus capsulatus Bath†

PubMed Central

Bergmann, David J.; Zahn, James A.; DiSpirito, Alan A.

1999-01-01

The polypeptide and structural gene for a high-molecular-mass c-type cytochrome, cytochrome c553O, was isolated from the methanotroph Methylococcus capsulatus Bath. Cytochrome c553O is a homodimer with a subunit molecular mass of 124,350 Da and an isoelectric point of 6.0. The heme c concentration was estimated to be 8.2 ± 0.4 mol of heme c per subunit. The electron paramagnetic resonance spectrum showed the presence of multiple low spin, S = 1/2, hemes. A degenerate oligonucleotide probe synthesized based on the N-terminal amino acid sequence of cytochrome c553O was used to identify a DNA fragment from M. capsulatus Bath that contains occ, the gene encoding cytochrome c553O. occ is part of a gene cluster which contains three other open reading frames (ORFs). ORF1 encodes a putative periplasmic c-type cytochrome with a molecular mass of 118,620 Da that shows approximately 40% amino acid sequence identity with occ and contains nine c-heme-binding motifs. ORF3 encodes a putative periplasmic c-type cytochrome with a molecular mass of 94,000 Da and contains seven c-heme-binding motifs but shows no sequence homology to occ or ORF1. ORF4 encodes a putative 11,100-Da protein. The four ORFs have no apparent similarity to any proteins in the GenBank database. The subunit molecular masses, arrangement and number of hemes, and amino acid sequences demonstrate that cytochrome c553O and the gene products of ORF1 and ORF3 constitute a new class of c-type cytochrome. PMID:9922265

Deviations from Born-Oppenheimer mass scaling in spectroscopy and ultracold molecular physics

NASA Astrophysics Data System (ADS)

Lutz, Jesse J.; Hutson, Jeremy M.

2016-12-01

We investigate Born-Oppenheimer breakdown (BOB) effects (beyond the usual mass scaling) for the electronic ground states of a series of homonuclear and heteronuclear alkali-metal diatoms, together with the Sr2 and Yb2 diatomics. Several widely available electronic structure software packages are used to calculate the leading contributions to the total isotope shift for commonly occurring isotopologs of each species. Computed quantities include diagonal Born-Oppenheimer corrections (mass shifts) and isotopic field shifts. Mass shifts dominate for light nuclei up to and including K, but field shifts contribute significantly for Rb and Sr and are dominant for Yb. We compare the ab initio mass-shift functions for Li2, LiK and LiRb with spectroscopically derived ground-state BOB functions from the literature. We find good agreement in the values of the functions for LiK and LiRb at their equilibrium geometries, but significant disagreement with the shapes of the functions for all 3 systems. The differences may be due to contributions of nonadiabatic terms to the empirical BOB functions. We present a semiclassical model for the effect of BOB corrections on the binding energies of near-threshold states and the positions of zero-energy Feshbach resonances.

Limitations of Mass Spectrometry-Based Peptidomic Approaches

NASA Astrophysics Data System (ADS)

Fricker, Lloyd D.

2015-12-01

Mass spectrometry-based peptidomic approaches are powerful techniques to detect and identify the peptide content of biological samples. The present study investigated the limitations of peptidomic approaches using trimethylammonium butyrate isotopic tags to quantify relative peptide levels and Mascot searches to identify peptides. Data were combined from previous studies on human cell lines or mouse tissues. The combined databases contain 2155 unique peptides ranging in mass from 444 to 8765 Da, with the vast majority between 1 and 3 kDa. The amino acid composition of the identified peptides generally reflected the frequency in the Eukaryotic proteome with the exception of Cys, which was not present in any of the identified peptides in the free-SH form but was detected at low frequency as a disulfide with Cys residues, a disulfide with glutathione, or as S-cyanocysteine. To test if the low detection rate of peptides smaller than 500 Da, larger than 3 kDa, or containing Cys was a limitation of the peptidomics procedure, tryptic peptides of known proteins were processed for peptidomics using the same approach used for human cell lines and mouse tissues. The identified tryptic peptides ranged from 516 to 2418 Da, whereas the theoretical digest ranged from 217 to 7559 Da. Peptides with Cys were rarely detected and, if present, the Cys was usually modified S-cyanocysteine. Additionally, peptides with mono- and di-iodo Tyr and His were identified. Taken together, there are limitations of peptidomic techniques, and awareness of these limitations is important to properly use and interpret results.

Insulin stimulates the tyrosine phosphorylation of a 61-kilodalton protein in rat adipocytes.

PubMed

Mooney, R A; Bordwell, K L

1992-03-01

Insulin stimulated the tyrosine phosphorylation of a 61-kilodalton (kDa) protein in rat adipocytes prelabeled for 2 h with [32P]orthophosphate. Tyrosine phosphorylation of this 61-kDa protein displayed very similar insulin concentration dependency to receptor autophosphorylation and tyrosine phosphorylation of a high molecular mass receptor substrate of 160 kDa. Phosphorylation of the 61-kDa protein was very rapid with maximum labeling attained at 30 sec, paralleling that of the other two proteins. Phosphoamino acid analysis revealed that each of the insulin-responsive phosphoproteins contained phosphoserine as well as phosphotyrosine, though the ratio of two phosphoamino acids recovered from each protein differed. The 61-kDa protein yielded relatively equal proportions of phosphoserine and phosphotyrosine. In contrast, the insulin receptor yielded relatively more label on phosphotyrosine than phosphoserine, whereas label incorporated into the 160-kDa protein was recovered primarily on phosphoserine. Cleveland peptide maps using either Staphylococcus aureus V8 proteinase or chymotrypsin revealed no similarities between the 61-kDa protein and the other tyrosine phosphorylated proteins. With subcellular fractionation, the 160-kDa protein was found in equal proportions in the high speed pellet (100,000 g) and supernatant. The 61-kDa protein had a similar distribution to that of the 160-kDa protein but was also detected in the low speed pellet (10,000 g). The insulin receptor was localized to the low speed pellet. In summary, rat adipocytes contain an insulin-dependent phosphotyrosyl protein of 61 kDa which is distinct from the more prominent high molecular mass receptor substrate. This 61-kDa protein has characteristics consistent with it being a substrate for the insulin receptor tyrosine kinase.

Differences in outer membrane proteins of the lymphogranuloma venereum and trachoma biovars of Chlamydia trachomatis

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

Batteiger, B.E.; Jones, R.B.

1985-11-01

The lymphogranuloma venereum (LGV) and trachoma biovars of Chlamydia trachomatis exhibit differences in biological properties both in vivo and in vitro. To identify analogous biochemical differences, the authors studied the molecular charges of chlamydial outer membrane proteins (OMPs) by means of isoelectric focusing and nonequilibrium pH gradient electrophoresis. Analysis of proteins of whole elementary bodies biosynthetically labeled with L-(35S)cysteine revealed that most chlamydial proteins were neutral or acidic. The major OMPs (MOMPs) of all strains tested were acidic and had apparent isoelectric points (pIs) that varied within narrow limits despite differences in molecular mass of up to 3,000 daltons (Da).more » However, a low-molecular-mass cysteine-rich OMP analogous to that previously described for Chlamydia psittaci varied consistently in molecular mass (12,500 versus 12,000 Da) and pI (5.4 versus 6.9) between LGV strains and trachoma strains, respectively. OMPs with a molecular mass of 60,000 Da in the trachoma biovar strains had pIs in the 7.3 to 7.7 range. However, analogous OMPs in the LGV strains existed as a doublet with a molecular mass of about 60,000 Da. These data indicate substantial differences in biochemical characteristics of analogous OMPs in the LGV and trachoma biovars. Such differences are the first structural differences described between LGV and trachoma strains which support their distinction into separate biovars and may be related to some of their biological differences.« less

From Supercomputer Modeling to Highest Mass Resolution in FT-ICR.

PubMed

N Nikolaev, Evgene; N Vladimirov, Gleb; Jertz, Roland; Baykut, Gökhan

2013-01-01

Understanding of behavior of ion ensembles inside FT-ICR cell based on the computer simulation of ion motion gives rise to the new ideas of cell designs. The recently introduced novel FT-ICR cell based on a Penning ion trap with specially shaped excitation and detection electrodes prevents distortion of ion cyclotron motion phases (normally caused by non-ideal electric trapping fields) by averaging the trapping DC electric field during the ion motion in the ICR cell. Detection times of 5 min resulting in resolving power close to 40,000,000 have been reached for reserpine at m/z 609 at a magnetic field of only 7 Tesla. Fine structures of resolved 13Cn isotopic cluster groups could be measured for molecular masses up to 5.7 kDa (insulin) with resolving power of 4,000,000 at 7 Tesla. Based on resolved fine structure patterns atomic compositions can be directly determined using a new developed algorithm for fine structure processing. Mass spectra of proteins and multimers of proteins reaching masses up to 186 kDa (enolase tetramer) could be measured with isotopic resolution. For instance, at 7 Tesla resolving power of 800,000 was achieved for enolase dimer (96 kDa) and 500,000 for molecular masses above 100 kDa. Experimental data indicate that there is practically no limit for the resolving power of this ICR cell except by collisional damping in the ultrahigh vacuum chamber.

Indole-diterpenoid profiles of Claviceps paspali and Claviceps purpurea from high-resolution Fourier transform Orbitrap mass spectrometry.

PubMed

Uhlig, Silvio; Egge-Jacobsen, Wolfgang; Vrålstad, Trude; Miles, Christopher O

2014-07-30

The biological activities most commonly associated with indole-diterpenoids are tremorgenicity in mammals and toxicity in insects through modulation of ion channels. The neurotoxic effects of some analogues are the cause of syndromes such as 'ryegrass staggers' and 'Paspalum staggers' in cattle and sheep. Our purpose was to obtain and interpret mass spectra of some pure Claviceps-related indole-diterpenoids (paspaline, paspalinine, paxilline, paspalitrems A and B) to facilitate identification of related compounds for which standards were not available. C. paspali-infected Paspalum dilatatum as well as C. purpurea sclerotia obtained from infected Phalaris arundinacea were extracted and the extracts separated via liquid chromatography. Low- and high-resolution mass spectra were then obtained of known and potentially unknown indole-diterpenoids. At least 20 different indole-diterpenoids were detected in the C. paspali extract with molecular masses ranging from 405 Da (C28H40NO) to 517 Da (C32H40NO5). The C. purpurea sclerotia were shown to contain several indole-diterpenoids with molecular masses ranging from 405 Da (C28H40NO) to 419 Da (C28H38NO2). This study demonstrates for the first time that C. purpurea may also produce indole-diterpenoids. This might explain why grazing of Phalaris spp. is occasionally connected with a tremorgenic syndrome in cattle, called 'phalaris staggers'. Copyright © 2014 John Wiley & Sons, Ltd.

Method for the elucidation of the elemental composition of low molecular mass chemicals using exact masses of product ions and neutral losses: application to environmental chemicals measured by liquid chromatography with hybrid quadrupole/time-of-flight mass spectrometry.

PubMed

Suzuki, Shigeru; Ishii, Tetsuko; Yasuhara, Akio; Sakai, Shinichi

2005-01-01

A method for elucidating the elemental compositions of low molecular weight chemicals, based primarily on mass measurements made using liquid chromatography (LC) with time-of-flight mass spectrometry (TOFMS) and quadrupole/time-of-flight mass spectrometry (LC/QTOFMS), was developed and tested for 113 chemicals of environmental interest with molecular masses up to approximately 400 Da. As the algorithm incorporating the method is not affected by differences in the instrument used, or by the ionization method and other ionization conditions, the method is useful not only for LC/TOFMS, but also for all kinds of mass spectra measured with higher accuracy and precision (uncertainties of a few mDa) employing all ionization methods and on-line separation techniques. The method involves calculating candidate compositions for intact ionized molecules (ionized forms of the sample molecule that have lost or gained no more than a proton, i.e., [M+H](+) or [M-H](-)) as well as for fragment ions and corresponding neutral losses, and eliminating those atomic compositions for the molecules that are inconsistent with the corresponding candidate compositions of fragment ions and neutral losses. Candidate compositions were calculated for the measured masses of the intact ionized molecules and of the fragment ions and corresponding neutral losses, using mass uncertainties of 2 and 5 mDa, respectively. Compositions proposed for the ionized molecule that did not correspond to the sum of the compositions of a candidate fragment ion and its corresponding neutral loss were discarded. One, 2-5, 6-10, 11-20, and >20 candidate compositions were found for 65%, 39%, 1%, 1%, and 0%, respectively, for the 124 ionized molecules formed from the 113 chemicals tested (both positive and negative ions were obtained from 11 of the chemicals). However, no candidate composition was found for 2% of the test cases (i.e., 3 chemicals), for each of which the measured mass of one of the product ions was in error by 5-6.7 mDa.

Two Bioactive Molecular Weight Fractions of a Conditioned Medium Enhance RPE Cell Survival on Age-Related Macular Degeneration and Aged Bruch's Membrane

PubMed Central

Sugino, Ilene K.; Sun, Qian; Springer, Carola; Cheewatrakoolpong, Noounanong; Liu, Tong; Li, Hong; Zarbin, Marco A.

2016-01-01

Purpose To characterize molecular weight fractions of bovine corneal endothelial cell conditioned medium (CM) supporting retinal pigment epithelium (RPE) cell survival on aged and age-related macular degeneration (AMD) Bruch's membrane. Methods CM was subject to size separation using centrifugal filters. Retentate and filtrate fractions were tested for bioactivity by analyzing RPE survival on submacular Bruch's membrane of aged and AMD donor eyes and behavior on collagen I-coated tissue culture wells. Protein and peptide composition of active fractions was determined by mass spectrometry. Results Two bioactive fractions, 3-kDa filtrate and a 10-50–kDa fraction, were necessary for RPE survival on aged and AMD Bruch's membrane. The 3-kDa filtrate, but not the 10-50–kDa fraction, supported RPE growth on collagen 1‐coated tissue culture plates. Mass spectrometry of the 10-50–kDa fraction identified 175 extracellular proteins, including growth factors and extracellular matrix molecules. Transforming growth factor (TGF)β-2 was identified as unique to active CM. Peptides representing 29 unique proteins were identified in the 3-KDa filtrate. Conclusions These results indicate there is a minimum of two bioactive molecules in CM, one found in the 3-kDa filtrate and one in the 10-50–kDa fraction, and that bioactive molecules in both fractions must be present to ensure RPE survival on Bruch's membrane. Mass spectrometry analysis suggested proteins to test in future studies to identify proteins that may contribute to CM bioactivity. Translational Relevance Results of this study are the first steps in development of an adjunct to cell-based therapy to ensure cell transplant survival and functionality in AMD patients. PMID:26933521

Three-dimensional structure of nylon hydrolase and mechanism of nylon-6 hydrolysis.

PubMed

Negoro, Seiji; Shibata, Naoki; Tanaka, Yusuke; Yasuhira, Kengo; Shibata, Hiroshi; Hashimoto, Haruka; Lee, Young-Ho; Oshima, Shohei; Santa, Ryuji; Oshima, Shohei; Mochiji, Kozo; Goto, Yuji; Ikegami, Takahisa; Nagai, Keisuke; Kato, Dai-Ichiro; Takeo, Masahiro; Higuchi, Yoshiki

2012-02-10

We performed x-ray crystallographic analyses of the 6-aminohexanoate oligomer hydrolase (NylC) from Agromyces sp. at 2.0 Å-resolution. This enzyme is a member of the N-terminal nucleophile hydrolase superfamily that is responsible for the degradation of the nylon-6 industry byproduct. We observed four identical heterodimers (27 kDa + 9 kDa), which resulted from the autoprocessing of the precursor protein (36 kDa) and which constitute the doughnut-shaped quaternary structure. The catalytic residue of NylC was identified as the N-terminal Thr-267 of the 9-kDa subunit. Furthermore, each heterodimer is folded into a single domain, generating a stacked αββα core structure. Amino acid mutations at subunit interfaces of the tetramer were observed to drastically alter the thermostability of the protein. In particular, four mutations (D122G/H130Y/D36A/E263Q) of wild-type NylC from Arthrobacter sp. (plasmid pOAD2-encoding enzyme), with a heat denaturation temperature of T(m) = 52 °C, enhanced the protein thermostability by 36 °C (T(m) = 88 °C), whereas a single mutation (G111S or L137A) decreased the stability by ∼10 °C. We examined the enzymatic hydrolysis of nylon-6 by the thermostable NylC mutant. Argon cluster secondary ion mass spectrometry analyses of the reaction products revealed that the major peak of nylon-6 (m/z 10,000-25,000) shifted to a smaller range, producing a new peak corresponding to m/z 1500-3000 after the enzyme treatment at 60 °C. In addition, smaller fragments in the soluble fraction were successively hydrolyzed to dimers and monomers. Based on these data, we propose that NylC should be designated as nylon hydrolase (or nylonase). Three potential uses of NylC for industrial and environmental applications are also discussed.

Characterization of aromatic organosulfur model compounds relevant to fossil fuels by using atmospheric pressure chemical ionization with CS2 and high-resolution tandem mass spectrometry.

PubMed

Tang, Weijuan; Sheng, Huaming; Jin, Chunfen; Riedeman, James S; Kenttämaa, Hilkka I

2016-04-15

The chemistry of desulfurization involved in processing crude oil is greatly dependent on the forms of sulfur in the oil. Sulfur exists in different chemical bonding environments in fossil fuels, including those in thiophenes and benzothiophenes, thiols, sulfides, and disulfides. In this study, the fragmentation behavior of the molecular ions of 17 aromatic organosulfur compounds with various functionalities was systematically investigated by using high-resolution tandem mass spectrometry. Multiple-stage tandem mass spectrometric experiments were carried out using a linear quadrupole ion trap (LQIT) equipped with an atmospheric pressure chemical ionization (APCI) source. (+)APCI/CS2 was used to generate stable dominant molecular ions for all the compounds studied except for three sulfides that also showed abundant fragment ions. The LQIT coupled with an orbitrap mass spectrometer was used for elemental composition analysis, which facilitated the identification of the neutral molecules lost during fragmentation. The characteristic fragment ions generated in MS(2) and MS(3) experiments provide clues for the chemical bonding environment of sulfur atoms in the examined compounds. Upon collision-induced dissociation (CID), the molecular ions can lose the sulfur atom in a variety of ways, including as S (32 Da), HS(•) (33 Da), H2 S (34 Da), CS (44 Da), (•) CHS (45 Da) and CH2 S (46 Da). These neutral fragments are not only indicative of the presence of sulfur, but also of the type of sulfur present in the compound. Generally, losses of HS(•) and H2 S were found to be associated with compounds containing saturated sulfur functionalities, while losses of S, CS and (•) CHS were more common for heteroaromatic sulfur compounds. High-resolution tandem mass spectrometry with APCI/CS2 ionization is a viable approach to determining the types of organosulfur compounds. It can potentially be applied to analysis of complex mixtures, which is beneficial to improving the desulfurization process of fossil fuels. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

Nutrition and health in hotel staff on different shift patterns.

PubMed

Seibt, R; Süße, T; Spitzer, S; Hunger, B; Rudolf, M

2015-08-01

Limited research is available that examines the nutritional behaviour and health of hotel staff working alternating and regular shifts. To analyse the nutritional behaviour and health of employees working in alternating and regular shifts. The study used an ex post facto cross-sectional analysis to compare the nutritional behaviour and health parameters of workers with alternating shifts and regular shift workers. Nutritional behaviour was assessed with the Food Frequency Questionnaire. Body dimensions (body mass index, waist hip ratio, fat mass and active cell mass), metabolic values (glucose, triglyceride, total cholesterol and low- and high-density lipoprotein), diseases and health complaints were included as health parameters. Participants worked in alternating (n = 53) and regular shifts (n = 97). The average age of subjects was 35 ± 10 years. There was no significant difference in nutritional behaviour, most surveyed body dimensions or metabolic values between the two groups. However, alternating shift workers had significantly lower fat mass and higher active cell mass but nevertheless reported more pronounced health complaints. Sex and age were also confirmed as influencing the surveyed parameters. Shift-dependent nutritional problems were not conspicuously apparent in this sample of hotel industry workers. Health parameters did not show significantly negative attributes for alternating shift workers. Conceivably, both groups could have the same level of knowledge on the health effects of nutrition and comparable opportunities to apply this. Further studies on nutritional and health behaviour in the hotel industry are necessary in order to create validated screening programmes. © The Author 2015. Published by Oxford University Press on behalf of the Society of Occupational Medicine. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Purification and characterization of a p-coumarate decarboxylase and a vinylphenol reductase from Brettanomyces bruxellensis.

PubMed

Godoy, Liliana; Martínez, Claudio; Carrasco, Nelson; Ganga, María Angélica

2008-09-30

The presence of Brettanomyces bruxellensis has been correlated with an increase of phenolic aromas in wine. The production of these aromas results from the metabolization of cinnamic acids, present in the wine, to their ethyl derivatives. Hence, the participation of two enzymes has been proposed: a p-coumarate decarboxylase (CD) and a vinylphenol reductase (VR). Both enzymes were purified and characterized from B. bruxellensis. In denaturing conditions, the CD enzyme had a molecular mass of 21 kDa, while in native conditions its mass was 41 kDa. The optimal activity was obtained at a temperature of 40 degrees C and a pH of 6.0. For p-coumaric acid, the Km value and Vmax were 1.22+/-0.08 mM and 98+/-0.15 micromol/min mg, respectively. The VR enzyme had a molecular mass of 37 kDa in SDS-PAGE, while in natural conditions its mass was 118 kDa. The Km value was > 3.37+/-2.05 mM and its Vmax was 107.62+/-50.38 micromol/min mg for NADPH used as a cofactor. Both enzymatic activities were stable at pH 3.4, but in the presence of ethanol the CD activity decreased drastically while the VR activity was more stable. This is the first report that shows the presence of a CD and a VR enzyme in B. bruxellensis.

Inertial mass sensing with low Q-factor vibrating microcantilevers

NASA Astrophysics Data System (ADS)

Adhikari, S.

2017-10-01

Mass sensing using micromechanical cantilever oscillators has been established as a promising approach. The scientific principle underpinning this technique is the shift in the resonance frequency caused by the additional mass in the dynamic system. This approach relies on the fact that the Q-factor of the underlying oscillator is high enough so that it does not significantly affect the resonance frequencies. We consider the case when the Q-factor is low to the extent that the effect of damping is prominent. It is shown that the mass sensing can be achieved using a shift in the damping factor. We prove that the shift in the damping factor is of the same order as that of the resonance frequency. Based on this crucial observation, three new approaches have been proposed, namely, (a) mass sensing using frequency shifts in the complex plane, (b) mass sensing from damped free vibration response in the time domain, and (c) mass sensing from the steady-state response in the frequency domain. Explicit closed-form expressions relating absorbed mass with changes in the measured dynamic properties have been derived. The rationale behind each new method has been explained using non-dimensional graphical illustrations. The new mass sensing approaches using damped dynamic characteristics can expand the current horizon of micromechanical sensing by incorporating a wide range of additional measurements.

Charge detection mass spectrometry: Instrumentation & applications to viruses

NASA Astrophysics Data System (ADS)

Pierson, Elizabeth E.

For over three decades, electrospray ionization (ESI) has been used to ionize non-covalent complexes and subsequently transfer the intact ion into the gas phase for mass spectrometry (MS) analysis. ESI generates a distribution of multiple charged ions, resulting in an m/z spectrum comprised of a series of peaks, known as a charge state envelope. To obtain mass information, the number of charges for each peak must be deduced. For smaller biological analytes like peptides, the charge states are sufficiently resolved and this process is straightforward. For macromolecular complexes exceeding ~100 kDa, this process is complicated by the broadening and shifting of charge states due to incomplete desolvation, salt adduction, and inherent mass heterogeneity. As the analyte mass approaches the MDa regime, the m/z spectrum is often comprised of a broad distribution of unresolved charge states. In such cases, mass determination is precluded. Charge detection mass spectrometry (CDMS) is an emerging MS technique for determining the masses of heterogeneous, macromolecular complexes. In CDMS, the m/z and z of single ions are measured concurrently so that mass is easily calculated. With this approach, deconvolution of an m/z spectrum is unnecessary. This measurement is carried out by passing macroions through a conductive cylinder. The induced image charge on the cylindrical detector provides information about m/z and z: the m/z is related to its time-of-flight through the detector, and the z is related to the intensity of the image charge. We have applied CDMS to study the self-assembly of virus capsids. Late-stage intermediates in the assembly of hepatitis B virus, a devastating human pathogen, have been identified. This is the first time that such intermediates have been detected and represent a significant advancement towards understanding virus capsid assembly. CDMS has also been used to identify oversized, non-icosahedral polymorphs in the assembly of woodchuck hepatitis virus capsids. Finally, CDMS has been used to characterize the purity of adeno-associated viral vectors for potential gene therapy applications.

The 29-kDa proteins phosphorylated ion thrombin-activated human platelets are forms of the estrogen receptor-related 27-kDa heat shock protein

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

Mendelsohn, M.E.; Yan Zhu; O'Neill, S.

Thrombin plays a critical role in platelet activation, hemostasis, and thrombosis. Cellular activation by thrombin leads to the phosphorylation of multiple proteins, most of which are unidentified. The authors have characterized several 29-kDa proteins that are rapidly phosphorylated following exposure of intact human platelets to thrombin. A murine monoclonal antibody raised to an unidentified estrogen receptor-related 29-kDa protein selectively recognized these proteins as well as a more basic, unphosphorylated 27-kDa protein. Cellular activation by thrombin led to a marked shift in the proportion of protein from the 27-kDa unphosphorylated form to the 29-kDa phosphoprotein species. Using this antibody, they isolatedmore » and sequenced a human cDNA clone encoding a protein that was identical to the mammalian 27-kDa heat shock protein (HSP27), a protein of uncertain function that is known to be phosphorylated to several forms and to be transcriptionally induced by estrogen. The 29-kDa proteins were confirmed to be phosphorylated forms of HSP27 by immunoprecipitation studies. Thus, the estrogen receptor-related protein is HSP27, and the three major 20-kDa proteins phosphorylated in thrombin-activated platelets are forms of HSP27. These data suggest a role for HSP27 in the signal transduction events of platelet activation.« less

Identification and Characterization of Two Temperature-Induced Surface-Associated Proteins of Streptococcus suis with High Homologies to Members of the Arginine Deiminase System of Streptococcus pyogenes

PubMed Central

Winterhoff, Nora; Goethe, Ralph; Gruening, Petra; Rohde, Manfred; Kalisz, Henryk; Smith, Hilde E.; Valentin-Weigand, Peter

2002-01-01

The present study was performed to identify stress-induced putative virulence proteins of Streptococcus suis. For this, protein expression patterns of streptococci grown at 32, 37, and 42°C were compared by one- and two-dimensional gel electrophoresis. Temperature shifts from 32 and 37 to 42°C induced expression of two cell wall-associated proteins with apparent molecular masses of approximately 47 and 53 kDa. Amino-terminal sequence analysis of the two proteins indicated homologies of the 47-kDa protein with an ornithine carbamoyltransferase (OCT) from Streptococcus pyogenes and of the 53-kDa protein with the streptococcal acid glycoprotein (SAGP) from S. pyogenes, an arginine deiminase (AD) recently proposed as a putative virulence factor. Cloning and sequencing the genes encoding the putative OCT and AD of S. suis, octS and adiS, respectively, revealed that they had 81.2 (octS) and 80.2% (adiS) identity with the respective genes of S. pyogenes. Both genes belong to the AD system, also found in other bacteria. Southern hybridization analysis demonstrated the presence of the adiS gene in all 42 serotype 2 and 9 S. suis strains tested. In 9 of these 42 strains, selected randomly, we confirmed expression of the AdiS protein, homologous to SAGP, by immunoblot analysis using a specific antiserum against the SAGP of S. pyogenes. In all strains AD activity was detected. Furthermore, by immunoelectron microscopy using the anti-S. pyogenes SAGP antiserum we were able to demonstrate that the AdiS protein is expressed on the streptococcal surface in association with the capsular polysaccharides but is not coexpressed with them. PMID:12446626

Characteristic and antioxidant activity of retorted gelatin hydrolysates from cobia (Rachycentron canadum) skin.

PubMed

Yang, Jing-Iong; Ho, Hsin-Yi; Chu, Yuh-Jwo; Chow, Chau-Jen

2008-09-01

Alkali-pretreated cobia (Rachycentron canadum) skin was extracted in a retort (121°C) for 30min to obtain a retorted skin gelatin hydrolysate (RSGH). The molecular mass distributions and antioxidant activities of cobia RSGH and enzyme-treated RSGHs (ET-RSGHs) derived from bromelain, papain, pancreatin, and trypsin digestion were then characterized. The molecular mass distribution of the RSGH ranged mainly between 20,000 and 700Da and those of ET-RSGHs ranged between 6500 and 700Da. The DPPH (α,α-diphenyl-β-picrylhydrazyl) radical scavenging effects (%) of 10mg/ml of RSGH and 10mg/ml of the four ET-RSGHs were 55% and 51-61%, respectively. The lipid peroxidation inhibition (%) of RSGH and ET-RSGHs (10mg/ml) were 58% and 60-71% on the fifth day in a linoleic acid model system, respectively. The 3Kd-ET-RSGHs, obtained by using a series of centrifugal ultrafiltration filters (molecular weight cut-offs of 10, 5, and 3kDa done sequentially with decreasing pore size), exhibited dramatically improved antioxidant activity, with most of the molecular mass ranging below 700Da. Compared to 10mg/ml of the RSGH, 10mg/ml of 3Kd-ET-RSGHs exhibited 45-65% more scavenging of DPPH radical and 24-38% more inhibition of lipid peroxidation. The peptides with molecular masses below 700Da in the ET-RSGHs or 3Kd-ET-RSGHs significantly affect the antioxidant properties. These peptides are composed of a small number of amino acids or free amino acids and have the potential to be added as antioxidants in foods. Copyright © 2008 Elsevier Ltd. All rights reserved.

Light-meson masses in an unquenched quark model

NASA Astrophysics Data System (ADS)

Chen, Xiaoyun; Ping, Jialun; Roberts, Craig D.; Segovia, Jorge

2018-05-01

We perform a coupled-channels calculation of the masses of light mesons with the quantum numbers I JP =-, (I ,J )=0 , 1, by including q q ¯ and (q q ¯)2 components in a nonrelativistic chiral quark model. The coupling between two- and four-quark configurations is realized through a 3P0 quark-pair creation model. With the usual form of this operator, the mass shifts are large and negative, an outcome which raises serious issues of validity for the quenched quark model. Herein, therefore, we introduce some improvements of the 3P0 operator in order to reduce the size of the mass shifts. By introducing two simple factors, physically well motivated, the coupling between q q ¯ and (q q ¯)2 components is weakened, producing mass shifts that are around 10%-20% of hadron bare masses.

Direct Midbrain Dopamine Input to the Suprachiasmatic Nucleus Accelerates Circadian Entrainment.

PubMed

Grippo, Ryan M; Purohit, Aarti M; Zhang, Qi; Zweifel, Larry S; Güler, Ali D

2017-08-21

Dopamine (DA) neurotransmission controls behaviors important for survival, including voluntary movement, reward processing, and detection of salient events, such as food or mate availability. Dopaminergic tone also influences circadian physiology and behavior. Although the evolutionary significance of this input is appreciated, its precise neurophysiological architecture remains unknown. Here, we identify a novel, direct connection between the DA neurons of the ventral tegmental area (VTA) and the suprachiasmatic nucleus (SCN). We demonstrate that D1 dopamine receptor (Drd1) signaling within the SCN is necessary for properly timed resynchronization of activity rhythms to phase-shifted light:dark cycles and that elevation of DA tone through selective activation of VTA DA neurons accelerates photoentrainment. Our findings demonstrate a previously unappreciated role for direct DA input to the master circadian clock and highlight the importance of an evolutionarily significant relationship between the circadian system and the neuromodulatory circuits that govern motivational behaviors. Copyright © 2017 Elsevier Ltd. All rights reserved.

Effects of repeated treatment with MDMA on working memory and behavioural flexibility in mice.

PubMed

Viñals, Xavier; Maldonado, Rafael; Robledo, Patricia

2013-03-01

Repeated administration of 3,4-methylenedioxymethamphetamine (MDMA) produces dopaminergic neurotoxicity in mice. However, it is still not clear whether this exposure induces deficits in cognitive processing related to specific subsets of executive functioning. We evaluated the effects of neurotoxic and non-neurotoxic doses of MDMA (0, 3 and 30 mg/kg, twice daily for 4 days) on working memory and attentional set-shifting in mice, and changes in extracellular levels of dopamine (DA) in the striatum. Treatment with MDMA (30 mg/kg) disrupted performance of acquired operant alternation, and this impairment was still apparent 5 days after the last drug administration. Decreased alternation was not related to anhedonia because no differences were observed between groups in the saccharin preference test under similar experimental conditions. Correct responding on delayed alternation was increased 1 day after repeated treatment with MDMA (30 mg/kg), probably because of general behavioural quiescence. Notably, the high dose regimen of MDMA impaired attentional set-shifting related to an increase in total perseveration errors. Finally, basal extracellular levels of DA in the striatum were not modified in mice repeatedly treated with MDMA with respect to controls. However, an acute challenge with MDMA (10 mg/kg) failed to increase DA outflow in mice receiving the highest MDMA dose (30 mg/kg), corroborating a decrease in the functionality of DA transporters. Seven days after this treatment, the effects of MDMA on DA outflow were recovered. These results suggest that repeated neurotoxic doses of MDMA produce lasting impairments in recall of alternation behaviour and reduce cognitive flexibility in mice. © 2012 The Authors, Addiction Biology © 2012 Society for the Study of Addiction.

Distributed data analysis in ATLAS

NASA Astrophysics Data System (ADS)

Nilsson, Paul; Atlas Collaboration

2012-12-01

Data analysis using grid resources is one of the fundamental challenges to be addressed before the start of LHC data taking. The ATLAS detector will produce petabytes of data per year, and roughly one thousand users will need to run physics analyses on this data. Appropriate user interfaces and helper applications have been made available to ensure that the grid resources can be used without requiring expertise in grid technology. These tools enlarge the number of grid users from a few production administrators to potentially all participating physicists. ATLAS makes use of three grid infrastructures for the distributed analysis: the EGEE sites, the Open Science Grid, and Nordu Grid. These grids are managed by the gLite workload management system, the PanDA workload management system, and ARC middleware; many sites can be accessed via both the gLite WMS and PanDA. Users can choose between two front-end tools to access the distributed resources. Ganga is a tool co-developed with LHCb to provide a common interface to the multitude of execution backends (local, batch, and grid). The PanDA workload management system provides a set of utilities called PanDA Client; with these tools users can easily submit Athena analysis jobs to the PanDA-managed resources. Distributed data is managed by Don Quixote 2, a system developed by ATLAS; DQ2 is used to replicate datasets according to the data distribution policies and maintains a central catalog of file locations. The operation of the grid resources is continually monitored by the Ganga Robot functional testing system, and infrequent site stress tests are performed using the Hammer Cloud system. In addition, the DAST shift team is a group of power users who take shifts to provide distributed analysis user support; this team has effectively relieved the burden of support from the developers.

Seasonal photosynthetic gas exchange and water-use efficiency in a constitutive CAM plant, the giant saguaro cactus (Carnegiea gigantea).

PubMed

Bronson, Dustin R; English, Nathan B; Dettman, David L; Williams, David G

2011-11-01

Crassulacean acid metabolism (CAM) and the capacity to store large quantities of water are thought to confer high water use efficiency (WUE) and survival of succulent plants in warm desert environments. Yet the highly variable precipitation, temperature and humidity conditions in these environments likely have unique impacts on underlying processes regulating photosynthetic gas exchange and WUE, limiting our ability to predict growth and survival responses of desert CAM plants to climate change. We monitored net CO(2) assimilation (A(net)), stomatal conductance (g(s)), and transpiration (E) rates periodically over 2 years in a natural population of the giant columnar cactus Carnegiea gigantea (saguaro) near Tucson, Arizona USA to investigate environmental and physiological controls over carbon gain and water loss in this ecologically important plant. We hypothesized that seasonal changes in daily integrated water use efficiency (WUE(day)) in this constitutive CAM species would be driven largely by stomatal regulation of nighttime transpiration and CO(2) uptake responding to shifts in nighttime air temperature and humidity. The lowest WUE(day) occurred during time periods with extreme high and low air vapor pressure deficit (D(a)). The diurnal with the highest D(a) had low WUE(day) due to minimal net carbon gain across the 24 h period. Low WUE(day) was also observed under conditions of low D(a); however, it was due to significant transpiration losses. Gas exchange measurements on potted saguaro plants exposed to experimental changes in D(a) confirmed the relationship between D(a) and g(s). Our results suggest that climatic changes involving shifts in air temperature and humidity will have large impacts on the water and carbon economy of the giant saguaro and potentially other succulent CAM plants of warm desert environments.

The Cuban scorpion Rhopalurus junceus (Scorpiones, Buthidae): component variations in venom samples collected in different geographical areas

PubMed Central

2013-01-01

Backgound The venom of the Cuban scorpion Rhopalurus junceus is poorly study from the point of view of their components at molecular level and the functions associated. The purpose of this article was to conduct a proteomic analysis of venom components from scorpions collected in different geographical areas of the country. Results Venom from the blue scorpion, as it is called, was collected separately from specimens of five distinct Cuban towns (Moa, La Poa, Limonar, El Chote and Farallones) of the Nipe-Sagua-Baracoa mountain massif and fractionated by high performance liquid chromatography (HPLC); the molecular masses of each fraction were ascertained by mass spectrometry analysis. At least 153 different molecular mass components were identified among the five samples analyzed. Molecular masses varied from 466 to 19755 Da. Scorpion HPLC profiles differed among these different geographical locations and the predominant molecular masses of their components. The most evident differences are in the relative concentration of the venom components. The most abundant components presented molecular weights around 4 kDa, known to be K+-channel specific peptides, and 7 kDa, known to be Na+-channel specific peptides, but with small molecular weight differences. Approximately 30 peptides found in venom samples from the different geographical areas are identical, supporting the idea that they all probably belong to the same species, with some interpopulational variations. Differences were also found in the presence of phospholipase, found in venoms from the Poa area (molecular weights on the order of 14 to 19 kDa). The only ubiquitous enzyme identified in the venoms from all five localities studied (hyaluronidase) presented the same 45 kD molecular mass, identified by gel electrophoresis analysis. Conclusions The venom of these scorpions from different geographical areas seem to be similar, and are rich in peptides that have of the same molecular masses of the peptides purified from other scorpions that affect ion-channel functions. PMID:23849540

Insecticidal components from field pea extracts: sequences of some variants of pea albumin 1b.

PubMed

Taylor, Wesley G; Sutherland, Daniel H; Olson, Douglas J H; Ross, Andrew R S; Fields, Paul G

2004-12-15

Methanol soluble insecticidal peptides with masses of 3752, 3757, and 3805 Da, isolated from crude extracts (C8 extracts) derived from the protein-enriched flour of commercial field peas [Pisum sativum (L.)], were purified by reversed phase chromatography and, after reduction and alkylation, were sequenced by matrix-assisted laser desorption/ionization (MALDI) time-of-flight mass spectrometry with the aid of various peptidases. These major peptides were variants of pea albumin 1b (PA1b) with methionine sulfoxide rather than methionine at position 12. Peptide 3752 showed additional variations at positions 29 (valine for isoleucine) and 34 (histidine for asparagine). A minor, 37 amino acid peptide with a molecular mass of 3788 Da was also sequenced and differed from a known PA1b variant at positions 1, 25, and 31. Sequence variants of PA1b with their molecular masses were compiled, and variants that matched the accurate masses of the experimental peptides were used to narrow the search. MALDI postsource decay experiments on pronase fragments helped to confirm the sequences. Whole and dehulled field peas gave insecticidal C8 extracts in the laboratory that were enriched in peptides with masses of 3736, 3741, and 3789 Da, as determined by high-performance liquid chromatography (HPLC) and electrospray ionization mass spectrometry. It was therefore concluded that oxidation of the methionine residues to methionine sulfoxide occurred primarily during the processing of dehulled peas in a mill.

Detection of Large Ions in Time-of-Flight Mass Spectrometry: Effects of Ion Mass and Acceleration Voltage on Microchannel Plate Detector Response

NASA Astrophysics Data System (ADS)

Liu, Ranran; Li, Qiyao; Smith, Lloyd M.

2014-08-01

In time-of-flight mass spectrometry (TOF-MS), ion detection is typically accomplished by the generation and amplification of secondary electrons produced by ions colliding with a microchannel plate (MCP) detector. Here, the response of an MCP detector as a function of ion mass and acceleration voltage is characterized, for singly charged peptide/protein ions ranging from 1 to 290 kDa in mass, and for acceleration voltages from 5 to 25 kV. A nondestructive inductive charge detector (ICD) employed in parallel with MCP detection provides a reliable reference signal to allow accurate calibration of the MCP response. MCP detection efficiencies were very close to unity for smaller ions at high acceleration voltages (e.g., angiotensin, 1046.5 Da, at 25 kV acceleration voltage), but decreased to ~11% for the largest ions examined (immunoglobulin G (IgG) dimer, 290 kDa) even at the highest acceleration voltage employed (25 kV). The secondary electron yield γ (average number of electrons produced per ion collision) is found to be proportional to mv3.1 (m: ion mass, v: ion velocity) over the entire mass range examined, and inversely proportional to the square root of m in TOF-MS analysis. The results indicate that although MCP detectors indeed offer superlative performance in the detection of smaller peptide/protein species, their performance does fall off substantially for larger proteins, particularly under conditions of low acceleration voltage.

Determination of Hyaluronan Molecular Mass Distribution in Human Breast Milk

PubMed Central

Yuan, Han; Amin, Ripal; Ye, Xin; De La Motte, Carol A.; Cowman, Mary K.

2015-01-01

Hyaluronan (HA) in human milk mediates host responses to microbial infection, via TLR4- and CD44-dependent signaling. Signaling by HA is generally size-specific. Because pure HA with average molecular mass (M) of 35 kDa can elicit a protective response in intestinal epithelial cells, it has been proposed that human milk HA may have a bioactive low M component. Here we report the size distribution of HA in human milk samples from twenty unique donors. A new method for HA analysis, employingion exchange (IEX) chromatography to fractionate HA by size, and specific quantification of each size fraction by competitive Enzyme Linked Sorbent Assay (ELSA), was developed. When separated into four fractions, milk HA with M ≤ 20 kDa, M ≈20-60 kDa, and M ≈ 60-110 kDa comprised an average of 1.5%, 1.4% and 2% of the total HA, respectively. The remaining 95% was HA with M≥110 kDa. Electrophoretic analysis of the higher M HA from thirteen samples showed nearly identical M distributions, with an average M of ∼440 kDa. This higher M HA component in human milk is proposed to bind to CD44 and to enhance human beta defensin 2 (HBD2) induction by the low M HA components. PMID:25579786

High Speed Intact Protein Characterization Using 4X Frequency Multiplication, Ion Trap Harmonization, and 21 Tesla FTICR-MS.

PubMed

Shaw, Jared B; Gorshkov, Mikhail V; Wu, Qinghao; Paša-Tolić, Ljiljana

2018-05-01

Mass spectrometric characterization of large biomolecules, such as intact proteins, requires the specificity afforded by ultrahigh resolution mass measurements performed at both the intact mass and product ion levels. Although the performance of time-of-flight mass analyzers is steadily increasing, the choice of mass analyzer for large biomolecules (e.g., proteins >50 kDa) is generally limited to the Fourier transform family of mass analyzers such as Orbitrap and ion cyclotron resonance (FTICR-MS), with the latter providing unmatched mass resolving power and measurement accuracy. Yet, protein analyses using FTMS are largely hindered by the low acquisition rates of spectra with ultrahigh resolving power. Frequency multiple detection schemes enable FTICR-MS to overcome this fundamental barrier and achieve resolving powers and acquisition speeds 4× greater than the limits imposed by magnetic field strength. Here we expand upon earlier work on the implementation of this technique for biomolecular characterization. We report the coupling of 21T FTICR-MS, 4X frequency multiplication, ion trapping field harmonization technology, and spectral data processing methods to achieve unprecedented acquisition rates and resolving power in mass spectrometry of large intact proteins. Isotopically resolved spectra of multiply charged ubiquitin ions were acquired using detection periods as short as 12 ms. Large proteins such as apo-transferrin (MW = 78 kDa) and monoclonal antibody (MW = 150 kDa) were isotopically resolved with detection periods of 384 and 768 ms, respectively. These results illustrate the future capability of accurate characterization of large proteins on time scales compatible with online separations.

The first trimeric Galanthus nivalis agglutinin-related lectin of Orchidaceae was found in Dendrobium pendulum: purification, characterization, and effects of stress factors.

PubMed

Siripipatthana, Patthraporn; Phaonakrop, Narumon; Roytrakul, Sittiruk; Senawong, Gulsiri; Mudalige-Jayawickrama, Rasika G; Sattayasai, Nison

2015-07-01

Trimeric Galanthus nivalis agglutinin-related lectin of Orchidaceae with two conformational forms was first studied in Dendrobium pendulum . It was highly expressed by stress factors. Using mannan-agarose column chromatography, a mannose-binding protein was purified from Dendrobium pendulum Roxb. pseudobulb. After heating in the presence of sodium dodecyl sulfate (SDS) with or without 2-mercaptoethanol, the protein showed one band with molecular mass of 14.0 kDa on SDS-polyacrylamide gel electrophoresis (PAGE). Without heating, three bands were found at positions of 14.0, 39.4, and 41.5 kDa, but a higher amount of 39.4 and 41.5 kDa protein bands were seen in the presence of 2-mercaptoethanol. Liquid chromatography-tandem mass spectrometry and database search indicated that the 14.0 kDa protein band contained three peptide fragments identical to parts of a lectin precursor from Dendrobiu m findleyanum Parish & Rchb.f. Native-PAGE and Ferguson plot showed that the purified protein had two native forms with molecular masses of 44.2 and 45.3 kDa, indicating three 14.0 kDa polypeptide subunits. The purified protein exhibited the agglutination activity with trypsinized chicken erythrocytes. It was then recognized as a Galanthus nivalis agglutinin-related lectin and named D. pendulum agglutinin (DPA). Using reverse transcription-polymerase chain reaction and DNA sequencing, the deduced amino acid sequence of DPA precursor showed the highest homology (96.4%) with a lectin precursor of D. findleyanum and contained three mannose-binding sites. Greater amounts of DPA were found when the pseudobulbs were treated with stress factors including ultraviolet light, abscisic acid, hydrogen peroxide, and acetylene gas.

Nanoscale mass detection based on vibrating piezoelectric ultrathin films under thermo-electro-mechanical loads

NASA Astrophysics Data System (ADS)

Asemi, H. R.; Asemi, S. R.; Farajpour, A.; Mohammadi, M.

2015-04-01

The potential applications of piezoelectric nanofilms (PNFs) and double-piezoelectric-nanofilm (DPNF) systems as nanoelectromechanical mass sensors are examined. The PNFs carrying multiple nanoparticles at arbitrary locations are modeled as rectangular nonlocal plates with attached concentrated masses. Using the nonlocal elasticity theory and Hamilton's principle, the differential equations of motion are derived for both PNF-based and DPNF-based nanosensors. The influences of small scale, initial stress and temperature change on the frequency shifts of the nanoelectromechanical sensors are taken into consideration. Explicit expressions are derived for the resonance frequencies of the nanosensors by employing the Galerkin method. The present results show that when the value of nonlocal parameter decreases, the frequency shifts of piezoelectric nanosensors increase. Further, the frequency shifts of DPNF-based mass sensors are always greater than those of PNF-based mass sensors. The present work would be helpful in the design of nanoelectromechanical mass sensors using PNFs.

Light-meson masses in an unquenched quark model

DOE PAGES

Chen, Xiaoyun; Ping, Jialun; Roberts, Craig D.; ...

2018-05-17

We perform a coupled-channels calculation of the masses of light mesons with the quantum numbers IJ P=-, (I,J) = 0,1, by includingmore » $$q\\bar{q}$$ and ($$q\\bar{q}$$) 2 components in a nonrelativistic chiral quark model. The coupling between two- and four-quark configurations is realized through a 3P 0 quark-pair creation model. With the usual form of this operator, the mass shifts are large and negative, an outcome which raises serious issues of validity for the quenched quark model. Therefore, we introduce some improvements of the 3P 0 operator in order to reduce the size of the mass shifts. By introducing two simple factors, physically well motivated, the coupling between $$q\\bar{q}$$ and ($$q\\bar{q}$$) 2 components is weakened, producing mass shifts that are around 10%–20% of hadron bare masses.« less

Light-meson masses in an unquenched quark model

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

Chen, Xiaoyun; Ping, Jialun; Roberts, Craig D.

We perform a coupled-channels calculation of the masses of light mesons with the quantum numbers IJ P=-, (I,J) = 0,1, by includingmore » $$q\\bar{q}$$ and ($$q\\bar{q}$$) 2 components in a nonrelativistic chiral quark model. The coupling between two- and four-quark configurations is realized through a 3P 0 quark-pair creation model. With the usual form of this operator, the mass shifts are large and negative, an outcome which raises serious issues of validity for the quenched quark model. Therefore, we introduce some improvements of the 3P 0 operator in order to reduce the size of the mass shifts. By introducing two simple factors, physically well motivated, the coupling between $$q\\bar{q}$$ and ($$q\\bar{q}$$) 2 components is weakened, producing mass shifts that are around 10%–20% of hadron bare masses.« less

Isolation and Characterization of Acetylated Derivative of Recombinant Insulin Lispro Produced in Escherichia coli.

PubMed

Szewczak, Joanna; Bierczyńska-Krzysik, Anna; Piejko, Marcin; Mak, Paweł; Stadnik, Dorota

2015-07-01

Insulin lispro is a rapid-acting insulin analogue produced by recombinant DNA technology. As a biosynthetic drug, the protein undergoes strict monitoring aiming for detection and characterization of impurities. The goal of this study was to isolate and identify a derivative of insulin lispro formed during biosynthesis. For this purpose, ion exchange chromatography in combination with endoproteinase Glu-C digestion, MALDI-TOF/TOF mass spectrometry and Edman sequencing were employed. Ion exchange chromatography analysis of related proteins in development batches of recombinant insulin lispro revealed the existence of unknown derivative in excess of the assumed limit. Its molecular mass was 42 Da higher than the theoretical mass of Lys(B31) insulin lispro--one of the expected process-related intermediates. Endoproteinase Glu-C cleavage enabled indication of the modified peptide. Tandem mass spectrometry (MS/MS) allowed to explore the location and type of the modification. The 42 amu shift was present in the mass of y-type ions, while b-type ions were in agreement with theoretical values. It suggested that the modification is present on B31 lysine. Further inquiry revealed the presence of two diagnostic ions for lysine acetylation at m/z 143.1 and 126.1. In addition, the peptide was isolated and sequenced by Edman degradation. Standards of phenylthiohydantoin derivatives of N-ε-acetyl-L-lysine and N-ε-trimethyl-L-lysine, not available commercially, were synthesized in the laboratory. The retention time of the modified residue confirmed its identity as N-ε-acetyl-L-lysine. The derivative of insulin lispro formed during biosynthesis of the drug was identified to be N-ε-acetyl-L-lysine (B31) insulin lispro.

Frequency Shift During Mass Properties Testing Using Compound Pendulum Method

NASA Technical Reports Server (NTRS)

Wolfe, David; Regan, Chris

2012-01-01

During mass properties testing on the X-48B Blended Wing Body aircraft (The Boeing Company, Chicago, Illinois) at the National Aeronautics and Space Administration Dryden Flight Research Center, Edwards, California, large inertia measurement errors were observed in results from compound pendulum swings when compared to analytical models. By comparing periods of oscillations as measured from an average over the test period versus the period of each oscillation, it was noticed that the frequency of oscillation was shifting significantly throughout the test. This phenomenon was only noticed during compound pendulum swings, and not during bifilar pendulum swings. The frequency shift was only visible upon extensive data analysis of the frequency for each oscillation, and did not appear in averaged frequency data over the test period. Multiple test articles, test techniques, and hardware setups were used in attempts to eliminate or identify the cause of the frequency shift. Plotting the frequency of oscillation revealed a region of minimal shift that corresponded to a larger amplitude range. This region of minimal shift provided the most accurate results compared to a known test article; however, the amplitudes that produce accurate inertia measurements are amplitudes larger than those generally accepted in mass properties testing. This paper examines two case studies of the frequency shift, using mass properties testing performed on a dummy test article, and on the X-48B Blended Wing Body aircraft.

Changing palaeoenvironments and tetrapod populations in the Daptocephalus Assemblage Zone (Karoo Basin, South Africa) indicate early onset of the Permo-Triassic mass extinction

NASA Astrophysics Data System (ADS)

Viglietti, Pia A.; Smith, Roger M. H.; Rubidge, Bruce S.

2018-02-01

Important palaeoenvironmental differences are identified during deposition of the latest Permian Daptocephalus Assemblage Zone (DaAZ) of the South African Beaufort Group (Karoo Supergoup), which is also divided into a Lower and Upper subzone. A lacustrine floodplain facies association showing evidence for higher water tables and subaqueous conditions on the floodplains is present in Lower DaAZ. The change to well-drained floodplain facies association in the Upper DaAZ is coincident with a faunal turnover as evidenced by the last appearance of the dicynodont Dicynodon lacerticeps, the therocephalian Theriognathus microps, the cynodont Procynosuchus delaharpeae, and first appearance of the dicynodont Lystrosaurus maccaigi within the Ripplemead member. Considering the well documented 3-phased extinction of Karoo tetrapods during the Permo-Triassic Mass Extinction (PTME), the facies transition between the Lower and Upper DaAZ represents earlier than previously documented palaeoenvironmental changes associated with the onset of this major global biotic crisis.

Neuroendocrine factors affecting the glycogen metabolism of purified Mytilus edulis glycogen cells: partial characterization of the putative glycogen mobilization hormone--demonstration of a factor that stimulates glycogen synthesis.

PubMed

Robbins, I; Lenoir, F; Mathieu, M

1991-04-01

A putative glycogen mobilizing hormone (GMH) from the marine mussel Mytilus edulis L. has been partially characterized. GMH activity is present in the cerebral ganglia and the hemolymph serum and promotes the mobilization of glycogen in isolated glycogen cells. The cerebral GMH is trypsin sensitive and partially heat labile and has an apparent molecular mass of greater than 20 kDa. Following fractionation of cerebral extracts by molecular mass, a second factor, with a molecular mass of ca. 1.5 kDa, was discovered. This factor stimulates post-incubation incorporation of 14C into glycogen in isolated glycogen cells.

Yolk proteins during ovary and egg development of mature female freshwater crayfish (Cherax quadricarinatus).

PubMed

Serrano-Pinto, Vania; Vazquez-Boucard, Celia; Villarreal-Colmenares, Humberto

2003-01-01

Vitellins from ovaries and eggs at different stages of development in freshwater crayfish (Cherax quadricarinatus) were examined by chromatography, PAGE and SDS-PAGE. With these methods, two forms of vitellin (Vt1 and Vt2) were observed in ovaries and eggs (stages I and V). In ovaries in secondary vitellogenesis, native molecular mass was 470 (Vt1) and 440 (Vt2) kDa. The electrophoretic pattern of the eggs proved to be more complex. The protein molecular mass depend on the development stage of the egg: stage I, 650 kDa (Vt1) and 440 kDa (Vt2); stage V, 390 kDa (Vt1) and 340 kDa (Vt2). The identified vitellins appear to be lipo-glycocarotenoprotein. A similar vitellin polypeptide composition was observed in the two forms of vitellin from ovaries and eggs in stage V. In ovaries the SDS-PAGE analysis showed four subunits with molecular weights of approximately 180, 120, 95 and 80 kDa (Vt1 and Vt2). The polypeptide composition in the two forms of vitellins in stage I and stage III eggs were different at 195, 190, 130 and 110 kDa (Vt1) and 116 and 107 kDa (Vt2). On the other hand, in stage V eggs, 110, 95, 87 and 75 kDa (Vt1 and Vt2) were identified. Two antibodies (Ab1 and Ab2) were prepared against the purified proteins of stage V eggs and their specificity was demonstrated by radial immunoprecipitation, and Western blotting analysis. Two forms of vitellins were also found in stage V eggs after chromatography on Sepharose CL-2B column and hydroxylapatite and polyacrylamide gel electrophoresis.

Effect of the mass center shift for force-free flexible spacecraft

NASA Technical Reports Server (NTRS)

Meirovitch, L.; Juang, J.-N.

1975-01-01

For a spinning flexible spacecraft the mass center generally shifts relative to the nominal undeformed position. It is thought that this shift of center complicates spacecraft stability analysis. It is proved, on the basis of results achieved by Meirovitch and Calico (1972), that for the general class of force-free single-spin flexible spacecraft it is possible to ignore this shift of center without affecting the stability criteria in any significant way. A new theorem on inequalities for quadratic forms is proved to demonstrate the validity of the stability analysis.

Poly(ethyleneimines) in dermal applications: biocompatibility and antimicrobial effects.

PubMed

Wiegand, Cornelia; Bauer, Marius; Hipler, Uta-Christina; Fischer, Dagmar

2013-11-01

Cationic polyamines, such as poly(ethyleneimines) (PEIs), may recommend themselves for antimicrobial applications as they can interact with microbial membranes resulting in their disruption. The purpose of the study was the assessment of biocompatibility and antibacterial activity of PEIs with different architectures (branched (b) and linear (l)) and molar masses (0.8-750 kDa). lPEI and bPEI exhibited a strong antibacterial activity against Staphylococcus aureus and Escherichia coli with a more pronounced effect on the Gram-positive bacteria. lPEIs further demonstrated a higher antibacterial efficacy compared to bPEIs but no significant differences between 5 and 25 kDa were observed. In accordance, antibacterial activity of bPEI did not specifically depend on molar mass. Only slightly lower minimal inhibitory concentrations (MIC) were observed at 5 kDa (S. aureus) and 25 kDa (E. coli) in the tests. As PEIs are compelling candidates for use in antimicrobial treatment, two basic aspects have to be investigated: treatment effectiveness and safety. PEIs clearly induced molecular weight dependent cytotoxic effects in vitro. PEIs with low molecular weight (0.8 and 5 kDa) exhibited higher biocompatibility. Nonetheless, the results confirmed a low genotoxic potential of lPEI and bPEIs. In conclusion, 2.5 kDa-lPEI and 0.8 kDa-bPEI can be recommended for use as antimicrobial polymers in dermal applications due to their high biocompatibility with concomitant antibacterial efficacy. Copyright © 2013 Elsevier B.V. All rights reserved.

Experimental Protein Structure Verification by Scoring with a Single, Unassigned NMR Spectrum.

PubMed

Courtney, Joseph M; Ye, Qing; Nesbitt, Anna E; Tang, Ming; Tuttle, Marcus D; Watt, Eric D; Nuzzio, Kristin M; Sperling, Lindsay J; Comellas, Gemma; Peterson, Joseph R; Morrissey, James H; Rienstra, Chad M

2015-10-06

Standard methods for de novo protein structure determination by nuclear magnetic resonance (NMR) require time-consuming data collection and interpretation efforts. Here we present a qualitatively distinct and novel approach, called Comparative, Objective Measurement of Protein Architectures by Scoring Shifts (COMPASS), which identifies the best structures from a set of structural models by numerical comparison with a single, unassigned 2D (13)C-(13)C NMR spectrum containing backbone and side-chain aliphatic signals. COMPASS does not require resonance assignments. It is particularly well suited for interpretation of magic-angle spinning solid-state NMR spectra, but also applicable to solution NMR spectra. We demonstrate COMPASS with experimental data from four proteins--GB1, ubiquitin, DsbA, and the extracellular domain of human tissue factor--and with reconstructed spectra from 11 additional proteins. For all these proteins, with molecular mass up to 25 kDa, COMPASS distinguished the correct fold, most often within 1.5 Å root-mean-square deviation of the reference structure. Copyright © 2015 Elsevier Ltd. All rights reserved.

Experimental Protein Structure Verification by Scoring with a Single, Unassigned NMR Spectrum

PubMed Central

Courtney, Joseph M.; Ye, Qing; Nesbitt, Anna E.; Tang, Ming; Tuttle, Marcus D.; Watt, Eric D.; Nuzzio, Kristin M.; Sperling, Lindsay J.; Comellas, Gemma; Peterson, Joseph R.; Morrissey, James H.; Rienstra, Chad M.

2016-01-01

Standard methods for de novo protein structure determination by nuclear magnetic resonance (NMR) require time-consuming data collection and interpretation efforts. Here we present a qualitatively distinct and novel approach, called Comparative, Objective Measurement of Protein Architectures by Scoring Shifts (COMPASS), which identifies the best structures from a set of structural models by numerical comparison with a single, unassigned 2D 13C-13C NMR spectrum containing backbone and side-chain aliphatic signals. COMPASS does not require resonance assignments. It is particularly well suited for interpretation of magic-angle spinning solid-state NMR spectra, but also applicable to solution NMR spectra. We demonstrate COMPASS with experimental data from four proteins—GB1, ubiquitin, DsbA, and the extracellular domain of human tissue factor—and with reconstructed spectra from 11 additional proteins. For all these proteins, with molecular mass up to 25 kDa, COMPASS distinguished the correct fold, most often within 1.5 Å root-mean-square deviation of the reference structure. PMID:26365800

Electron Cyclotron Current Drive Efficiency in General Tokamak Geometry and Its Application to Advanced Tokamak Plasmas

NASA Astrophysics Data System (ADS)

Lin-Liu, Y. R.; Chan, V. S.; Luce, T. C.; Prater, R.

1998-11-01

Owing to relativistic mass shift in the cyclotron resonance condition, a simple and accurate interpolation formula for estimating the current drive efficiency, such as those(S.C. Chiu et al.), Nucl. Fusion 29, 2175 (1989).^,(D.A. Ehst and C.F.F. Karney, Nucl. Fusion 31), 1933 (1991). commonly used in FWCD, is not available in the case of ECCD. In this work, we model ECCD using the adjoint techniques. A semi-analytic adjoint function appropriate for general tokamak geometry is obtained using Fisch's relativistic collision model. Predictions of off-axis ECCD qualitatively and semi-quantitatively agrees with those of Cohen,(R.H. Cohen, Phys. Fluids 30), 2442 (1987). currently implemented in the raytracing code TORAY. The dependences of the current drive efficiency on the wave launch configuration and the plasma parameters will be presented. Strong absorption of the wave away from the resonance layer is shown to be an important factor in optimizing the off-axis ECCD for application to advanced tokamak operations.

Mass-independent isotope fractionation of Mo, Ru, Cd, and Te

NASA Astrophysics Data System (ADS)

Fujii, T.; Moynier, F.; Albarède, F.

2006-12-01

The variation of the mean charge distribution in the nucleus with the neutron number of different isotopes induces a tenuous shift of the nuclear field. The mass fractionation induced during phase changes is irregular, notably with 'staggering' between odd and even masses, and becomes increasingly non-linear for neutron-rich isotopes. A strong correlation is observed between the deviation of the isotopic effects from the linear dependence with mass and the corresponding nuclear charge radii. We first demonstrated on a number of elements the existence of such mass-independent isotope fractionation in laboratory experiments of solvent extraction with a macrocyclic compound. The isotope ratios were analyzed by multiple-collector inductively coupled plasma mass spectrometry with a typical precision of <100 ppm. The isotopes of odd and even atomic masses are enriched in the solvent to an extent that closely follows the variation of their nuclear charge radii. The present results fit Bigeleisen's (1996) model, which is the standard mass-dependent theory modified to include a correction term named the nuclear field shift effect. For heavy elements like uranium, the mass-independent effect is important enough to dominate the mass-dependent effect. We subsequently set out to compare the predictions of Bigeleisen's theory with the isotopic anomalies found in meteorites. Some of these anomalies are clearly inconsistent with nucleosynthetic effects (either s- or r-processes). Isotopic variations of Mo and Ru in meteorites, especially in Allende (CV3), show a clear indication of nucleosynthetic components. However, the mass-independent anomaly of Ru observed in Murchison (CM2) is a remarkable exception which cannot be explained by the nucleosynthetic model, but fits the nuclear field shift theory extremely well. The abundances of the even atomic mass Te isotopes in the leachates of carbonaceous chondrites, Allende, Murchison, and Orgueil, fit a mass-dependent law well, but the odd atomic mass isotope ^{125}Te clearly deviates from this correlation. The nuclear field shift theory shows that there is no effect on ^{130}Te but that the ^{125}Te anomaly is real. Carbonaceous chondrites do not reveal significant isotope fractionation of Cd isotopes, but a nuclear field shift effect is clearly present in type-3 (unequilibrated) ordinary chondrites. The nuclear field shift effect is temperature dependent and is probably more frequent in nature than commonly thought. It remains, together with nucleosynthetric anomalies, perfectly visible through the normalization of isotopic ratios to a reference value. In meteorites, this effect may originate both during condensation/evaporation processes in the nebular gas and during the metamorphism of the meteorite parent bodies.

Ribozyme-mediated signal augmentation on a mass-sensitive biosensor.

PubMed

Knudsen, Scott M; Lee, Joonhyung; Ellington, Andrew D; Savran, Cagri A

2006-12-20

Mass-based detection methods such as the quartz crystal microbalance (QCM) offer an attractive option to label-based methods; however the sensitivity is generally lower by comparison. In particular, low-molecular-weight analytes can be difficult to detect based on mass addition alone. In this communication, we present the use of effector-dependent ribozymes (aptazymes) as reagents for augmenting small ligand detection on a mass-sensitive device. Two distinct aptazymes were chosen: an L1-ligase-based aptazyme (L1-Rev), which is activated by a small peptide (MW approximately 2.4 kDa) from the HIV-1 Rev protein, and a hammerhead cleavase-based aptazyme (HH-theo3) activated by theophylline (MW = 180 Da). Aptazyme activity was observed in real time, and low-molecular-weight analyte detection has been successfully demonstrated with both aptazymes.

Protected Amine Labels: A Versatile Molecular Scaffold for Multiplexed Nominal Mass and Sub-Da Isotopologue Quantitative Proteomic Reagents

PubMed Central

Ficarro, Scott B.; Biagi, Jessica M.; Wang, Jinhua; Scotcher, Jenna; Koleva, Rositsa I.; Card, Joseph D.; Adelmant, Guillaume; He, Huan; Askenazi, Manor; Marshall, Alan G.; Young, Nicolas L.; Gray, Nathanael S.; Marto, Jarrod A.

2014-01-01

We assemble a versatile molecular scaffold from simple building blocks to create binary and multiplexed stable isotope reagents for quantitative mass spectrometry. Termed Protected Amine Labels (PAL), these reagents offer multiple analytical figures of merit including, (i) robust targeting of peptide N-termini and lysyl side chains, (ii) optimal mass spectrometry ionization efficiency through regeneration of primary amines on labeled peptides, (iii) an amino acid-based mass tag that incorporates heavy isotopes of carbon, nitrogen, and oxygen to ensure matched physicochemical and MS/MS fragmentation behavior among labeled peptides, and (iv) a molecularly efficient architecture, in which the majority of hetero-atom centers can be used to synthesize a variety of nominal mass and sub-Da isotopologue stable isotope reagents. We demonstrate the performance of these reagents in well-established strategies whereby up to four channels of peptide isotopomers, each separated by 4 Da are quantified in MS-level scans with accuracies comparable to current commercial reagents. In addition we utilize the PAL scaffold to create isotopologue reagents in which labeled peptide analogs differ in mass based on the binding energy in carbon and nitrogen nuclei, thereby allowing quantification based on MS or MS/MS spectra. We demonstrate accurate quantification for reagents that support 6-plex labeling and propose extension of this scheme to 9-channels based on a similar PAL scaffold. Finally we provide exemplar data that extends the application of isotopologe-based quantification reagents to medium resolution, quadrupole time-of-flight mass spectrometers. PMID:24496597

The plastid ribosomal proteins. Identification of all the proteins in the 30 S subunit of an organelle ribosome (chloroplast).

PubMed

Yamaguchi, K; von Knoblauch, K; Subramanian, A R

2000-09-15

Identification of all the protein components of a plastid (chloroplast) ribosomal 30 S subunit has been achieved, using two-dimensional gel electropholesis, high performance liquid chromatography purification, N-terminal sequencing, polymerase chain reaction-based screening of cDNA library, nucleotide sequencing, and mass spectrometry (electrospray ionization, matrix-assisted laser desorption/ionization time-of-flight, and reversed-phase HPLC coupled with electrospray ionization mass spectrometry). 25 proteins were identified, of which 21 are orthologues of all Escherichia coli 30 S ribosomal proteins (S1-S21), and 4 are plastid-specific ribosomal proteins (PSRPs) that have no homologues in the mitochondrial, archaebacterial, or cytosolic ribosomal protein sequences in data bases. 12 of the 25 plastid 30 S ribosomal proteins (PRPs) are encoded in the plastid genome, whereas the remaining 13 are encoded by the nuclear genome. Post-translational transit peptide cleavage sites for the maturation of the 13 cytosolically synthesized PRPs, and post-translational N-terminal processing in the maturation of the 12 plastid synthesized PRPs are described. Post-translational modifications in several PRPs were observed: alpha-N-acetylation of S9, N-terminal processings leading to five mature forms of S6 and two mature forms of S10, C-terminal and/or internal modifications in S1, S14, S18, and S19, leading to two distinct forms differing in mass and/or charge (the corresponding modifications are not observed in E. coli). The four PSRPs in spinach plastid 30 S ribosomal subunit (PSRP-1, 26.8 kDa, pI 6.2; PSRP-2, 21.7 kDa, pI 5.0; PSRP-3, 13.8 kDa, pI 4.9; PSRP-4, 5.2 kDa, pI 11.8) comprise 16% (67.6 kDa) of the total protein mass of the 30 S subunit (429.3 kDa). PSRP-1 and PSRP-3 show sequence similarities with hypothetical photosynthetic bacterial proteins, indicating their possible origins in photosynthetic bacteria. We propose the hypothesis that PSRPs form a "plastid translational regulatory module" on the 30 S ribosomal subunit structure for the possible mediation of nuclear factors on plastid translation.

An integrative strategy for quantitative analysis of the N-glycoproteome in complex biological samples.

PubMed

Wang, Ji; Zhou, Chuang; Zhang, Wei; Yao, Jun; Lu, Haojie; Dong, Qiongzhu; Zhou, Haijun; Qin, Lunxiu

2014-01-15

The complexity of protein glycosylation makes it difficult to characterize glycosylation patterns on a proteomic scale. In this study, we developed an integrated strategy for comparatively analyzing N-glycosylation/glycoproteins quantitatively from complex biological samples in a high-throughput manner. This strategy entailed separating and enriching glycopeptides/glycoproteins using lectin affinity chromatography, and then tandem labeling them with 18O/16O to generate a mass shift of 6 Da between the paired glycopeptides, and finally analyzing them with liquid chromatography-mass spectrometry (LC-MS) and the automatic quantitative method we developed based on Mascot Distiller. The accuracy and repeatability of this strategy were first verified using standard glycoproteins; linearity was maintained within a range of 1:10-10:1. The peptide concentration ratios obtained by the self-build quantitative method were similar to both the manually calculated and theoretical values, with a standard deviation (SD) of 0.023-0.186 for glycopeptides. The feasibility of the strategy was further confirmed with serum from hepatocellular carcinoma (HCC) patients and healthy individuals; the expression of 44 glycopeptides and 30 glycoproteins were significantly different between HCC patient and control serum. This strategy is accurate, repeatable, and efficient, and may be a useful tool for identification of disease-related N-glycosylation/glycoprotein changes.

/sup 3/H)forskolin. Direct photoaffinity labeling of the erythrocyte D-glucose transporter

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

Shanahan, M.F.; Morris, D.P.; Edwards, B.M.

1987-05-05

Irradiation of erythrocyte ghosts in the presence of (/sup 3/H)forskolin resulted in a concentration-dependent, covalent incorporation of radiolabel into several of the major membrane protein bands. Most of the incorporation occurred in four regions of the gel. Peak 1 (216 kDa) was a sharp peak near the top of the gel in the region corresponding to spectrin. Peak 2 appeared to be associated with band 3 (89 kDa), while a third peak occurred around the position of band 4.2 (76 kDa). The fourth region of labeling was a broad area between 43-75 kDa which corresponds to the region of themore » glucose transporter. Forskolin labeling of this region was inhibited by cytochalasin B and D-glucose, but not L-glucose. Extraction of extrinsic membrane proteins resulted in a loss of radiolabeled protein from the 216- and 76-kDa regions. Treatment of membranes labeled with either cytochalasin B or forskolin with endo-beta-galactosidase resulted in identical shifts of the 43 to 75-kDa peaks to 42 kDa. Similarly, trypsinization of membranes photolabeled with either cytochalasin B or forskolin resulted in the generation of a 17-kDa radiolabeled fragment in both cases. Photoincorporation of (/sup 3/H)cytochalasin B into the glucose transporter was blocked in a concentration-dependent manner by unlabeled forskolin.« less

Spontaneous emission and atomic line shift in causal perturbation theory

NASA Astrophysics Data System (ADS)

Marzlin, Karl-Peter; Fitzgerald, Bryce

2018-04-01

We derive a spontaneous emission rate and line shift for two-level atoms coupled to the radiation field using causal perturbation theory. In this approach, employing the theory of distribution splitting prevents the occurrence of divergent integrals. Our method confirms the result for an atomic decay rate but suggests that the cutoff frequency for the atomic line shift is determined by the atomic mass, rather than the Bohr radius or electron mass.

Forward Masking of the Speech-Evoked Auditory Brainstem Response.

PubMed

Hodge, Sarah E; Menezes, Denise C; Brown, Kevin D; Grose, John H

2018-02-01

The hypothesis tested was that forward masking of the speech-evoked auditory brainstem response (sABR) increases peak latency as an inverse function of masker-signal interval (Δt), and that the overall persistence of forward masking is age dependent. Older listeners exhibit deficits in forward masking. If forward-masked sABRs provide an objective measure of the susceptibility of speech sounds to prior stimulation, then this provides a novel approach to examining the age dependence of temporal processing. A /da/ stimulus forward masked by speech-shaped noise (Δt = 4-64 ms) was used to measure sABRs in 10 younger and nine older participants. Forward masking of subsegments of the /da/ stimulus (Δt = 16 ms) and click trains (Δt = 0-64 ms) was also measured. Forward-masked sABRs from young participants showed an increase in latency with decreasing Δt for the initial peak. Latency shifts for later peaks were smaller and more uniform. None of the peak latencies returned to baseline by Δt = 64 ms. Forward-masked /da/ subsegments showed peak latency shifts that did not depend simply on peak position, while forward-masked click trains showed latency shifts that were dependent on click position. The sABRs from older adults were less robust but confirmed the viability of the approach. Forward masking of the sABR provides an objective measure of the susceptibility of the auditory system to prior stimulation. Failure of recovery functions to return to baseline suggests an interaction between forward masking by the prior masker and temporal effects within the stimulus itself.

Spontaneous Mass and Charge Losses from Single Multi-Megadalton Ions Studied by Charge Detection Mass Spectrometry

NASA Astrophysics Data System (ADS)

Keifer, David Z.; Alexander, Andrew W.; Jarrold, Martin F.

2017-03-01

Spontaneous mass and charge losses from individual multi-megadalton ions have been observed with charge detection mass spectrometry (CDMS) by trapping single hepatitis B virus (HBV) capsids for 3 s. Gradual increases in the oscillation frequency of single ions in the ion trap are attributed mainly to mass loss (probably solvent, water, and/or salt). The total mass lost during the 3 s trapping period peaks at around 20 kDa for 4 MDa HBV T = 4 capsids. Discrete frequency drops punctuate the gradual increases in the oscillation frequencies. The drops are attributed to a sudden loss of charge. In most cases a single positive charge is lost along with some mass (on average around 1000 Da). Charge loss occurs for over 40% of the trapped ions. It usually occurs near the beginning of the trapping event, and it occurs preferentially in regions of the trap with strong electric fields, indicating that external electric fields promote charge loss. This process may contribute to the decrease in m/ z resolution that often occurs with megadalton ions.

Dynamic fluctuations in dopamine efflux in the prefrontal cortex and nucleus accumbens during risk-based decision making.

PubMed

St Onge, Jennifer R; Ahn, Soyon; Phillips, Anthony G; Floresco, Stan B

2012-11-21

Mesocorticolimbic dopamine (DA) has been implicated in cost/benefit decision making about risks and rewards. The prefrontal cortex (PFC) and nucleus accumbens (NAc) are two DA terminal regions that contribute to decision making in distinct manners. However, how fluctuations of tonic DA levels may relate to different aspects of decision making remains to be determined. The present study measured DA efflux in the PFC and NAc with microdialysis in well trained rats performing a probabilistic discounting task. Selection of a small/certain option always delivered one pellet, whereas another, large/risky option yielded four pellets, with probabilities that decreased (100-12.5%) or increased (12.5-100%) across four blocks of trials. Yoked-reward groups were also included to control for reward delivery. PFC DA efflux during decision making decreased or increased over a session, corresponding to changes in large/risky reward probabilities. Similar profiles were observed from yoked-rewarded rats, suggesting that fluctuations in PFC DA reflect changes in the relative rate of reward received. NAc DA efflux also showed decreasing/increasing trends over the session during both tasks. However, DA efflux was higher during decision making on free- versus forced-choice trials and during periods of greater reward uncertainty. Moreover, changes in NAc DA closely tracked shifts in choice biases. These data reveal dynamic and dissociable fluctuations in PFC and NAc DA transmission associated with different aspects of risk-based decision making. PFC DA may signal changes in reward availability that facilitates modification of choice biases, whereas NAc DA encodes integrated signals about reward rates, uncertainty, and choice, reflecting implementation of decision policies.

Proteoform analysis of lipocalin-type prostaglandin D-synthase from human cerebrospinal fluid by isoelectric focusing and superficially porous liquid chromatography with Fourier transform mass spectrometry.

PubMed

Zhang, Junmei; Corbett, John R; Plymire, Daniel A; Greenberg, Benjamin M; Patrie, Steven M

2014-05-01

Lipocalin-type prostaglandin D-synthase (L-PGDS) in cerebrospinal fluid contributes to the maturation and maintenance of the CNS. L-PGDS PTMs may contribute to pathobiology of different CNS diseases, but methods to monitor its proteoforms are limited. Herein, we combined off-gel IEF and superficially porous LC (SPLC) with Fourier transform MS to characterize common cerebrospinal fluid L-PGDS proteoforms. Across 3D physiochemical space (pI, hydrophobicity, and mass), 217 putative proteoforms were observed from 21 to 24 kDa and pI 5-10. Glycoprotein accurate mass information, combined with MS/MS analysis of peptides generated from 2D-fractionated proteoforms, enabled the putative assignment of 208 proteoforms with varied PTM positional occupants. Fifteen structurally related N-glycans at N29 and N56 were observed, with different N-glycan compositional variants being preferred on each amino acid. We also observed that sialic acid content was a major factor for pI shifts between L-PGDS proteoforms. Other putative PTMs characterized include a core-1 HexHexNAc-O-glycan at S7, acetylation at K16 and K138, sulfonation at S41 and T142, and dioxidation at C43 and C145. The IEF-SPLC-MS platform presented provides 30-40× improved peak capacity versus conventional 2DE and shows potential for repeatable proteoform analysis of surrogate PTM-based biomarkers. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Fostering Informed Decisions: Impact of a Decision Aid Among Men Registered to Undergo Mass Screening for Prostate Cancer in a Randomized Controlled Trial

PubMed Central

Williams, Randi M.; Davis, Kimberly M.; Luta, George; Edmond, Sara N.; Dorfman, Caroline S.; Schwartz, Marc D.; Lynch, John; Ahaghotu, Chiledum; Taylor, Kathryn L.

2013-01-01

Objective Screening asymptomatic men for prostate cancer is controversial and informed decision making is recommended. Within two prostate cancer screening programs, we evaluated the impact of a print-based decision aid (DA) on decision-making outcomes. Methods Men (N=543) were 54.9 (SD=8.1) years old and 61% were African-American. The 2 (booklet type: DA vs. usual care (UC)) × 2 (delivery mode: Home vs. Clinic) randomized controlled trial assessed decisional and screening outcomes at baseline, two-months, and 13-months. Results Intention-to-treat linear regression analyses using generalized estimating equations revealed that DA participants reported improved knowledge relative to UC (B=.41, p<.05). For decisional conflict, per-protocol analyses revealed a group by time interaction (B = −.69, p<.05), indicating that DA participants were less likely to report decisional conflict at two-months compared to UC participants (OR=.49, 95% CI: .26–.91, p<.05). Conclusion This is the first randomized trial to evaluate a DA in the context of free mass screening, a challenging setting in which to make an informed decision. The DA was highly utilized by participants, improved knowledge and reduced decisional conflict. Practice implications These results are valuable in understanding ways to improve the decisions of men who seek screening and can be easily implemented within many settings. PMID:23357414

Identification of caleosin and oleosin in oil bodies of pine pollen.

PubMed

Pasaribu, Buntora; Chen, Chii-Shiarng; Liao, Yue Ken; Jiang, Pei-Luen; Tzen, Jason T C

2017-02-01

Unique proteins including steroleosin, caleosin, oleosin-L, and oleosin-G have been identified in seed oil bodies of pine (Pinus massoniana). In this study, mature pollen grains with wing-like bladders were collected from pine (Pinus elliottii). Ultrastructural studies showed that oil bodies were present in pollen grains, but not the attached bladders, and the presence of oil bodies was further confirmed by fluorescent staining with BODIPY 493/503. Stable oil bodies were successfully purified from pine pollen grains, and analyzed to be mainly composed of triacylglycerols. Putative oleosin and caleosin in pine pollen oil bodies were detected by immunoassaying with antibodies against sesame seed caleosin and lily pollen oleosin. Complete cDNA fragments encoding these two pollen oil-body proteins were obtained by PCR cloning. Sequence alignment showed that pine pollen caleosin (27 kDa) was highly homologous to pine seed caleosin (28 kDa) except for the lack of an appendix of eight residues at the C-terminus in accord with the 1 kDa difference in their molecular masses. Pine pollen oleosin (15 kDa) was highly homologous to pine seed oleosin-G (14 kDa) except for an insertion of eight residues at the N-terminus in accord with the 1 kDa difference in their molecular masses. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

DETERMINATION OF ION AND NEUTRAL LOSS COMPOSITIONS AND DECONVOLUTION OF PRODUCT ION MASS SPECTRA USING AN ORTHOGONAL ACCELERATION, TIME-OF-FLIGHT MASS SPECTROMETER AND AN ION CORRELATION PROGRAM

EPA Science Inventory

Exact masses of monoisotopic ions and the relative isotopic abundances (RIAs) of ions greater in mass by 1 and 2 Da than the monoisotopic ion are independent and complementary physical properties useful for istinguishing among ion compositions possible for a given nominal mass. U...

The c15 ring of the Spirulina platensis F-ATP synthase: F1/F0 symmetry mismatch is not obligatory

PubMed Central

Pogoryelov, Denys; Yu, Jinshu; Meier, Thomas; Vonck, Janet; Dimroth, Peter; Muller, Daniel J

2005-01-01

The oligomeric c ring of the F-ATP synthase from the alkaliphilic cyanobacterium Spirulina platensis was isolated and characterized. Mass spectroscopy analysis indicated a mass of 8,210 Da, reflecting that of a c monomer. The mass increased by 206 Da after treatment with the c-subunit-specific inhibitor dicyclohexylcarbodiimide (DCCD), which indicated modification of the ion-binding carboxylate by DCCD. Atomic force microscopy topographs of c rings from S. platensis showed 15 symmetrically assembled subunits. The c15-mer reported here is the largest c ring that is isolated and does not show the classical c-ring mismatch to the three-fold symmetry of the F1 domain. PMID:16170308

Periastron shifts of stellar orbits near the Galactic Center

NASA Astrophysics Data System (ADS)

Rubilar, G. F.; Eckart, A.

2001-07-01

The presence of a 2.9+/-0.4 million solar mass object in the central stellar cluster of the Milky Way has recently been demonstrated via measurements of the stellar proper motions and radial velocities. This mass is located at the position of the compact radio source Sagittarius A* (Sgr A*) at a distance of Ro=8.0 kpc and is most likely present in the form of a massive black hole (BH). Some of the stars have a projected distance to Sgr A* of <=0.005 pc and have proper motion velocities of up to 1400 km s-1. Recent measurements indicate that their orbits show significant curvatures indicating that the stars indeed orbit the central compact object. Detailed measurements of the stellar orbits close to Sgr A* will allow us to precisely determine the distribution of this mass. With an increased point source sensitivity due to the combination of large telescope apertures, adaptive optics, and - in the very near future - NIR interferometry it is likely that stars with orbital time scales of the order of one year will be detected. Theses sources, however, will most likely not be on simple Keplerian orbits. The effects of measurable prograde relativistic and retrograde Newtonian periastron shifts will result in rosetta shaped orbits. A substantial Newtonian periastron rotation can already be expected if only a few percent of the central mass are extended. We discuss the conditions under which an extended mass can (over-) compensate the relativistic periastron shift. We also demonstrate that measuring a single periastron shift is not sufficient to determine the distribution of an extended mass component. A periastron shift will allow us to determine the inclination of the stellar orbits and to derive inclination corrected shift values. These have to be acquired for three stars on orbits with different energy or angular momentum in order to unambiguously solve for the compactness, extent and shape of any extended mass contribution.

Colloid characterization and in situ release in shallow groundwater under different hydrogeology conditions.

PubMed

Zhou, Jingjing; Liu, Dan; Zhang, Wenjing; Chen, Xuequn; Huan, Ying; Yu, Xipeng

2017-06-01

Changes to groundwater hydrodynamics and chemistry can lead to colloid release that can have a major impact on the groundwater environment. To analyze the effects of colloid release caused by artificial groundwater recharge, field and laboratory tests on colloid characterization and colloid release were conducted. The field tests were carried out at an artificial recharge test site in Shandong Province. In the field investigation, one recharge water sample and five groundwater samples were collected and filtered through three levels of ultrafiltration membranes, with pore sizes of 0.45 μm, 100 kDa, and 50 kDa. The field results indicated that the colloid mass concentrations in groundwater retained between membranes with pore sizes of 100 kDa-0.45 μm and 50 kDa-100 kDa were 19 and 62 mg/L, respectively. In recharge water, the colloid mass concentrations retained by 100-kDa-0.45-μm and 50-kDa-100-kDa membranes were 3 and 99 mg/L, respectively. Colloids detected on the ultrafiltration membranes were mainly inorganic between 100 kDa and 0.45 μm, and mainly organic between 50 and 100 kDa. Based on the field colloid investigation results, the organic colloid was chosen in the laboratory experiments to reveal its release behavior under different conditions. Porous media diameter, flux, ionic strength (IS), and ion valence were changed to determine their influences on organic colloid concentration outflow from undisturbed porous media. The experiment's results indicate that decreasing the diameter, and increasing the flux, ionic strength, and the number of divalent cations, can promote organic colloid release. The organic colloid release rate in the early stage was high and is thus likely to affect the quality of groundwater. The results provide a useful scientific basis for minimizing changes to hydrodynamic and hydrochemical conditions during artificial recharge, thus safeguarding groundwater quality.

Reducing body myopathy and desmin storage in skeletal muscle: morphological and biochemical findings.

PubMed

Bertini, E; Salviati, G; Apollo, F; Ricci, E; Servidei, S; Broccolini, A; Papacci, M; Tonali, P

1994-01-01

We describe clinical, morphological and biochemical findings of a patient with reducing body myopathy (RBM). This 15-year-old patient was affected by severe limb-girdle progressive myopathy with asymmetric distribution. Muscle biopsy showed many fibers with cytoplasmic polymorphic masses, which stained dark purple with modified Gomori's trichrome, associated with proliferation of cytoplasmic bodies. Cytoplasmic polymorphic masses showed marked reducing activity with menadione-nitro blue tetrazolium reaction. Ultrastructurally, there was great amount of highly electron-dense tubular-filamentous structures of 16-17 nm in diameter. Immunohistochemistry showed that many fibers were positive for desmin. Sodium dodecyl sulfate-electrophoresis disclosed an increase in two bands of approximately 53 and 70 kDa, and Western blot demonstrated that the 53-kDa band was desmin. It was not possible to characterize the 70-kDa protein further.

The molecular mass of dextran used to modify magnetite nanoparticles affects insulin amyloid aggregation

NASA Astrophysics Data System (ADS)

Siposova, Katarina; Pospiskova, Kristyna; Bednarikova, Zuzana; Safarik, Ivo; Safarikova, Mirka; Kubovcikova, Martina; Kopcansky, Peter; Gazova, Zuzana

2017-04-01

Protein transformation from its soluble state into amyloid aggregates is associated with amyloid-related diseases. Amyloid deposits of insulin fibrils have been found in the sites of subcutaneous insulin application in patients with prolonged diabetes. Using atomic force microscopy and ThT fluorescence assay we have investigated the interference of insulin amyloid aggregation with superparamagnetic Fe3O4-based nanoparticles (SPIONs) coated with dextran (DEX); molecular mass of dextran was equal to 15-20, 40 or 70 kDa. The obtained data indicate that all three types of dextran coated nanoparticles (NP-FeDEXs) are able to inhibit insulin fibrillization and to destroy amyloid fibrils. The extent of anti-amyloid activities depends on the properties of NP-FeDEXs, mainly on the size of nanoparticles which is determined by molecular mass of dextran molecules. The most effective inhibiting activity was observed for the smallest nanoparticles coated with 15-20 kDa dextran. Contrary, the highest destroying activity was observed for the largest NP-FeDEX (70 kDa dextran).

A HUPO test sample study reveals common problems in mass spectrometry-based proteomics

PubMed Central

Bell, Alexander W.; Deutsch, Eric W.; Au, Catherine E.; Kearney, Robert E.; Beavis, Ron; Sechi, Salvatore; Nilsson, Tommy; Bergeron, John J.M.

2009-01-01

We carried out a test sample study to try to identify errors leading to irreproducibility, including incompleteness of peptide sampling, in LC-MS-based proteomics. We distributed a test sample consisting of an equimolar mix of 20 highly purified recombinant human proteins, to 27 laboratories for identification. Each protein contained one or more unique tryptic peptides of 1250 Da to also test for ion selection and sampling in the mass spectrometer. Of the 27 labs, initially only 7 labs reported all 20 proteins correctly, and only 1 lab reported all the tryptic peptides of 1250 Da. Nevertheless, a subsequent centralized analysis of the raw data revealed that all 20 proteins and most of the 1250 Da peptides had in fact been detected by all 27 labs. The centralized analysis allowed us to determine sources of problems encountered in the study, which include missed identifications (false negatives), environmental contamination, database matching, and curation of protein identifications. Improved search engines and databases are likely to increase the fidelity of mass spectrometry-based proteomics. PMID:19448641

Size-Sorting Combined with Improved Nanocapillary-LC-MS for Identification of Intact Proteins up to 80 kDa

PubMed Central

Vellaichamy, Adaikkalam; Tran, John C.; Catherman, Adam D.; Lee, Ji Eun; Kellie, John F.; Sweet, Steve M.M.; Zamdborg, Leonid; Thomas, Paul M.; Ahlf, Dorothy R.; Durbin, Kenneth R.; Valaskovic, Gary A.; Kelleher, Neil L.

2010-01-01

Despite the availability of ultra-high resolution mass spectrometers, methods for separation and detection of intact proteins for proteome-scale analyses are still in a developmental phase. Here we report robust protocols for on-line LC-MS to drive high-throughput top-down proteomics in a fashion similar to bottom-up. Comparative work on protein standards showed that a polymeric stationary phase led to superior sensitivity over a silica-based medium in reversed-phase nanocapillary-LC, with detection of proteins >50 kDa routinely accomplished in the linear ion trap of a hybrid Fourier-Transform mass spectrometer. Protein identification was enabled by nozzle-skimmer dissociation (NSD) and detection of fragment ions with <5 ppm mass accuracy for highly-specific database searching using custom software. This overall approach led to identification of proteins up to 80 kDa, with 10-60 proteins identified in single LC-MS runs of samples from yeast and human cell lines pre-fractionated by their molecular weight using a gel-based sieving system. PMID:20073486

Optimization of high molecular weight pullulan production by Aureobasidium pullulans in batch fermentations.

PubMed

Gibson, Larry H; Coughlin, Robert W

2002-01-01

Of five strains of Aureobasidium pullulans studied, NRRL Y-2311-1 yielded the highest titer (26.2 g/L) of pullulan and formed the lowest amount of melanin-like pigment. Sucrose was superior to glucose as the carbon and energy source on the basis of yield and titer of pullulan produced. Pullulan titer was higher (26.2 vs 5.1 g/L), biomass concentration was lower (6.9 vs 12.7 g/L), and DO was lower (0 vs 60% of saturation) when the fermenter was agitated by a marine propeller compared to Rushton impellers. Pullulan produced by strain NRRL Y-2311-1 ranged in weight-average molar mass (M(w)) from 486 KDa and number-average molar mass (M(n)) from 220 Da on day 1 of growth to 390 KDa and 690 Da on day 6; M(w) declined by about 35% from day 1 to day 3, the day of maximum pullulan titer. For the other strains, the ranges of molar mass on the day of maximum pullulan titer were 338-614 KDa (M(w)) and 100-6820 Da (M(n)).

Chemometric Analysis of Gas Chromatography – Mass Spectrometry Data using Fast Retention Time Alignment via a Total Ion Current Shift Function

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

Nadeau, Jeremy S.; Wright, Bob W.; Synovec, Robert E.

2010-04-15

A critical comparison of methods for correcting severely retention time shifted gas chromatography-mass spectrometry (GC-MS) data is presented. The method reported herein is an adaptation to the Piecewise Alignment Algorithm to quickly align severely shifted one-dimensional (1D) total ion current (TIC) data, then applying these shifts to broadly align all mass channels throughout the separation, referred to as a TIC shift function (SF). The maximum shift varied from (-) 5 s in the beginning of the chromatographic separation to (+) 20 s toward the end of the separation, equivalent to a maximum shift of over 5 peak widths. Implementing themore » TIC shift function (TIC SF) prior to Fisher Ratio (F-Ratio) feature selection and then principal component analysis (PCA) was found to be a viable approach to classify complex chromatograms, that in this study were obtained from GC-MS separations of three gasoline samples serving as complex test mixtures, referred to as types C, M and S. The reported alignment algorithm via the TIC SF approach corrects for large dynamic shifting in the data as well as subtle peak-to-peak shifts. The benefits of the overall TIC SF alignment and feature selection approach were quantified using the degree-of-class separation (DCS) metric of the PCA scores plots using the type C and M samples, since they were the most similar, and thus the most challenging samples to properly classify. The DCS values showed an increase from an initial value of essentially zero for the unaligned GC-TIC data to a value of 7.9 following alignment; however, the DCS was unchanged by feature selection using F-Ratios for the GC-TIC data. The full mass spectral data provided an increase to a final DCS of 13.7 after alignment and two-dimensional (2D) F-Ratio feature selection.« less

Effects of Housing on Methamphetamine-Induced Neurotoxicity and Spatial Learning and Memory.

PubMed

Gutierrez, Arnold; Jablonski, Sarah A; Amos-Kroohs, Robyn M; Barnes, Anna C; Williams, Michael T; Vorhees, Charles V

2017-07-19

Severe stress potentiates methamphetamine (MA) neurotoxicity. However, whether moderate stress increases or decreases the neurotoxic effects of MA is unknown. We assessed the effects of MA (4 × 10 mg/kg at 2 h intervals) in combination with prior barren-cage housing in adult male Sprague-Dawley rats on monoamines and glial fibrillary acid protein (GFAP) in one cohort and spatial learning and memory in the Morris water maze in another cohort. MA reduced dopamine (DA) and serotonin (5-HT) in the neostriatum and nucleus accumbens, 5-HT in the hippocampus, and increased GFAP in neostriatum and nucleus accumbens compared with saline controls. In neostriatum, barren-cage housing protected against MA-induced increases in GFAP, but it did not prevent DA and 5-HT reductions, although it did increase hippocampal norepinephrine. MA impaired spatial learning during acquisition, reversal, and shift phases and impaired reference memory on reversal and shift probe trials. Barren-cage housing enhanced performance during acquisition but not during reversal or shift or on probe trials. The data indicate that prior barren-cage housing moderates MA-induced neostriatal astrogliosis and initial spatial learning, but has no protective effect when the platform is smaller and relocated and therefore requires cognitive flexibility in relearning.

Trypsin and MALDI matrix pre-coated targets simplify sample preparation for mapping proteomic distributions within biological tissues by imaging mass spectrometry

PubMed Central

Zubair, Faizan; Laibinis, Paul E.; Swisher, William G.; Yang, Junhai; Spraggins, Jeffrey M.; Norris, Jeremy L.; Caprioli, Richard M.

2017-01-01

Prefabricated surfaces containing α-cyano-4-hydroxycinnamic acid and trypsin have been developed to facilitate enzymatic digestion of endogenous tissue proteins prior to matrix-assisted laser desorption/ionization (MALDI) imaging mass spectrometry (IMS). Tissue sections are placed onto slides that were previously coated with α-cyano-4-hydroxycinnamic acid and trypsin. After incubation to promote enzymatic digestion, the tissue is analyzed by MALDI IMS to determine the spatial distribution of the tryptic fragments. The peptides detected in the MALDI IMS dataset were identified by Liquid chromatography-tandem mass spectrometry/mass spectrometry. Protein identification was further confirmed by correlating the localization of unique tryptic fragments originating from common parent proteins. Using this procedure, proteins with molecular weights as large as 300 kDa were identified and their distributions were imaged in sections of rat brain. In particular, large proteins such as myristoylated alanine-rich C-kinase substrate (29.8 kDa) and spectrin alpha chain, non-erythrocytic 1 (284 kDa) were detected that are not observed without trypsin. The pre-coated targets simplify workflow and increase sample throughput by decreasing the sample preparation time. Further, the approach allows imaging at higher spatial resolution compared with robotic spotters that apply one drop at a time. PMID:27676701

Purification and characterization of myrosinase from horseradish (Armoracia rusticana) roots.

PubMed

Li, Xian; Kushad, Mosbah M

2005-06-01

Myrosinase (beta-thioglucoside glucohydrolase; EC 3.2.3.147) from horseradish (Armoracia rusticana) roots was purified to homogeneity by ammonium sulfate fractionation, Q-sepharose, and concanavalin A sepharose affinity chromatography. The purified protein migrated as a single band with a mass of about 65 kDa on SDS-polyacrylamide gel electrophoresis. Using LC-MS/MS, this band was identified as myrosinase. Western blot analysis, using the anti-myrosinase monoclonal antibody 3D7, showed a single band of about 65 kDa for horseradish crude extract and for the purified myrosinase. The native molecular mass of the purified myrosinase was estimated, using gel filtration, to be about 130 kDa. Based on these data, it appeared that myrosinase from horseradish root consists of two subunits of similar molecular mass of about 65 kDa. The enzyme exhibited high activity at broad pH (pH 5.0-8.0) and temperature (37 and 45 degrees C). The purified enzyme remained stable at 4 degrees C for more than 1 year. Using sinigrin as a substrate, the Km and Vmax values for the purified enzyme were estimated to be 0.128 mM and 0.624 micromol min(-1), respectively. The enzyme was strongly activated by 0.5 mM ascorbic acid and was able to breakdown intact glucosinolates in a crude extract of broccoli.

Effect of shift in growth temperature on tolerance of psychrotrophic and mesophilic strains of Bacillus cereus to heat and sodium chloride.

PubMed

Mahakarnchanakul, W; Beuchat, L R

1999-01-01

A shift in growth temperature of a psychrotrophic (F3802A/84) strain and a mesophilic strain (B4ac-1) of Bacillus cereus grown at 30 degrees C for 10 h, then at 37 degrees C or 40 degrees C for 14 h, enhanced thermotolerance. Sodium chloride, at concentrations of 2.0 and 4.0% in brain heart infusion (BHI) broth, had no effect on thermotolerance of strain B4ac-1 heated at 50 degrees C, whereas the same concentrations of NaCl caused a decrease in thermotolerance of strain F3802A/84 heated at 48 degrees C. A downshift in growth temperature from 30 degrees C to 10 degrees C followed by incubation for 3 to 9 days increased thermotolerance of strain F3802A/84 but not strain B4ac-1 heated in BHI broth containing 2.0 or 4.0% NaCl compared to thermotolerance in BHI broth containing 0.5% NaCl. Protein analysis using one-dimensional gel electrophoresis revealed an increase in proteins with molecular weights of 54, 50, 44, and 42 kDa in cells of strain F3802A/84 and 83 and 69 kDa in cells of strain B4ac-1 subjected to an upshift in growth temperature from 30 degrees C to 37 degrees C or 40 degrees C, respectively. A downshift in growth temperature from 30 degrees C to 10 degrees C resulted in substantial amounts of proteins with molecular weights of 63, 40, and 29 kDa produced by strain F3802A/84 and 63 kDa to be produced by strain B4ac-1. Proteins produced in response to upshift or downshift in growth temperature are suspected to play an important role in heat resistance of the psychrotrophic and mesophilic strains of B. cereus examined in this study. Changes in resistance to heat or refrigeration temperatures, as well as tolerance to NaCl, as affected by previous exposure of cells to temperature shifts may influence the ability of B. cereus to grow in minimally processed foods during distribution and storage.

Origin of the DA and non-DA white dwarf stars

NASA Technical Reports Server (NTRS)

Shipman, Harry L.

1989-01-01

Various proposals for the bifurcation of the white dwarf cooling sequence are reviewed. 'Primordial' theories, in which the basic bifurcation of the white dwarf sequence is rooted in events predating the white dwarf stage of stellar evolution, are discussed, along with the competing 'mixing' theories in which processes occurring during the white dwarf stage are responsible for the existence of DA or non-DA stars. A new proposal is suggested, representing a two-channel scenario. In the DA channel, some process reduces the hydrogen layer mass to the value of less than 10 to the -7th. The non-DA channel is similar to that in the primordial scenario. These considerations suggest that some mechanism operates in both channels to reduce the thickness of the outermost layer of the white dwarf. It is also noted that accretion from the interstellar medium has little to do with whether a particular white dwarf becomes a DA or a non-DA star.

Short-chain ubiquitination is associated with the degradation rate of a cell-surface-resident bile salt export pump (BSEP/ABCB11).

PubMed

Hayashi, Hisamitsu; Sugiyama, Yuichi

2009-01-01

The reduced expression of the bile salt export pump (BSEP/ABCB11) at the canalicular membrane is associated with cholestasis-induced hepatotoxicity due to the accumulation of bile acids in hepatocytes. We demonstrated previously that 4-phenylbutyrate (4PBA) treatment, a U.S. Food and Drug Administration-approved drug for the treatment of urea cycle disorders, induces the cell-surface expression of BSEP by prolonging the degradation rate of cell-surface-resident BSEP. On the other hand, BSEP mutations, E297G and D482G, found in progressive familial intrahepatic cholestasis type 2 (PFIC2), reduced it by shortening the degradation rate of cell-surface-resident BSEP. Therefore, to help the development of the medical treatment of cholestasis, we investigated the underlying mechanism by which 4PBA and PFIC2-type mutations affect the BSEP degradation from cell surface, focusing on short-chain ubiquitination. In Madin-Darby canine kidney II (MDCK II) cells expressing BSEP and rat canalicular membrane vesicles, the molecular mass of the mature form of BSEP/Bsep shifted from 170 to 190 kDa after ubiquitin modification (molecular mass, 8 kDa). Ubiquitination susceptibility of BSEP/Bsep was reduced in vitro and in vivo by 4PBA treatment and, conversely, was enhanced by BSEP mutations E297G and D482G. Moreover, biotin-labeling studies using MDCK II cells demonstrated that the degradation of cell-surface-resident chimeric protein fusing ubiquitin to BSEP was faster than that of BSEP itself. In conclusion, BSEP/Bsep is modified with two to three ubiquitins, and its ubiquitination is modulated by 4PBA treatment and PFIC2-type mutations. Modulation of short-chain ubiquitination can regulate the change in the degradation rate of cell-surface-resident BSEP by 4PBA treatment and PFIC2-type mutations.

Study of an Unusual Advanced Glycation End-Product (AGE) Derived from Glyoxal Using Mass Spectrometry

NASA Astrophysics Data System (ADS)

Lopez-Clavijo, Andrea F.; Duque-Daza, Carlos A.; Romero Canelon, Isolda; Barrow, Mark P.; Kilgour, David; Rabbani, Naila; Thornalley, Paul J.; O'Connor, Peter B.

2014-04-01

Glycation is a post-translational modification (PTM) that affects the physiological properties of peptides and proteins. In particular, during hyperglycaemia, glycation by α-dicarbonyl compounds generate α-dicarbonyl-derived glycation products also called α-dicarbonyl-derived advanced glycation end products. Glycation by the α-dicarbonyl compound known as glyoxal was studied in model peptides by MS/MS using a Fourier transform ion cyclotron resonance mass spectrometer. An unusual type of glyoxal-derived AGE with a mass addition of 21.98436 Da is reported in peptides containing combinations of two arginine-two lysine, and one arginine-three lysine amino acid residues. Electron capture dissociation and collisionally activated dissociation results supported that the unusual glyoxal-derived AGE is formed at the guanidino group of arginine, and a possible structure is proposed to illustrate the 21.9843 Da mass addition.

It's a Trap! A Review of MOMA and Other Ion Traps in Space or Under Development

NASA Technical Reports Server (NTRS)

Arevalo, R., Jr.; Brinckerhoff, W. B.; Mahaffy, P. R.; van Amerom, F. H. W.; Danell, R. M.; Pinnick, V. T.; Li, X.; Hovmand, L.; Getty, S. A.; Goesmann, F.;

The 30 kDa protein co-purified with chick liver glutathione S-transferases is a carbonyl reductase.

PubMed

Tsai, S P; Wang, L Y; Yeh, H I; Tam, M F

1996-02-08

An unidentified 30 kDa protein was co-purified with chick liver glutathione S-transferases from S-hexylglutathione affinity column. The protein was isolated to apparent homogeneity with chromatofocusing. The molecular mass of the protein was determined to be 30 277 +/- 3 dalton by mass spectrometry. The protein was digested with Achromobacter proteinase I. Amino-acid sequence analyses of the resulting peptides show a high degree of identity with those of human carbonyl reductase. The protein is active with menadione as substrate. Thus, it is identified as chick liver carbonyl reductase.

SODA: Smart Objects, Dumb Archives

NASA Technical Reports Server (NTRS)

Nelson, Michael L.; Maly, Kurt; Zubair, Mohammad; Shen, Stewart N. T.

2004-01-01

We present the Smart Object, Dumb Archive (SODA) model for digital libraries (DLs). The SODA model transfers functionality traditionally associated with archives to the archived objects themselves. We are exploiting this shift of responsibility to facilitate other DL goals, such as interoperability, object intelligence and mobility, and heterogeneity. Objects in a SODA DL negotiate presentation of content and handle their own terms and conditions. In this paper we present implementations of our smart objects, buckets, and our dumb archive (DA). We discuss the status of buckets and DA and how they are used in a variety of DL projects.

Backbone and sidechain methyl Ile (δ1), Leu and Val chemical shift assignments of RDE-4 (1-243), an RNA interference initiation protein in C. elegans.

PubMed

Chiliveri, Sai Chaitanya; Kumar, Sonu; Marelli, Udaya Kiran; Deshmukh, Mandar V

2012-10-01

The RNAi pathway of several organisms requires presence of double stranded RNA binding proteins for functioning of Dicer in gene regulation. In C. elegans, a double stranded RNA binding protein, RDE-4 (385 aa, 44 kDa) recognizes long exogenous dsRNA and initiates the RNAi pathway. We have achieved complete backbone and stereospecific methyl sidechain Ile (δ1), Leu and Val chemical shifts of first 243 amino acids of RDE-4, namely RDE-4ΔC.

Rapid high-throughput characterisation, classification and selection of recombinant mammalian cell line phenotypes using intact cell MALDI-ToF mass spectrometry fingerprinting and PLS-DA modelling.

PubMed

Povey, Jane F; O'Malley, Christopher J; Root, Tracy; Martin, Elaine B; Montague, Gary A; Feary, Marc; Trim, Carol; Lang, Dietmar A; Alldread, Richard; Racher, Andrew J; Smales, C Mark

2014-08-20

Despite many advances in the generation of high producing recombinant mammalian cell lines over the last few decades, cell line selection and development is often slowed by the inability to predict a cell line's phenotypic characteristics (e.g. growth or recombinant protein productivity) at larger scale (large volume bioreactors) using data from early cell line construction at small culture scale. Here we describe the development of an intact cell MALDI-ToF mass spectrometry fingerprinting method for mammalian cells early in the cell line construction process whereby the resulting mass spectrometry data are used to predict the phenotype of mammalian cell lines at larger culture scale using a Partial Least Squares Discriminant Analysis (PLS-DA) model. Using MALDI-ToF mass spectrometry, a library of mass spectrometry fingerprints was generated for individual cell lines at the 96 deep well plate stage of cell line development. The growth and productivity of these cell lines were evaluated in a 10L bioreactor model of Lonza's large-scale (up to 20,000L) fed-batch cell culture processes. Using the mass spectrometry information at the 96 deep well plate stage and phenotype information at the 10L bioreactor scale a PLS-DA model was developed to predict the productivity of unknown cell lines at the 10L scale based upon their MALDI-ToF fingerprint at the 96 deep well plate scale. This approach provides the basis for the very early prediction of cell lines' performance in cGMP manufacturing-scale bioreactors and the foundation for methods and models for predicting other mammalian cell phenotypes from rapid, intact-cell mass spectrometry based measurements. Copyright © 2014 Elsevier B.V. All rights reserved.

Production of Hypoallergenic Antibacterial Peptides from Defatted Soybean Meal in Membrane Bioreactor: A Bioprocess Engineering Study with Comprehensive Product Characterization

PubMed Central

2017-01-01

Summary Hypoallergenic antibacterial low-molecular-mass peptides were produced from defatted soybean meal in a membrane bioreactor. In the first step, soybean meal proteins were digested with trypsin in the bioreactor, operated in batch mode. For the tryptic digestion of soybean meal protein, optimum initial soybean meal concentration of 75 g/L, temperature of 40 °C and pH=9.0 were determined. After enzymatic digestion, low-molecular-mass peptides were purified with cross-flow flat sheet membrane (pore size 100 µm) and then with tubular ceramic ultrafiltration membrane (molecular mass cut-off 5 kDa). Effects of transmembrane pressure and the use of a static turbulence promoter to reduce the concentration polarization near the ultrafiltration membrane surface were examined and their positive effects were proven. For the filtration with ultrafiltration membrane, transmembrane pressure of 3·105 Pa with 3-stage discontinuous diafiltration was found optimal. The molecular mass distribution of purified peptides using ultrafiltration membrane was determined by a liquid chromatography–electrospray ionization quadrupole time-of-flight mass spectrometry setup. More than 96% of the peptides (calculated as relative frequency) from the ultrafiltration membrane permeate had the molecular mass M≤1.7 kDa and the highest molecular mass was found to be 3.1 kDa. The decrease of allergenic property due to the tryptic digestion and membrane filtration was determined by an enzyme-linked immunosorbent assay and it was found to exceed 99.9%. It was also found that the peptides purified in the ultrafiltration membrane promoted the growth of Pediococcus acidilactici HA6111-2 and they possessed antibacterial activity against Bacillus cereus. PMID:29089846

Development and Evaluation of a Reverse-Entry Ion Source Orbitrap Mass Spectrometer

NASA Astrophysics Data System (ADS)

Poltash, Michael L.; McCabe, Jacob W.; Patrick, John W.; Laganowsky, Arthur; Russell, David H.

2018-05-01

As a step towards development of a high-resolution ion mobility mass spectrometer using the orbitrap mass analyzer platform, we describe herein a novel reverse-entry ion source (REIS) coupled to the higher-energy C-trap dissociation (HCD) cell of an orbitrap mass spectrometer with extended mass range. Development of the REIS is a first step in the development of a drift tube ion mobility-orbitrap MS. The REIS approach retains the functionality of the commercial instrument ion source which permits the uninterrupted use of the instrument during development as well as performance comparisons between the two ion sources. Ubiquitin (8.5 kDa) and lipid binding to the ammonia transport channel (AmtB, 126 kDa) protein complex were used as model soluble and membrane proteins, respectively, to evaluate the performance of the REIS instrument. Mass resolution obtained with the REIS is comparable to that obtained using the commercial ion source. The charge state distributions for ubiquitin and AmtB obtained on the REIS are in agreement with previous studies which suggests that the REIS-orbitrap EMR retains native structure in the gas phase.

Visual measurement of the evaporation process of a sessile droplet by dual-channel simultaneous phase-shifting interferometry.

PubMed

Sun, Peng; Zhong, Liyun; Luo, Chunshu; Niu, Wenhu; Lu, Xiaoxu

2015-07-16

To perform the visual measurement of the evaporation process of a sessile droplet, a dual-channel simultaneous phase-shifting interferometry (DCSPSI) method is proposed. Based on polarization components to simultaneously generate a pair of orthogonal interferograms with the phase shifts of π/2, the real-time phase of a dynamic process can be retrieved with two-step phase-shifting algorithm. Using this proposed DCSPSI system, the transient mass (TM) of the evaporation process of a sessile droplet with different initial mass were presented through measuring the real-time 3D shape of a droplet. Moreover, the mass flux density (MFD) of the evaporating droplet and its regional distribution were also calculated and analyzed. The experimental results show that the proposed DCSPSI will supply a visual, accurate, noncontact, nondestructive, global tool for the real-time multi-parameter measurement of the droplet evaporation.

Antigenic profile and localization of Clonorchis sinensis proteins in the course of infection

PubMed Central

Kim, Tae Yun; Song, Kye-Yong; Sohn, Woon-Mok; Kang, Shin-Yong

2001-01-01

In the course of Clonorchis sinensis infection, antigens presented to the hosts may be in a close relation to growth of the fluke. The antigenic proteins stimulating IgG antibody production were chronologically identified by immunoblot and localized by immunohistochemical staining. In the early stage of infection until 12 weeks post-infection (PI), antigens were proteins with molecular mass larger than 34 kDa which were derived from the tegument, testes and intrauterine eggs. After 20 weeks PI, antigens recognized were 29, 27 and 26 kDa proteins from the intestine, excretory bladder and reproductive organs. It is suggested that the tegumental proteins are the most potent antigens and the excretory-secretory proteins with middle molecular mass of 26-45 kDa contribute to the high level production of antibodies after 20 weeks of the C. sinensis infection. PMID:11775331

A Silicon Nanomembrane Detector for Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS) of Large Proteins

PubMed Central

Park, Jonghoo; Blick, Robert H.

2013-01-01

We describe a MALDI-TOF ion detector based on freestanding silicon nanomembrane technology. The detector is tested in a commercial MALDI-TOF mass spectrometer with equimolar mixtures of proteins. The operating principle of the nanomembrane detector is based on phonon-assisted field emission from these silicon nanomembranes, in which impinging ion packets excite electrons in the nanomembrane to higher energy states. Thereby the electrons can overcome the vacuum barrier and escape from the surface of the nanomembrane via field emission. Ion detection is demonstrated of apomyoglobin (16,952 Da), aldolase (39,212 Da), bovine serum albumin (66,430 Da), and their equimolar mixtures. In addition to the three intact ions, a large number of fragment ions are also revealed by the silicon nanomembrane detector, which are not observable with conventional detectors. PMID:24152929

A silicon nanomembrane detector for matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) of large proteins.

PubMed

Park, Jonghoo; Blick, Robert H

2013-10-11

We describe a MALDI-TOF ion detector based on freestanding silicon nanomembrane technology. The detector is tested in a commercial MALDI-TOF mass spectrometer with equimolar mixtures of proteins. The operating principle of the nanomembrane detector is based on phonon-assisted field emission from these silicon nanomembranes, in which impinging ion packets excite electrons in the nanomembrane to higher energy states. Thereby the electrons can overcome the vacuum barrier and escape from the surface of the nanomembrane via field emission. Ion detection is demonstrated of apomyoglobin (16,952 Da), aldolase (39,212 Da), bovine serum albumin (66,430 Da), and their equimolar mixtures. In addition to the three intact ions, a large number of fragment ions are also revealed by the silicon nanomembrane detector, which are not observable with conventional detectors.

Dopamine disposition in the presynaptic process regulates the severity of methamphetamine-induced neurotoxicity.

PubMed

Kuhn, Donald M; Francescutti-Verbeem, Dina M; Thomas, David M

2008-10-01

Methamphetamine (METH) is well known for its ability to cause damage to dopamine (DA) nerve endings of the striatum. The mechanisms by which METH causes neurotoxicity are not fully understood, but likely candidates are increased oxidative and nitrosative stress and mitochondrial dysfunction. Microglial activation is also emerging as an important element of the METH neurotoxic cascade, and it appears that extensive cross-talk between these cells and DA nerve endings is an early event in this process. It may seem paradoxical, but DA itself is also thought to be an essential factor in the neuronal damaging effects of METH, but issues relating to its precise role in this regard remain unanswered. We present in this overview a summary of studies that tested how alterations in the disposition of presynaptic DA (injections of reserpine, L-DOPA, or clorgyline) modulate METH neurotoxicity. In all cases, these drugs significantly increased the magnitude of microglial activation as well as the severity of damage to striatal DA nerve endings caused by METH. The enhancement of METH effects in striatum by reserpine, L-DOPA, and clorgyline persisted for 14 days and showed no evidence of recovery. These data establish that subtle shifts in the newly synthesized pool of DA can cause substantial changes in the severity of METH-induced neurotoxicity. DA released into the synapse by METH is very likely the source of downstream reactants that provoke microglial activation and the ensuing damage to DA nerve endings.

A Low-Noise, Wideband Preamplifier for a Fourier-Transform Ion Cyclotron Resonance Mass Spectrometer

PubMed Central

Mathur, Raman; Knepper, Ronald W.; O'Connor, Peter B.

2009-01-01

FTMS performance parameters such as limits of detection, dynamic range, sensitivity, and even mass accuracy and resolution can be greatly improved by enhancing its detection circuit. An extended investigation of significant design considerations for optimal signal-to-noise ratio in an FTMS detection circuit are presented. A low noise amplifier for an FTMS is developed based on the discussed design rules. The amplifier has a gain of ≈ 3500 and a bandwidth of 10 kHz - 1 MHz corresponding to m/z range of 100 Da to 10 kDa (at 7 Tesla). The performance of the amplifier was tested on a MALDI-FTMS, and has demonstrated a 25-fold reduction in noise in a mass spectrum of C60 compared to that of a commercial amplifier. PMID:18029195

DA white dwarfs from the LSS-GAC survey DR1: the preliminary luminosity and mass functions and formation rate

NASA Astrophysics Data System (ADS)

Rebassa-Mansergas, A.; Liu, X.-W.; Cojocaru, R.; Yuan, H.-B.; Torres, S.; García-Berro, E.; Xiang, M.-X.; Huang, Y.; Koester, D.; Hou, Y.; Li, G.; Zhang, Y.

2015-06-01

Modern large-scale surveys have allowed the identification of large numbers of white dwarfs. However, these surveys are subject to complicated target selection algorithms, which make it almost impossible to quantify to what extent the observational biases affect the observed populations. The LAMOST (Large Sky Area Multi-Object Fiber Spectroscopic Telescope) Spectroscopic Survey of the Galactic anticentre (LSS-GAC) follows a well-defined set of criteria for selecting targets for observations. This advantage over previous surveys has been fully exploited here to identify a small yet well-characterized magnitude-limited sample of hydrogen-rich (DA) white dwarfs. We derive preliminary LSS-GAC DA white dwarf luminosity and mass functions. The space density and average formation rate of DA white dwarfs we derive are 0.83 ± 0.16 × 10-3 pc-3 and 5.42 ± 0.08 × 10-13 pc-3 yr-1, respectively. Additionally, using an existing Monte Carlo population synthesis code we simulate the population of single DA white dwarfs in the Galactic anticentre, under various assumptions. The synthetic populations are passed through the LSS-GAC selection criteria, taking into account all possible observational biases. This allows us to perform a meaningful comparison of the observed and simulated distributions. We find that the LSS-GAC set of criteria is highly efficient in selecting white dwarfs for spectroscopic observations (80-85 per cent) and that, overall, our simulations reproduce well the observed luminosity function. However, they fail at reproducing an excess of massive white dwarfs present in the observed mass function. A plausible explanation for this is that a sizable fraction of massive white dwarfs in the Galaxy are the product of white dwarf-white dwarf mergers.

Lipid, Detergent, and Coomassie Blue G-250 Affect the Migration of Small Membrane Proteins in Blue Native Gels

PubMed Central

Crichton, Paul G.; Harding, Marilyn; Ruprecht, Jonathan J.; Lee, Yang; Kunji, Edmund R. S.

2013-01-01

Blue native gel electrophoresis is a popular method for the determination of the oligomeric state of membrane proteins. Studies using this technique have reported that mitochondrial carriers are dimeric (composed of two ∼32-kDa monomers) and, in some cases, can form physiologically relevant associations with other proteins. Here, we have scrutinized the behavior of the yeast mitochondrial ADP/ATP carrier AAC3 in blue native gels. We find that the apparent mass of AAC3 varies in a detergent- and lipid-dependent manner (from ∼60 to ∼130 kDa) that is not related to changes in the oligomeric state of the protein, but reflects differences in the associated detergent-lipid micelle and Coomassie Blue G-250 used in this technique. Higher oligomeric state species are only observed under less favorable solubilization conditions, consistent with aggregation of the protein. Calibration with an artificial covalent AAC3 dimer indicates that the mass observed for solubilized AAC3 and other mitochondrial carriers corresponds to a monomer. Size exclusion chromatography of purified AAC3 in dodecyl maltoside under blue native gel-like conditions shows that the mass of the monomer is ∼120 kDa, but appears smaller on gels (∼60 kDa) due to the unusually high amount of bound negatively charged dye, which increases the electrophoretic mobility of the protein-detergent-dye micelle complex. Our results show that bound lipid, detergent, and Coomassie stain alter the behavior of mitochondrial carriers on gels, which is likely to be true for other small membrane proteins where the associated lipid-detergent micelle is large when compared with the mass of the protein. PMID:23744064

Urtica dioica pollen allergy: Clinical, biological, and allergomics analysis.

PubMed

Tiotiu, Angelica; Brazdova, Andrea; Longé, Cyril; Gallet, Patrice; Morisset, Martine; Leduc, Virginie; Hilger, Christiane; Broussard, Cédric; Couderc, Rémy; Sutra, Jean-Pierre; Sénéchal, Hélène; Poncet, Pascal

2016-11-01

The most emblematic members of Urticaceae at allergic risk level are wall pellitories (Parietaria), whereas nettle (Urtica) pollen is considered as poorly allergenic. No allergen from nettle pollen has yet been characterized, whereas 4 are listed for Parietaria pollen by the International Union of Immunological Societies. Clinical and biological profiles of 2 adult men who developed symptoms against nettle pollen and/or leaves were studied. To characterize the allergic reaction and identify the potential nettle pollen sensitizing allergens. IgE-mediated reaction to nettle pollen extract was evaluated by skin prick test, immunoassay, nasal provocation, and basophil activation test. To characterize specific nettle pollen allergens, an allergomic (IgE immunoproteomic) analysis was performed combining 1- and 2-dimensional electrophoresis, IgE immunoblots of nettle pollen extract, identification of allergens by mass spectrometry, and database queries. The results of biological and immunochemical analyses revealed that the allergic rhinitis was due to Urtica dioica pollen in both patients. The allergomic analysis of nettle pollen extract allowed the characterization of 4 basic protein allergens: a thaumatin-like protein (osmotin) with a relative molecular mass of 27 to 29 kDa, a pectinesterase (relative molecular mass, 40 kDa), and 2 other basic proteins with relative molecular masses of 14 to 16 kDa and 43 kDa. There is no or only very weak allergen associations between pellitory and nettle pollen. Exposure to nettle pollen can be responsible of allergic symptoms, and several allergens were characterized. Unravelling the allergens of this underestimated allergy might help to improve diagnosis and care for patients, to predict cross-reactivities and design adapted specific immunotherapy. Copyright © 2016 American College of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

Turkish scorpion Buthacus macrocentrus: general characterization of the venom and description of Bu1, a potent mammalian Na⁺-channel α-toxin.

PubMed

Caliskan, F; Quintero-Hernández, V; Restano-Cassulini, R; Batista, C V F; Zamudio, F Z; Coronas, F I; Possani, L D

2012-03-01

The venom of the scorpion Buthacus macrocentrus of Turkey was fractionated by high performance liquid chromatography (HPLC) and its mass finger print analysis was obtained by spectrometry. More than 70 different fractions were obtained, allowing the determination of the molecular masses of at least 60 peptides ranging between 648 and 44,336 Da. The venom is enriched with peptides containing molecular masses between 3200-4500 Da, and 6000-7500 Da. They very likely correspond to K⁺-channel and Na⁺-channel specific peptides, respectively, as expected from venoms of scorpions of the family Buthidae, already determined for other species. The major component obtained from HPLC was shown to be lethal to mice and was further purified and characterized. It contains 65 amino acid residues maintained closely packed by 4 disulfide bridges, and shows a molecular weight of 7263 Da. Additionally, a cDNA from the venomous glands of this scorpion was used in conjunction with sequence data from Edman degradation and mass spectrometry for cloning the gene that codes for Bu1 as we named this toxin. This gene codes for a 67 amino acid residues peptide, where the two last are eliminated post-translationally for production of an amidated C-terminal arginine. Its sequence is closely related to toxins from the species Leiurus quinquestriatus, as revealed by a phylogenetic tree analysis. Electrophysiological results conducted with Bu1 using patch-clamp techniques indicate that it modifies the Na⁺ currents, in a similar way as other well known α-scorpion toxins. These results support the conclusion that this species of scorpions is dangerous to humans, having an epidemiological interest for the country. Copyright © 2012 Elsevier Ltd. All rights reserved.

Dopamine physiology in the basal ganglia of male zebra finches during social stimulation.

PubMed

Ihle, Eva C; van der Hart, Marieke; Jongsma, Minke; Tecott, Larry H; Doupe, Allison J

2015-06-01

Accumulating evidence suggests that dopamine (DA) is involved in altering neural activity and gene expression in a zebra finch cortical-basal ganglia circuit specialized for singing, upon the shift between solitary singing and singing as a part of courtship. Our objective here was to sample changes in the extracellular concentrations of DA in Area X of adult and juvenile birds, to test the hypothesis that DA levels would change similarly during presentation of a socially salient stimulus in both age groups. We used microdialysis to sample the extracellular milieu of Area X in awake, behaving adult and juvenile male zebra finches, and analysed the dialysate using high-performance liquid chromatography coupled with electrochemical detection. The extracellular levels of DA in Area X increased significantly during both female presentation to adult males and tutor presentation to juvenile males. DA levels were not correlated with the time spent singing. We also reverse-dialysed Area X with pharmacologic agents that act either on DA systems directly or on norepinephrine, and found that all of these agents significantly increased DA levels (3- to 10-fold) in Area X. These findings suggest that changes in extracellular DA levels can be stimulated similarly by very different social contexts (courtship and interaction with tutor), and influenced potently by dopaminergic and noradrenergic drugs. These results raise the possibility that the arousal level or attentional state of the subject (rather than singing behavior) is the common feature eliciting changes in extracellular DA concentration. © 2015 The Authors. European Journal of Neuroscience published by Federation of European Neuroscience Societies and John Wiley & Sons Ltd.

Aversive Stimuli Differentially Modulate Real-Time Dopamine Transmission Dynamics within the Nucleus Accumbens Core and Shell

PubMed Central

Badrinarayan, Aneesha; Wescott, Seth A.; Vander Weele, Caitlin M.; Saunders, Benjamin T.; Couturier, Brenann E.; Maren, Stephen

2012-01-01

Although fear directs adaptive behavioral responses, how aversive cues recruit motivational neural circuitry is poorly understood. Specifically, while it is known that dopamine (DA) transmission within the nucleus accumbens (NAc) is imperative for mediating appetitive motivated behaviors, its role in aversive behavior is controversial. It has been proposed that divergent phasic DA transmission following aversive events may correspond to segregated mesolimbic dopamine pathways; however, this prediction has never been tested. Here, we used fast-scan cyclic voltammetry to examine real-time DA transmission within NAc core and shell projection systems in response to a fear-evoking cue. In male Sprague Dawley rats, we first demonstrate that a fear cue results in decreased DA transmission within the NAc core, but increased transmission within the NAc shell. We examined whether these changes in DA transmission could be attributed to modulation of phasic transmission evoked by cue presentation. We found that cue presentation decreased the probability of phasic DA release in the core, while the same cue enhanced the amplitude of release events in the NAc shell. We further characterized the relationship between freezing and both changes in DA as well as local pH. Although we found that both analytes were significantly correlated with freezing in the NAc across the session, changes in DA were not strictly associated with freezing while basic pH shifts in the core more consistently followed behavioral expression. Together, these results provide the first real-time neurochemical evidence that aversive cues differentially modulate distinct DA projection systems. PMID:23136417

The Exploration of Peptide Biomarkers in Malignant Pleural Effusion of Lung Cancer Using Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry

PubMed Central

Xu, Jing; Xu, Bin; Tang, Chuanhao; Li, Xiaoyan; Qin, Haifeng; Wang, Weixia; Wang, Hong; Wang, Zhongyuan; Li, Liangliang; Li, Zhihua; Gao, Hongjun

2017-01-01

Background. Diagnoses of malignant pleural effusion (MPE) are a crucial problem in clinics. In our study, we compared the peptide profiles of MPE and tuberculosis pleural effusion (TPE) to investigate the value of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) in diagnosis of MPE. Material and Methods. The 46 MPE and 32 TPE were randomly assigned to training set and validation set. Peptides were isolated by weak cation exchange magnetic beads and peaks in the m/z range of 800–10000 Da were analyzed. Comparing the peptide profile between 30 MPE and 22 TPE samples in training set by ClinProTools software, we screened the specific biomarkers and established a MALDI-TOF-MS classification of MPE. Finally, the other 16 MPE and 10 TPE were included to verify the model. We additionally determined carcinoembryonic antigen (CEA) in MPE and TPE samples using electrochemiluminescent immunoassay method. Results. Five peptide peaks (917.37 Da, 4469.39 Da, 1466.5 Da, 4585.21 Da, and 3216.87 Da) were selected to separate MPE and TPE by MALDI-TOF-MS. The sensitivity, specificity, and accuracy of the classification were 93.75%, 100%, and 96.15%, respectively, after blinded test. The sensitivity of CEA was significantly lower than MALDI-TOF-MS classification (P = 0.035). Conclusions. The results indicate MALDI-TOF-MS is a potential method for diagnosing MPE. PMID:28386154

The Exploration of Peptide Biomarkers in Malignant Pleural Effusion of Lung Cancer Using Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry.

PubMed

Xu, Jing; Xu, Bin; Tang, Chuanhao; Li, Xiaoyan; Qin, Haifeng; Wang, Weixia; Wang, Hong; Wang, Zhongyuan; Li, Liangliang; Li, Zhihua; Gao, Hongjun; He, Kun; Liu, Xiaoqing

2017-01-01

Background . Diagnoses of malignant pleural effusion (MPE) are a crucial problem in clinics. In our study, we compared the peptide profiles of MPE and tuberculosis pleural effusion (TPE) to investigate the value of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) in diagnosis of MPE. Material and Methods . The 46 MPE and 32 TPE were randomly assigned to training set and validation set. Peptides were isolated by weak cation exchange magnetic beads and peaks in the m / z range of 800-10000 Da were analyzed. Comparing the peptide profile between 30 MPE and 22 TPE samples in training set by ClinProTools software, we screened the specific biomarkers and established a MALDI-TOF-MS classification of MPE. Finally, the other 16 MPE and 10 TPE were included to verify the model. We additionally determined carcinoembryonic antigen (CEA) in MPE and TPE samples using electrochemiluminescent immunoassay method. Results . Five peptide peaks (917.37 Da, 4469.39 Da, 1466.5 Da, 4585.21 Da, and 3216.87 Da) were selected to separate MPE and TPE by MALDI-TOF-MS. The sensitivity, specificity, and accuracy of the classification were 93.75%, 100%, and 96.15%, respectively, after blinded test. The sensitivity of CEA was significantly lower than MALDI-TOF-MS classification ( P = 0.035). Conclusions . The results indicate MALDI-TOF-MS is a potential method for diagnosing MPE.

Structural characterization of hemoglobins from Monilifera and Frenulata tubeworms (Siboglinids): first discovery of giant hexagonal-bilayer hemoglobin in the former "Pogonophora" group.

PubMed

Meunier, Cédric; Andersen, Ann C; Bruneaux, Matthieu; Le Guen, Dominique; Terrier, Peran; Leize-Wagner, Emmanuelle; Zal, Franck

2010-01-01

Siboglinids are symbiotic polychete annelids having hemoglobins as essential oxygen- and sulfide-carriers for their endosymbiotic bacteria. We analyzed the structure of the hemoglobins from two species of siboglinids: the monilifera Sclerolinum contortum and the frenulata Oligobrachia webbi (i.e. haakonmosbiensis) from Norwegian cold seeps. Measured by Multi-Angle Laser Light Scattering (MALLS), Sclerolinum shows a 3190+/-50 kDa hexagonal bilayer hemoglobin (HBL-Hb) and a 461+/-46 kDa ring-Hb, just as vestimentifera, whereas Oligobrachia has a 409+/-3.7 kDa ring-Hb only. Electrospray Ionization-Mass Spectrometry (ESI-MS) showed Sclerolinum HBL-Hb composed of seven monomeric globins (15-16 kDa), three disulfide-bonded globin heterodimers and three linkers. The heterodimers always contain globin-b (15814.4+/-1.5 Da). Sclerolinum ring-Hb is composed of globins and dimers with identical masses as its HBL-Hb, but lacks linkers. Oligobrachia ring-Hb has three globin monomers (14-15 kDa) only, with no disulfide-bonded dimers. Comparison of Sclerolinum hemoglobins between Storegga and Haakon Mosby Mud Volcano, using the normalized height of deconvoluted ESI-MS peaks, shows differences in globin monomers abundances that could reflect genetic differences or differential gene expression between distinct seep populations. The discovery of HBL-Hb in Sclerolinum is a new element supporting the hypothesis of monilifera being phylogenetically more closely related to vestimentifera, than to frenulata.

Purification and characterization of two enone reductases from Saccharomyces cerevisiae.

PubMed

Wanner, P; Tressl, R

1998-07-01

Two enone reductases catalyzing irreversibly the enantioselective reduction of alpha,beta-unsaturated carbonyls have been purified 165-fold and 257-fold, respectively, from the cytosolic fraction of Saccharomyces cerevisiae by means of streptomycin sulfate treatment, Sephadex G-25 filtration, DEAE-Sepharose CL-6B chromatography, blue Sepharose CL-6B chromatography and Superdex 200 preparation-grade filtration. One enzyme (E I) was NADPH-dependent, showed a molecular mass of 75 kDa and decomposed under denaturing electrophoretic conditions into two subunits of 34 kDa and 37 kDa. The other enzyme (E II) was NADH linked and the molecular mass estimated by means of Superdex 200 preparation-grade filtration, was 130 kDa. The enzyme decomposed into subunits of 56 kDa and 64 kDa on SDS/PAGE. Both enzymes were most active at pH 4.8 and accepted 1-octen-3-one, 1-hexen-3-one, 3-alken-2-ones, 4-alken-3-ones, 2-cyclohexen-1-ones, 2-alkenals, 2,4-alkadienals, 2-phenyl-2-alkenals, and 2-alkyl-2-alkenals as substrates. Both enzymes showed their highest activities with 1-hexen-3-one and 1-octen-3-one, respectively. The Km values for 1-octen-3-one were estimated as 0.54 mM (E I) and 0.20 mM (E II), respectively. Both enzymes catalyzed the enantioselective reduction of cis- and trans-2-phenyl-2-butenal into (R)-2-phenylbutanal.

Profiling modifications for glioblastoma proteome using ultra-tolerant database search: Are the peptide mass shifts biologically relevant or chemically induced?

PubMed

Tarasova, Irina A; Chumakov, Peter M; Moshkovskii, Sergei A; Gorshkov, Mikhail V

2018-05-17

Peptide mass shifts were profiled using ultra-tolerant database search strategy for shotgun proteomics data sets of human glioblastoma cell lines demonstrating strong response to the type I interferon (IFNα-2b) treatment. The main objective of this profiling was revealing the cell response to IFN treatment at the level of protein modifications. To achieve this objective, statistically significant changes in peptide mass shift profiles between IFN treated and untreated glioblastoma samples were analyzed. Detailed analysis of MS/MS spectra allowed further interpretation of the observed mass shifts and differentiation between post-translational and artifact modifications. Malignant cells typically acquire increased sensitivity to viruses due to the deregulated antiviral mechanisms. Therefore, a viral therapy is considered as one of the promising approaches to treat cancer. However, recent studies have demonstrated that malignant cells can preserve intact antiviral mechanisms, e.g. interferon signaling, and develop resistance to virus infection in response to interferon treatment. Post translational modifications, e.g. tyrosine phosphorylation, are the interferon signaling drivers. Thus, comprehensive characterization of modifications is crucially important, yet, most challenging problem in cancer proteomics. Here, we report on the application of the recently introduced ultra-tolerant search strategy for profiling peptide modifications in the human glioblastoma cell lines demonstrating strong response to the type I interferon (IFNα-2b) treatment. The specific aim of the study was identification of statistically significant changes in peptide mass shift profiles between IFN treated and untreated glioblastoma samples, as well as determination of whether these shifts represent the biologically relevant modification. Copyright © 2018 Elsevier B.V. All rights reserved.

Effect of polarization force on head-on collision between multi-solitons in dusty plasma

NASA Astrophysics Data System (ADS)

Singh, Kuldeep; Sethi, Papihra; Saini, N. S.

2018-03-01

Head-on collision among dust acoustic (DA) multi-solitons in a dusty plasma with ions featuring non-Maxwellian hybrid distribution under the effect of the polarization force is investigated. The presence of the non-Maxwellian ions leads to eloquent modifications in the polarization force. Specifically, an increase in the superthermality index of ions (via κi) and nonthermal parameter (via α) diminishes the polarization parameter. By employing the extended Poincaré-Lighthill-Kuo method, two sided KdV equations are derived. The Hirota direct method is used to obtain multi-soliton solutions for each KdV equation, and all of them move along the same direction where the fastest moving soliton eventually overtakes the others. The expressions for collisional phase shifts after head-on collision of two, four, and six-(DA) solitons are derived under the influence of polarization force. It is found that the effect of polarization force and the presence of non-Maxwellian ions have an emphatic influence on the phase shifts after the head-on collision of DA rarefactive multi-solitons. In a small amplitude limit, the impact of polarization force on time evolution of multi-solitons is also illustrated. It is intensified that the present theoretical pronouncements actually effectuate in laboratory experiments and in space/astrophysical environments, in particular in Saturn's magnetosphere and comet tails.

Identification and Characterization of Human Proteoforms by Top-Down LC-21 Tesla FT-ICR Mass Spectrometry.

PubMed

Anderson, Lissa C; DeHart, Caroline J; Kaiser, Nathan K; Fellers, Ryan T; Smith, Donald F; Greer, Joseph B; LeDuc, Richard D; Blakney, Greg T; Thomas, Paul M; Kelleher, Neil L; Hendrickson, Christopher L

2017-02-03

Successful high-throughput characterization of intact proteins from complex biological samples by mass spectrometry requires instrumentation capable of high mass resolving power, mass accuracy, sensitivity, and spectral acquisition rate. These limitations often necessitate the performance of hundreds of LC-MS/MS experiments to obtain reasonable coverage of the targeted proteome, which is still typically limited to molecular weights below 30 kDa. The National High Magnetic Field Laboratory (NHMFL) recently installed a 21 T FT-ICR mass spectrometer, which is part of the NHMFL FT-ICR User Facility and available to all qualified users. Here we demonstrate top-down LC-21 T FT-ICR MS/MS of intact proteins derived from human colorectal cancer cell lysate. We identified a combined total of 684 unique protein entries observed as 3238 unique proteoforms at a 1% false discovery rate, based on rapid, data-dependent acquisition of collision-induced and electron-transfer dissociation tandem mass spectra from just 40 LC-MS/MS experiments. Our identifications included 372 proteoforms with molecular weights over 30 kDa detected at isotopic resolution, which substantially extends the accessible mass range for high-throughput top-down LC-MS/MS.

Triosephosphate isomerase and filamin C share common epitopes as novel allergens of Procambarus clarkii

USDA-ARS?s Scientific Manuscript database

Triosephosphate isomerase (TIM) is a key enzyme in glycolysis and has been identified as an allergen in saltwater products. In this study, TIM with a molecular mass of 28 kDa was purified from the freshwater crayfish (Procambarus clarkii) muscle. A 90-kDa protein that showed IgG/IgE cross-reactivity...

RISING FROM THE ASHES: MID-INFRARED RE-BRIGHTENING OF THE IMPOSTOR SN 2010da IN NGC 300

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

Lau, Ryan M.; Ressler, Michael E.; Kasliwal, Mansi M.

2016-10-20

We present multi-epoch mid-infrared (IR) photometry and the optical discovery observations of the “impostor” supernova (SN) 2010da in NGC 300 using new and archival Spitzer Space Telescope images and ground-based observatories. The mid-infrared counterpart of SN 2010da was detected as Spitzer Infrared Intensive Transient Survey (SPIRITS) 14bme in the SPIRITS, an ongoing systematic search for IR transients. Before erupting on 2010 May 24, the SN 2010da progenitor exhibited a constant mid-IR flux at 3.6 and only a slight ∼10% decrease at 4.5 μ m between 2003 November and 2007 December. A sharp increase in the 3.6 μ m flux followedmore » by a rapid decrease measured ∼150 days before and ∼80 days after the initial outburst, respectively, reveal a mid-IR counterpart to the coincident optical and high luminosity X-ray outbursts. At late times, after the outburst (∼2000 days), the 3.6 and 4.5 μ m emission increased to over a factor of two times the progenitor flux and is currently observed (as of 2016 Feb) to be fading, but still above the progenitor flux. We attribute the re-brightening mid-IR emission to continued dust production and increasing luminosity of the surviving system associated with SN 2010da. We analyze the evolution of the dust temperature ( T {sub d} ∼ 700–1000 K), mass ( M {sub d} ∼ 0.5–3.8 × 10{sup −7} M {sub ⊙}), luminosity ( L {sub IR} ∼ 1.3–3.5 × 10{sup 4} L {sub ⊙}), and the equilibrium temperature radius ( R {sub eq} ∼ 6.4–12.2 au) in order to resolve the nature of SN 2010da. We address the leading interpretation of SN 2010da as an eruption from a luminous blue variable high-mass X-ray binary (HMXB) system. We propose that SN 2010da is instead a supergiant (sg)B[e]-HMXB based on similar luminosities and dust masses exhibited by two other known sgB[e]-HMXB systems. Additionally, the SN 2010da progenitor occupies a similar region on a mid-IR color–magnitude diagram (CMD) with known sgB[e] stars in the Large Magellanic Cloud. The lower limit estimated for the orbital eccentricity of the sgB[e]-HMXB ( e > 0.82) from X-ray luminosity measurements is high compared to known sgHMXBs and supports the claim that SN 2010da may be associated with a newly formed HMXB system.« less

Affinity purification of angiotensin type 2 receptors from N1E-115 cells: evidence for agonist-induced formation of multimeric complexes.

PubMed

Siemens, I R; Yee, D K; Reagan, L P; Fluharty, S J

1994-01-01

The murine neuroblastoma N1E-115 cell line possesses type 1 and type 2 angiotensin II (AngII) receptor subtypes. In vitro differentiation of these cells substantially increases the density of the AT2-receptor subtype, whereas the density of the AT1 receptors remains unchanged. In the present study, we report that the zwitterionic detergent 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS) selectively solubilized AT2 receptors from N1E-115 cell membranes and that these receptors could be purified further to near homogeneity by affinity chromatography. More specifically, the presence of an agonist (AngII) during affinity purification of AT2 receptors resulted in the elution of high (110-kDa) and low (66-kDa) molecular mass proteins as determined by gel electrophoresis under nonreducing conditions. In contrast, when the nonselective antagonist Sar1,Ile8-AngII was used during purification, only the lower 66-kDa protein was observed. Affinity purification in the presence of the peptide and nonpeptide AT2-receptor antagonists CGP42112A and PD123319 also resulted in elution of the same 66-kDa protein, but unlike that in the presence of Sar1,Ile8-AngII, some of the high molecular weight site was observed as well. On the other hand, Losartan, an AT1-receptor antagonist, was completely ineffective in eluting any AngII receptors from the affinity column, further confirming their AT2 identity. After agonist elution, the 110-kDa band dissociated into two low molecular mass bands of 66 kDa and 54 kDa when sodium dodecyl sulfate-gel electrophoresis was run under reducing conditions.(ABSTRACT TRUNCATED AT 250 WORDS)

ESI-MS2 and Anti-inflammatory Studies of Cyclopropanic Triterpenes. UPLC-ESI-MS and MS2 Search of Related Metabolites from Donella ubanguiensis.

PubMed

Sandjo, Louis P; Nascimento, Marcus V P Dos Santos; da Silva, Layzon A L; Munhoz, Antonio C M; Pollo, Luiz A E; Biavatti, Maique W; Ngadjui, Bonaventure T; Opatz, Till; Fröde, Tania S

2017-01-01

Triterpenes are one of the largest secondary metabolites groups spread in the plant kingdom with various skeletons. These metabolites have showed various bioactivities including anti-inflammatory activity. The study aims to explore the mass spectrometry fragmentation of donellanic acids A-C (DA A-C), three compounds identified from Donella ubanguiensis; in addition, the fragmentation behaviour of these metabolites will serve as a fingerprint to search and characterise triterpenes congeners in fruits, bark and wood crude extracts of D. ubanguiensis. This work was prompted by the anti-inflammatory activity on leukocyte migration, exudate concentrations and myeloperoxidase activity obtained for DA A-B. The bioactivity was performed on mouse model of pleurisy induced by carrageenan and the parameters were analysed by veterinarian automated cell counter and colorimetric assays. While the tandem mass analyses of DA A-C were carried out by a direct infusion ESI-QTOF-MS/MS, the extracts were studied by UPLC-ESI-QTOF-MS and UPLC-ESI-QTOF-MS/MS. DA A displayed interesting anti-inflammatory activity by inhibiting leukocyte migration, exudate concentrations and myeloperoxidase activity (p < 0.05) while DA B was weakly active (p > 0.05). Moreover, the diagnostic of the MS 2 behaviour of DA A-C in conjunction with the chromatograms and the obtained MS 2 data of the crude extract led to the characterisation of three cyclopropane triterpenes (T1-T3) and six saponins (T4-T9) from the fruits, the bark, and the wood extracts. Donella species deserve more investigation since metabolites related to the anti-inflammatory compound (DA A) could be identified. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

Investigation of an enhanced resolution triple quadrupole mass spectrometer for high-throughput liquid chromatography/tandem mass spectrometry assays.

PubMed

Yang, Liyu; Amad, Ma'an; Winnik, Witold M; Schoen, Alan E; Schweingruber, Hans; Mylchreest, Iain; Rudewicz, Patrick J

2002-01-01

Triple quadrupole mass spectrometers, when operated in multiple reaction monitoring (MRM) mode, offer a unique combination of sensitivity, specificity, and dynamic range. Consequently, the triple quadrupole is the workhorse for high-throughput quantitation within the pharmaceutical industry. However, in the past, the unit mass resolution of quadrupole instruments has been a limitation when interference from matrix or metabolites cannot be eliminated. With recent advances in instrument design, triple quadrupole instruments now afford mass resolution of less than 0.1 Dalton (Da) full width at half maximum (FWHM). This paper describes the evaluation of an enhanced resolution triple quadrupole mass spectrometer for high-throughput bioanalysis with emphasis on comparison of selectivity, sensitivity, dynamic range, precision, accuracy, and stability under both unit mass (1 Da FWHM) and enhanced (

The Label Matters: μPET Imaging of the Biodistribution of Low Molar Mass 89Zr and 18F-Labeled Poly(2-ethyl-2-oxazoline).

PubMed

Glassner, Mathias; Palmieri, Luca; Monnery, Bryn D; Verbrugghen, Thomas; Deleye, Steven; Stroobants, Sigrid; Staelens, Steven; Wyffels, Leonie; Hoogenboom, Richard

2017-01-09

Poly(2-alkyl-2-oxazoline)s (PAOx) have received increasing interest for biomedical applications. Therefore, it is of fundamental importance to gain an in-depth understanding of the biodistribution profile of PAOx. We report the biodistribution of poly(2-ethyl-2-oxazoline) (PEtOx) with a molar mass of 5 kDa radiolabeled with PET isotopes 89 Zr and 18 F. 18 F-labeled PEtOx is prepared by the strain-promoted azide-alkyne cycloaddition (SPAAC) of [ 18 F]fluoroethylazide to bicyclo[6.1.0]non-4-yne (BCN)-functionalized PEtOx as many common labeling strategies were found to be unsuccessful for PEtOx. 89 Zr-labeled PEtOx is prepared using desferrioxamine end-groups as a chelator. Five kDa PEtOx shows a significantly faster blood clearance compared to PEtOx of higher molar mass while uptake in the liver is lower, indicating a minor contribution of the liver in excretion of the 5 kDa PEtOx. While [ 18 F]-PEtOx displays a rapid and efficient clearance from the kidneys, 5 kDa [ 89 Zr]-Df-PEtOx is not efficiently cleared over the time course of the study, which is most likely caused by trapping of 89 Zr-labeled metabolites in the renal tubules and not the polymer itself, demonstrating the importance of selecting the appropriate label for biodistribution studies.

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) coupled to XAD fractionation: Method to algal organic matter characterization.

PubMed

Nicolau, Rudy; Leloup, Maud; Lachassagne, Delphine; Pinault, Emilie; Feuillade-Cathalifaud, Geneviève

2015-05-01

This work is focused on the development of an analytical procedure for the improvement of the Organic Matter structure characterization, particularly the algal matter. Two fractions of algal organic matter from laboratory cultures of algae (Euglena gracilis) and cyanobacteria (Microcystis aeruginosa) were extracted with XAD resins. The fractions were studied using laser desorption ionization (LDI) and Matrix-Assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF). A comparison with the natural organic matter characteristics from commercial humic acids and fulvic acids extracted from Suwannee River was performed. Results show that algal and natural organic matters have unique quasi-polymeric structures. Significant repeating patterns were identified. Different fractions extracted from organic matter with common origin had common structures. Thus, 44, 114 and 169Da peaks separation for fractions from E. gracilis organic matter and 28, 58 and 100Da for M. aeruginosa ones were clearly observed. Using the developed protocol, a structural scheme and organic matter composition were obtained. The range 600-2000Da contained more architectural composition differences than the range 100-600Da, suggesting that organic matter is composed of an assembly of common small molecules. Associated to specific monomers, particular patterns were common to all samples but assembly and resulting structure were unique for each organic matter. Thus, XAD fractionation coupled to mass spectroscopy allowed determining a specific fingerprint for each organic matter. Copyright © 2015 Elsevier B.V. All rights reserved.

Mass shift of charmonium states in p bar A collision

NASA Astrophysics Data System (ADS)

Wolf, György; Balassa, Gábor; Kovács, Péter; Zétényi, Miklós; Lee, Su Houng

2018-05-01

The masses of the low lying charmonium states, namely, the J / Ψ, Ψ (3686), and Ψ (3770) are shifted downwards due to the second order Stark effect. In p bar +Au collisions at 6-10 GeV we study their in-medium propagation. The time evolution of the spectral functions of these charmonium states is studied with a Boltzmann-Uehling-Uhlenbeck (BUU) type transport model. We show that their in-medium mass shift can be observed in the dilepton spectrum. Therefore, by observing the dileptonic decay channel of these low lying charmonium states, especially for Ψ (3686), we can gain information about the magnitude of the gluon condensate in nuclear matter. This measurement could be performed at the upcoming PANDA experiment at FAIR.

Mechanism of membrane damage by El Tor hemolysin of Vibrio cholerae O1.

PubMed

Ikigai, H; Akatsuka, A; Tsujiyama, H; Nakae, T; Shimamura, T

1996-08-01

El Tor hemolysin (ETH; molecular mass, 65 kDa) derived from Vibrio cholerae O1 spontaneously assembled oligomeric aggregates on the membranes of rabbit erythrocyte ghosts and liposomes. Membrane-associated oligomers were resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting into two to nine bands with apparent molecular masses of 170 to 350 kDa. ETH assembled oligomers on a liposomal membrane consisting of phosphatidylcholine and cholesterol, but not on a membrane of phosphatidylcholine alone. Cholesterol could be replaced with diosgenin or ergosterol but not with 5alpha-cholestane-3-one, suggesting that sterol is essential for the oligomerization. The treatment of carboxyfluorescein-encapsulated liposomes with ETH caused a rapid release of carboxyfluorescein into the medium. Because dextrin 20 (molecular mass, 900 Da) osmotically protected ETH-mediated hemolysis, this hemolysis is likely to be caused by pore formation on the membrane. The pore size(s) estimated from osmotic protection assays was in the range of 1.2 to 1.6 nm. The pore formed on a rabbit erythrocyte membrane was confirmed morphologically by electron microscopy. Thus, we provide evidence that ETH damages the target by the assembly of hemolysin oligomers and pore formation on the membrane.

Mechanism of membrane damage by El Tor hemolysin of Vibrio cholerae O1.

PubMed Central

Ikigai, H; Akatsuka, A; Tsujiyama, H; Nakae, T; Shimamura, T

1996-01-01

El Tor hemolysin (ETH; molecular mass, 65 kDa) derived from Vibrio cholerae O1 spontaneously assembled oligomeric aggregates on the membranes of rabbit erythrocyte ghosts and liposomes. Membrane-associated oligomers were resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting into two to nine bands with apparent molecular masses of 170 to 350 kDa. ETH assembled oligomers on a liposomal membrane consisting of phosphatidylcholine and cholesterol, but not on a membrane of phosphatidylcholine alone. Cholesterol could be replaced with diosgenin or ergosterol but not with 5alpha-cholestane-3-one, suggesting that sterol is essential for the oligomerization. The treatment of carboxyfluorescein-encapsulated liposomes with ETH caused a rapid release of carboxyfluorescein into the medium. Because dextrin 20 (molecular mass, 900 Da) osmotically protected ETH-mediated hemolysis, this hemolysis is likely to be caused by pore formation on the membrane. The pore size(s) estimated from osmotic protection assays was in the range of 1.2 to 1.6 nm. The pore formed on a rabbit erythrocyte membrane was confirmed morphologically by electron microscopy. Thus, we provide evidence that ETH damages the target by the assembly of hemolysin oligomers and pore formation on the membrane. PMID:8757822

The albumin of the brown trout (Salmo trutta) is a glycoprotein.

PubMed

Metcalf, V J; Brennan, S O; Chambers, G K; George, P M

1998-07-28

The albumin from an Atlantic salmonid, the brown trout (Salmo trutta), is 1730 Da higher in molecular mass than the albumin from a Pacific salmonid, the chinook salmon (Oncorhynchus tshawytscha), at 65230 Da. Digestion with neuraminidase revealed that purified brown trout albumin contained sialic acid while chinook salmon albumin did not. Concanavalin A-sepharose affinity chromatography was used to purify a glycopeptide from a total tryptic digest of brown trout albumin. The mass of this glycopeptide (3815 Da) was determined by mass spectrometry, and the sequence largely confirmed by N-terminal sequencing. The identified sequence of IAHCCNQSYSM-, contains an Asn-Gln-Ser glycosylation site and is identical to residues 475-485 derived from the cDNA of the albumin from the Atlantic salmon, the closest relative of the brown trout. Glycosylation of albumin is very unusual, and has not been identified in either reptilian or mammalian albumins. The finding of a glycoalbumin in salmonids, ancient members of the teleost fish subclass, coupled with evidence of albumin glycosylation in the oldest vertebrates, agnathans, as well as amphibians, suggests that albumin was originally a glycoprotein, but lost this modification sometime between the divergence of amphibians and reptiles.

The butter flavorant, diacetyl, exacerbates β-amyloid cytotoxicity.

PubMed

More, Swati S; Vartak, Ashish P; Vince, Robert

2012-10-15

Diacetyl (DA), an ubiquitous butter-flavoring agent, was found to influence several aspects of amyloid-β (Aβ) aggregation--one of the two primary pathologies associated with Alzheimer's disease. Thioflavin T fluorescence and circular dichroism spectroscopic measurements revealed that DA accelerates Aβ¹⁻⁴² aggregation into soluble and ultimately insoluble β-pleated sheet structures. DA was found to covalently bind to Arg⁵ of Aβ¹⁻⁴² through proteolytic digestion-mass spectrometric experiments. These biophysical and chemical effects translated into the potentiation of Aβ¹⁻⁴² cytotoxicity by DA toward SH-SY5Y cells in culture. DA easily traversed through a MDR1-MDCK cell monolayer, an in vitro model of the blood-brain barrier. Additionally, DA was found not only to be resistant to but also inhibitory toward glyoxalase I, the primary initiator of detoxification of amyloid-promoting reactive dicarbonyl species that are generated naturally in large amounts by neuronal tissue. In light of the chronic exposure of industry workers to DA, this study raises the troubling possibility of long-term neurological toxicity mediated by DA.

Porins of Pseudomonas fluorescens MFO as fibronectin-binding proteins.

PubMed

Rebière-Huët, J; Guérillon, J; Pimenta, A L; Di Martino, P; Orange, N; Hulen, C

2002-09-24

Bacterial adherence is a complex phenomenon involving specific interactions between receptors, including matricial fibronectin, and bacterial ligands. We show here that fibronectin and outer membrane proteins of Pseudomonas fluorescens were able to inhibit adherence of P. fluorescens to fibronectin-coated wells. We identified at least six fibronectin-binding proteins with molecular masses of 70, 55, 44, 37, 32 and 28 kDa. The presence of native (32 kDa) and heat-modified forms (37 kDa) of OprF was revealed by immuno-analysis and the 44-kDa band was composed of three proteins, their N-terminal sequences showing homologies with Pseudomonas aeruginosa porins (OprD, OprE1 and OprE3).

Cytosolic distributions of highly toxic metals Cd and Tl and several essential elements in the liver of brown trout (Salmo trutta L.) analyzed by size exclusion chromatography and inductively coupled plasma mass spectrometry.

PubMed

Dragun, Zrinka; Krasnići, Nesrete; Kolar, Nicol; Filipović Marijić, Vlatka; Ivanković, Dušica; Erk, Marijana

2018-05-15

Cytosolic distributions of nonessential metals Cd and Tl and seven essential elements among compounds of different molecular masses were studied in the liver of brown trout (Salmo trutta) from the karstic Krka River in Croatia. Analyses were done by size exclusion high performance liquid chromatography and high resolution inductively coupled plasma mass spectrometry. Common feature of Cd and Tl, as highly toxic elements, was their distribution within only two narrow peaks. The increase of cytosolic Cd concentrations was reflected in marked increase of Cd elution within low molecular mass peak (maximum at ∼15 kDa), presumably containing metallothioneins (MTs), which indicated successful Cd detoxification in brown trout liver under studied exposure conditions. Contrary, the increase of cytosolic Tl concentrations was reflected in marked increase of Tl elution within high molecular mass peak (maximum at 140 kDa), which probably indicated incomplete Tl detoxification. Common feature of the majority of studied essential elements was their distribution within more peaks, often broad and not well resolved, which is consistent with their numerous physiological functions. Among observed associations of essential metals/nonmetal to proteins, the following could be singled out: Cu and Zn association to MTs, Fe association to storage protein ferritin, and Se association to compounds of very low molecular masses (<5 kDa). The obtained results present the first step towards identification of metal-binding compounds in hepatic cytosol of brown trout, and thus a significant contribution to better understanding of metal fate in the liver of that important bioindicator species. Copyright © 2018 Elsevier Ltd. All rights reserved.

Mass spectrometry based on a coupled Cooper-pair box and nanomechanical resonator system

NASA Astrophysics Data System (ADS)

Jiang, Cheng; Chen, Bin; Li, Jin-Jin; Zhu, Ka-Di

2011-10-01

Nanomechanical resonators (NRs) with very high frequency have a great potential for mass sensing with unprecedented sensitivity. In this study, we propose a scheme for mass sensing based on the NR capacitively coupled to a Cooper-pair box (CPB) driven by two microwave currents. The accreted mass landing on the resonator can be measured conveniently by tracking the resonance frequency shifts because of mass changes in the signal absorption spectrum. We demonstrate that frequency shifts induced by adsorption of ten 1587 bp DNA molecules can be well resolved in the absorption spectrum. Integration with the CPB enables capacitive readout of the mechanical resonance directly on the chip.

Matrix-assisted laser desorption ionisation time-of-flight mass spectrometry rapid detection of carbapenamase activity in Acinetobacter baumannii isolates.

PubMed

Abouseada, Noha; Raouf, May; El-Attar, Eman; Moez, Pacinte

2017-01-01

Carbapenamase-producing Acinetobacter baumannii are an increasing threat in hospitals and Intensive Care Units. Accurate and rapid detection of carbapenamase producers has a great impact on patient improvement and aids in implementation of infection control measures. In this study, we describe the use of matrix-assisted laser desorption ionisation time-of-flight mass spectrometry (MALDI TOF MS) to identify carbapenamase-producing A. baumannii isolates in up to 3 h. Isolates and Methods: A total of 50 A. baumannii isolates (of which 39 were carabapenamase producers) were tested using MALDI TOF MS. Isolates were incubated for 3 h with 0.25 mg/ml up to 2 mg/ml of imipenem (IMP) at 37°C. Supernatants were analysed by MALDI TOF to analyse peaks corresponding to IMP (300 Da) and an IMP metabolite (254 Da) using UltrafleXtreme (Bruker Daltonics, Bremen, Germany). All carbapenamase-producing isolates were evidenced by the disappearance or reduction in intensity of the 300 Da peak of IPM and the appearance of a 254 Da peak of the IPM metabolite. In isolates that did not produce carbapenamase, the IPM 300 Da peak remained intact. MALDI TOF is a promising tool in the field of diagnostic microbiology that has the ability to transfer identification and antimicrobial susceptibility testing time from days to hours.

Characterization of the Lymantria dispar nucleopolyhedrovirus 25K FP gene

Treesearch

David S. Bischoff; James M. Slavicek

1996-01-01

The Lymantria dispar nucleopolyhedrovirus (LdMNPV) gene encoding the 25K FP protein has been cloned and sequenced. The 25KFP gene codes for a 217 amino acid protein with a predicted molecular mass of 24870 Da. Expression of the 25K FP protein in a rabbit reticulocyte system generated a 27 kDa protein, in close agreement with the...

Purification and characterization of protein PC, a component of glycine reductase from Eubacterium acidaminophilum.

PubMed

Schräder, T; Andreesen, J R

1992-05-15

Protein PC of the glycine reductase from Eubacterium acidaminophilum was purified to homogeneity by chromatography on phenyl-Sepharose and Sepharose S. The apparent molecular mass of the native protein, which showed an associating/dissociating behaviour, was about 420 kDa. Sodium dodecyl sulfate/polyacrylamide gel electrophoresis of protein PC revealed two protein bands corresponding to 48 and 57 kDa, indicating an alpha 4 beta 4 composition. The smaller subunit was identified as an acetyl-group-transferring protein, the 57-kDa protein was hydrophobic. N-terminal amino acid sequences were determined for both subunits. Antibodies raised against the 48-kDa subunit showed cross-reactions with extracts of E. acidaminophilum grown on different substrates and with extracts from other glycine-utilizing anaerobic bacteria such as Clostridium purinolyticum, C. sticklandii, and C. sporogenes. The respective protein from the former two organisms corresponded in molecular mass. When protein PA was chemically carboxymethylated by iodo[2-14C]acetate and incubated with protein PC, acetyl phosphate was a reaction product, thus establishing it as the product of the glycine reductase reaction by using homogeneous preparations of these two proteins from E. acidaminophilum.

Biosynthesis and intracellular transport of the receptor for platelet-derived growth factor.

PubMed Central

Claesson-Welsh, L; Rönnstrand, L; Heldin, C H

1987-01-01

The biosynthesis of the receptor for platelet-derived growth factor (PDGF) was examined in metabolically labeled human foreskin fibroblasts. The receptor was synthesized as a 145-kDa precursor, which, when incubated with endo-beta-N-acetylglucosaminidase H (endo H), underwent a 15-kDa decrease in molecular mass. This indicates that the size of the core protein is about 130 kDa and that the 145-kDa form represents a receptor precursor carrying high-mannose N-linked oligosaccharide groups. Within 15 min after synthesis, the receptor was converted to a 165-kDa form. This form was entirely resistant to endo H treatment and probably represents a receptor molecule that has undergone further posttranslational modification, including O-linked glycosylation. Subsequently, within 30 min, a molecule of 170 kDa--i.e., the size of the mature receptor--appeared. A slightly larger molecule, of 175 kDa, which could be immunoprecipitated from PDGF-stimulated 32P-labeled cells, probably represents a receptor further modified by autophosphorylation. The 170-kDa molecule had an isoelectric point of about 4.5. Addition of PDGF increased the turnover rate of the 170-kDa PDGF receptor. Images PMID:2827155

Red light-induced formation of ubiquitin-phytochrome conjugates: Identification of possible intermediates of phytochrome degradation

PubMed Central

Shanklin, John; Jabben, Merten; Vierstra, Richard D.

1987-01-01

Phytochrome is the photoreceptor that controls red light-mediated morphogenesis in higher plants. It exists in two photointerconvertible forms, a red light-absorbing form, Pr, and a far-red light-absorbing form, Pfr. Because photoconversion of Pr to Pfr by a brief light pulse decreases the in vivo half-life of this chromoprotein by a factor of ≈100, this system offers a unique way to modulate the turnover rate of a specific protein and hence study the mechanisms responsible for selective protein degradation. In etiolated oat [Avena sativa (L.)] seedlings, degradation of phytochrome as Pfr follows zero-order kinetics as measured both spectrally and by ELISA, with 50% of Pfr lost in ≈130 min at 27°C. Immunoblot analysis of the destruction process with anti-oat phytochrome immunoglobulins reveals that degradation involves the loss of the 124-kDa phytochrome monomer and that proteolytic intermediates of apparent molecular mass lower than 124 kDa do not accumulate to detectable levels in vivo (<0.015% of total phytochrome). The latter observation suggests that proteolytic breakdown of the protein is extremely rapid. However, a series of polypeptides with higher apparent molecular mass and recognized by anti-phytochrome immunoglobulins (principally 129 and 134 kDa) appears after photoconversion to Pfr. These polypeptides represent no more than a few percent of the total immunologically detectable phytochrome pool and have incremental differences in apparent molecular mass of 5 kDa. They appear within 5 min after Pfr formation, reach maximal levels between 90 and 180 min, and decline thereafter. These polypeptides and others of apparent molecular mass up to 160 kDa are also detectable with immunoglobulins directed against either oat or human ubiquitin, indicating that they are ubiquitin-phytochrome conjugates. Since ubiquitin conjugation is involved in intracellular protein turnover and since formation and degradation of Pfr-ubiquitin conjugates coincide with the turnover of Pfr, these data suggest that the Pfr form of phytochrome is degraded via a ubiquitin-dependent proteolytic pathway. Images PMID:16593800

The left ventricle as a mechanical engine: from Leonardo da Vinci to the echocardiographic assessment of peak power output-to-left ventricular mass.

PubMed

Dini, Frank L; Guarini, Giacinta; Ballo, Piercarlo; Carluccio, Erberto; Maiello, Maria; Capozza, Paola; Innelli, Pasquale; Rosa, Gian M; Palmiero, Pasquale; Galderisi, Maurizio; Razzolini, Renato; Nodari, Savina

2013-03-01

The interpretation of the heart as a mechanical engine dates back to the teachings of Leonardo da Vinci, who was the first to apply the laws of mechanics to the function of the heart. Similar to any mechanical engine, whose performance is proportional to the power generated with respect to weight, the left ventricle can be viewed as a power generator whose performance can be related to left ventricular mass. Stress echocardiography may provide valuable information on the relationship between cardiac performance and recruited left ventricular mass that may be used in distinguishing between adaptive and maladaptive left ventricular remodeling. Peak power output-to-mass, obtained during exercise or pharmacological stress echocardiography, is a measure that reflects the number of watts that are developed by 100 g of left ventricular mass under maximal stimulation. Power output-to-mass may be calculated as left ventricular power output per 100 g of left ventricular mass: 100× left ventricular power output divided by left ventricular mass (W/100 g). A simplified formula to calculate power output-to-mass is as follows: 0.222 × cardiac output (l/min) × mean blood pressure (mmHg)/left ventricular mass (g). When the integrity of myocardial structure is compromised, a mismatch becomes apparent between maximal cardiac power output and left ventricular mass; when this occurs, a reduction of the peak power output-to-mass index is observed.

Mass-spectrometric identification of T-kininogen I/thiostatin as an acute-phase inflammatory protein suppressed by curcumin and capsaicin.

PubMed

Joe, Bina; Nagaraju, Anitha; Gowda, Lalitha R; Basrur, Venkatesha; Lokesh, Belur R

2014-01-01

Curcumin and capsaicin are dietary xenobiotics with well-documented anti-inflammatory properties. Previously, the beneficial effect of these spice principles in lowering chronic inflammation was demonstrated using a rat experimental model for arthritis. The extent of lowering of arthritic index by the spice principles was associated with a significant shift in macrophage function favoring the reduction of pro-inflammatory molecules such as reactive oxygen species and production and release of anti-inflammatory metabolites of arachidonic acid. Beyond the cellular effects on macrophage function, oral administration of curcumin and capsaicin caused alterations in serum protein profiles of rats injected with adjuvant to develop arthritis. Specifically, a 72 kDa acidic glycoprotein, GpA72, which was elevated in pre-arthritic rats, was significantly lowered by feeding either curcumin or capsaicin to the rats. Employing the tandem mass spectrometric approach for direct sequencing of peptides, here we report the identification of GpA72 as T-kininogen I also known as Thiostatin. Since T-kininogen I is an early acute-phase protein, we additionally tested the efficiency of curcumin and capsaicin to mediate the inflammatory response in an acute phase model. The results demonstrate that curcumin and capsaicin lower the acute-phase inflammatory response, the molecular mechanism for which is, in part, mediated by pathways associated with the lowering of T-kininogen I.

Synthesis, structure, vapour pressure and deposition of ZnO thin film by plasma assisted MOCVD technique using a novel precursor bis[(pentylnitrilomethylidine) (pentylnitrilomethylidine-μ-phenalato)]dizinc(II)

NASA Astrophysics Data System (ADS)

Chandrakala, C.; Sravanthi, P.; Raj Bharath, S.; Arockiasamy, S.; George Johnson, M.; Nagaraja, K. S.; Jeyaraj, B.

2017-02-01

A novel binuclear zinc schiff's base complex bis[(pentylnitrilomethylidine)(pentylnitrilomethylidine-μ-phenalato)]dizinc(II) (hereafter referred as ZSP) was prepared and used as a precursor for the deposition of ZnO thin film by MOCVD. The dynamic TG run of ZSP showed sufficient volatility and good thermal stability. The temperature dependence of vapour pressure measured by transpiration technique yielded a value of 55.8 ± 2.3 kJ mol-1 for the enthalpy of sublimation (ΔH°sub) in the temperature range of 423-503 K. The crystal structure of ZSP was solved by single crystal XRD which exhibits triclinic crystal system with the space group of Pī. The molecular mass of ZSP was determined by mass spectrometry which yielded the m/z value of 891 and 445 Da corresponding to its dimeric as well as monomeric form. The complex ZSP was further characterized by FT-IR and NMR. The demonstration of ZnO thin film deposition was carried out by using plasma assisted MOCVD. The thin film XRD confirmed the highly oriented (002) ZnO thin films on Si(100) substrate. The uniformity and composition of the thin film were analyzed by SEM/EDX. The band gap of ZnO thin film measurement indicated the blue shift with the value of 3.79 eV.

An integrative strategy for quantitative analysis of the N-glycoproteome in complex biological samples

PubMed Central

2014-01-01

Background The complexity of protein glycosylation makes it difficult to characterize glycosylation patterns on a proteomic scale. In this study, we developed an integrated strategy for comparatively analyzing N-glycosylation/glycoproteins quantitatively from complex biological samples in a high-throughput manner. This strategy entailed separating and enriching glycopeptides/glycoproteins using lectin affinity chromatography, and then tandem labeling them with 18O/16O to generate a mass shift of 6 Da between the paired glycopeptides, and finally analyzing them with liquid chromatography-mass spectrometry (LC-MS) and the automatic quantitative method we developed based on Mascot Distiller. Results The accuracy and repeatability of this strategy were first verified using standard glycoproteins; linearity was maintained within a range of 1:10–10:1. The peptide concentration ratios obtained by the self-build quantitative method were similar to both the manually calculated and theoretical values, with a standard deviation (SD) of 0.023–0.186 for glycopeptides. The feasibility of the strategy was further confirmed with serum from hepatocellular carcinoma (HCC) patients and healthy individuals; the expression of 44 glycopeptides and 30 glycoproteins were significantly different between HCC patient and control serum. Conclusions This strategy is accurate, repeatable, and efficient, and may be a useful tool for identification of disease-related N-glycosylation/glycoprotein changes. PMID:24428921

The binary Feige 24 - The mass, radius, and gravitational redshift of the DA white dwarf

NASA Technical Reports Server (NTRS)

Vennes, Stephane; Shipman, Harry L.; Thorstensen, John R.; Thejll, Peter

1991-01-01

Observations are reported which refine the binary ephemeris of the Feige 24 system, which contains a peculiar hot DA white dwarf and an M dwarf with an atmosphere illuminated by extreme ultraviolet radiation from the white dwarf. With the new ephemeris and a set of IUE high-dispersion spectra, showing phase-dependent redshifted C IV, N V, and Si IV resonance lines, the orbital velocity, and hence the mass (0.54 + or - 0.20 solar masses), and the gravitational redshift of the white dwarf (14.1 + or - 5.2 km/s) are determined independently. It is shown that the measured Einstein redshift is consistent with an estimated radius for the white dwarf obtained from a model atmosphere solid angle and a parallax measurement. This radius is twice the Hamada-Salpeter radius for the given mass and offers a prospect to investigate the presence of a massive hydrogen envelope in that white dwarf star.

[The proteomic profiling of blood serum of children with gastroesophageal reflux disease].

PubMed

Korkotashvili, L V; Kolesov, S A; Jukova, E A; Vidmanova, T A; Kankova, N Yu; Bashurova, I A; Sidorova, A M; Kulakova, E V

2015-03-01

The mass-spectra of proteome of blood serum from healthy children and children with gastroesophageal reflux disease were received. The technology platform including direct proteome mass-spectrometer profiling after pre-fractional rectification using magnetic particles MB WCX was applied. The significant differences in mass-spectra were established manifesting in detection of more mass-spectrometer peaks and higher indicators of their intensity and area in group of healthy children. The study detected 39 particular peptides and low-molecular proteins predominantly intrinsic to healthy or ill children. It was established that two peptides with molecular mass 925 and 909 Da. are registered only in healthy patients and have no traces in group ofpatients with gastroesophageal reflux disease. The peptide 1564 Da is detected only in blood of children with gastroesophageal reflux disease and totally is absent in healthy children. The research data permitted to reveal specific patterns (signatures) of low-molecular proteins and peptides specific for blood serum of healthy children and patients with gastroesophageal reflux disease. The results testify the availability of singularities in metabolism of low-molecular proteins and can be used as a basis for development of minimally invasive mass-spectrometer system for its diagnostic.

Development and Evaluation of a Reverse-Entry Ion Source Orbitrap Mass Spectrometer.

PubMed

Poltash, Michael L; McCabe, Jacob W; Patrick, John W; Laganowsky, Arthur; Russell, David H

2018-05-23

As a step towards development of a high-resolution ion mobility mass spectrometer using the orbitrap mass analyzer platform, we describe herein a novel reverse-entry ion source (REIS) coupled to the higher-energy C-trap dissociation (HCD) cell of an orbitrap mass spectrometer with extended mass range. Development of the REIS is a first step in the development of a drift tube ion mobility-orbitrap MS. The REIS approach retains the functionality of the commercial instrument ion source which permits the uninterrupted use of the instrument during development as well as performance comparisons between the two ion sources. Ubiquitin (8.5 kDa) and lipid binding to the ammonia transport channel (AmtB, 126 kDa) protein complex were used as model soluble and membrane proteins, respectively, to evaluate the performance of the REIS instrument. Mass resolution obtained with the REIS is comparable to that obtained using the commercial ion source. The charge state distributions for ubiquitin and AmtB obtained on the REIS are in agreement with previous studies which suggests that the REIS-orbitrap EMR retains native structure in the gas phase. Graphical Abstract ᅟ.

The Roles of Phasic and Tonic Dopamine in Tic Learning and Expression.

PubMed

Maia, Tiago V; Conceição, Vasco A

2017-09-15

Tourette syndrome (TS) prominently involves dopaminergic disturbances, but the precise nature of those disturbances has remained elusive. A substantial body of empirical work and recent computational models have characterized the specific roles of phasic and tonic dopamine (DA) in action learning and selection, respectively. Using insights from this work and models, we suggest that TS involves increases in both phasic and tonic DA, which produce increased propensities for tic learning and expression, respectively. We review the evidence from reinforcement-learning and habit-learning studies in TS, which supports the idea that TS involves increased phasic DA responses; we also review the evidence that tics engage the habit-learning circuitry. On the basis of these findings, we suggest that tics are exaggerated, maladaptive, and persistent motor habits reinforced by aberrant, increased phasic DA responses. Increased tonic DA amplifies the tendency to execute learned tics and also provides a fertile ground of motor hyperactivity for tic learning. We review evidence suggesting that antipsychotics may counter both the increased propensity for tic expression, by increasing excitability in the indirect pathway, and the increased propensity for tic learning, by shifting plasticity in the indirect pathway toward long-term potentiation (and possibly also through more complex mechanisms). Finally, we review evidence suggesting that low doses of DA agonists that effectively treat TS decrease both phasic and tonic DA, thereby also reducing the propensity for both tic learning and tic expression, respectively. Copyright © 2017 Society of Biological Psychiatry. Published by Elsevier Inc. All rights reserved.

Retention behavior of isomeric polycyclic aromatic sulfur heterocycles in gas chromatography on stationary phases of different selectivity

PubMed Central

Wilson, Walter B.; Sander, Lane C.; Oña-Ruales, Jorge O.; Mössner, Stephanie G.; Sidisky, Leonard M.; Lee, Milton L.; Wise, Stephen A.

2017-01-01

Retention indices for 48 polycyclic aromatic sulfur heterocycles (PASHs) were determined using gas chromatography with three different stationary phases: a 50% phenyl phase, a 50% liquid crystalline dimethylpolysiloxane (LC-DMPS) phase, and an ionic liquid (IL) phase. Correlations between the retention behavior on the three stationary phases and PASH geometry (L/B and T, i.e., length-to-breadth ratio and thickness, respectively) were investigated for the following four isomer sets: (1) 4 three-ring molecular mass (MM) 184 Da PASHs, (2) 13 four-ring MM 234 Da PASHs, (3) 10 five-ring MM 258 Da PASHs, and (4) 20 five-ring MM 284 Da PASHs. Correlation coefficients for retention on the 50% LC-DMPS vs L/B ranged from r = 0.50 (MM 284 Da) to r = 0.77 (MM 234 Da). Correlation coefficients for retention on the IL phase vs L/B ranged from r = 0.31 (MM 234 Da) to r = 0.54 (MM 284 Da). Correlation coefficients for retention on the 50% phenyl vs L/B ranged from r = 0.14 (MM 258 Da) to r = 0.59 (MM 284 Da). Several correlation trends are discussed in detail for the retention behavior of PASH on the three stationary phases. PMID:28089272

Retention behavior of isomeric polycyclic aromatic sulfur heterocycles in gas chromatography on stationary phases of different selectivity.

PubMed

Wilson, Walter B; Sander, Lane C; Oña-Ruales, Jorge O; Mössner, Stephanie G; Sidisky, Leonard M; Lee, Milton L; Wise, Stephen A

2017-02-17

Retention indices for 48 polycyclic aromatic sulfur heterocycles (PASHs) were determined using gas chromatography with three different stationary phases: a 50% phenyl phase, a 50% liquid crystalline dimethylpolysiloxane (LC-DMPS) phase, and an ionic liquid (IL) phase. Correlations between the retention behavior on the three stationary phases and PASH geometry (L/B and T, i.e., length-to-breadth ratio and thickness, respectively) were investigated for the following four isomer sets: (1) 4 three-ring molecular mass (MM) 184Da PASHs, (2) 13 four-ring MM 234Da PASHs, (3) 10 five-ring MM 258Da PASHs, and (4) 20 five-ring MM 284Da PASHs. Correlation coefficients for retention on the 50% LC-DMPS vs L/B ranged from r=0.50 (MM 284Da) to r=0.77 (MM 234Da). Correlation coefficients for retention on the IL phase vs L/B ranged from r=0.31 (MM 234Da) to r=0.54 (MM 284Da). Correlation coefficients for retention on the 50% phenyl vs L/B ranged from r=0.14 (MM 258Da) to r=0.59 (MM 284Da). Several correlation trends are discussed in detail for the retention behavior of PASH on the three stationary phases. Published by Elsevier B.V.

Identification of phosphoproteins associated with maintenance of transformed state in temperature-sensitive Rous sarcoma-virus infected cells by proteomic analysis

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

Yamaoka, Kazuko; Imajoh-Ohmi, Shinobu; Fukuda, Hiroyuki

2006-07-07

To identify phosphotyrosine-containing proteins essential for maintaining the transformed state, we studied the tyrosine phosphorylation profile of temperature-sensitive mutant of Rous sarcoma virus, tsNY68, infected cells (68N7). Shifting the temperature from 39 {sup o}C (nonpermissive) to 32 {sup o}C (permissive) markedly increased the expression of phosphotyrosine-containing cell membrane proteins of {approx}40 kDa, as assessed by SDS-PAGE. Membrane and nuclear proteins were separated by two-dimensional gel electrophoresis and immunoblotted with anti-phosphotyrosine antibody. Proteins showing temperature-dependent changes in phosphorylation profile were subjected to in-gel digestion with trypsin and analyzed by mass spectrometry. Five proteins were identified: heterogeneous nuclear ribonucleoprotein (hnRNP) A3, hnRNPmore » A2, annexin II, phosphoglycerate mutase 1, and triosephosphate isomerase 1. hnRNP A3 was phosphorylated at serine residues and had both serine and tyrosine phosphorylated sites. These results suggest an important complementary role for proteomics in identifying molecular abnormalities associated with tumor progression that may be attractive candidates for tumor diagnosis.« less

Characterization and enzymatic hydrolysis of hydrothermally treated β-1,3-1,6-glucan from Aureobasidium pullulans.

PubMed

Hirabayashi, Katsuki; Kondo, Nobuhiro; Hayashi, Sachio

2016-12-01

The chemical structure of hydrothermally treated β-1,3-1,6-glucan from Aureobasidium pullulans was characterized using techniques such as gas chromatography/mass spectrometry (GC/MS) and nuclear magnetic resonance (NMR). The chemical shifts of anomeric carbons observed in the 13 C-NMR spectra suggested the presence of single flexible chains of polysaccharide in the sample. β-1,3-1,6-Glucan from A. pullulans became water-soluble, with an average molecular weight of 128,000 Da after hydrothermal treatment, and the solubility in water was approximately 10% (w/w). Sample (3% w/v) was completely hydrolyzed to glucose by enzymatic reaction with Lysing enzymes from Trichoderma harzianum. Gentiobiose (Glcβ1 → 6Glc) and glucose were released as products during the reaction, and the maximum yield of gentiobiose was approximately 70% (w/w). The molar ratio of gentiobiose to glucose after 1 h reaction suggested that the sample is likely highly branched. Sample (3% w/v) was also hydrolyzed to glucose by Uskizyme from Trichoderma sp., indicating that it is very sensitive to enzymatic hydrolysis.

Shift work and age as interactive predictors of body mass index among offshore workers.

PubMed

Parkes, Katharine R

2002-02-01

This study investigated shift pattern (day shifts versus day-night rotation) and its interactions with age, and with years of shiftwork exposure, as predictors of body mass index (BMI). Survey data were collected from offshore personnel working day shifts (N=787) or day-night shifts (N=787); information was obtained about shift pattern and years of shiftwork exposure, height, weight, demographic factors, and smoking habits. Hierarchical multiple regression was used to test a model in which BMI was predicted by additive and interactive effects of shift pattern, age, and exposure years with control for confounding variables. In a multivariate analysis (controlling for job type, education and smoking), BMI was predicted by the main effects of age and years of shiftwork exposure. Shift pattern was not significant as a main effect, but it interacted significantly with the curvilinear age term and with the linear and curvilinear components of shiftwork exposure. In the day shift group, age but not exposure predicted BMI; the opposite was true of the day-night shift group. The increase in BMI with an increase in age and exposure years was steeper for the day-night shift group than for the day shift group. The significant interaction effects found in this study were consistent with the view that continued exposure to day-night shift work gives rise to increases in BMI, over and above the normative effects of ageing on BMI shown by day-shift workers.

Effects of Pubertal Cannabinoid Administration on Attentional Set-Shifting and Dopaminergic Hyper-Responsivity in a Developmental Disruption Model of Schizophrenia

PubMed Central

Guimarães, Francisco S.; Grace, Anthony A.

2015-01-01

Background: Adolescent exposure to cannabinoids in vulnerable individuals is proposed to be a risk factor for psychiatric conditions later in life, particularly schizophrenia. Evidence from studies in animals has indicated that a combination of repeated pubertal cannabinoid administration with either neonatal prefrontocortical lesion, isolation rearing, or chronic NMDA receptor antagonism administration induces enhanced schizophrenia-like behavioral disruptions. The effects of adolescent exposure to CB1 receptor agonists, however, have not been tested in a developmental disruption model of schizophrenia. Methods: This was tested in the methylazoxymethanol (MAM) model, in which repeated treatment with the synthetic cannabinoid agonist WIN 55,212-2 (WIN; 1.2mg/kg) was extended over 25 days throughout puberty (postnatal days 40–65) in control and MAM rats. The rats received 20 injections, which were delivered irregularly to mimic the human condition. Adult rats were tested for attentional set-shifting task and locomotor response to amphetamine, which was compared with in vivo recording from ventral tegmental area (VTA) dopamine (DA) neurons. Results: MAM-treated rats showed impairment in the attentional set-shifting task, augmented locomotor response to amphetamine administration, and an increased number of spontaneously active DA neurons in the VTA. Interestingly, pubertal WIN treatment in normal animals induced similar changes at adulthood as those observed in MAM-treated rats, supporting the notion that adolescence exposure to cannabinoids may represent a risk factor for developing schizophrenia-like signs at adulthood. However, contrary to expectations, pubertal WIN administration did not exacerbate the behavioral and electrophysiological changes in MAM-treated rats beyond that observed in WIN-treated saline rats (Sal). Indeed, WIN treatment actually attenuated the locomotor response to amphetamine in MAM rats without impacting DA neuron activity states. Conclusions: Taken together, the present results indicate that the impact of cannabinoids during puberty/adolescence on schizophrenia models is more complex than may be predicted. PMID:25522381

Enhanced peroxydisulfate electrochemiluminescence for dopamine biosensing based on Au nanoparticle decorated reduced graphene oxide.

PubMed

Yan, Yuting; Liu, Qian; Wang, Kun; Jiang, Ling; Yang, Xingwang; Qian, Jing; Dong, Xiaoya; Qiu, Baijing

2013-12-07

This work reports a novel strategy to amplify the electrochemiluminescence (ECL) signal of peroxydisulfate solution based on the Au nanoparticle decorated reduced graphene oxide (Au NP-RGO), and further an ECL biosensor for sensitive and selective detection of dopamine (DA) was constructed. Due to the synergistic amplification of Au NPs and RGO, the ECL signal of peroxydisulfate solution on the Au NP-RGO modified electrode was about 5-fold enhanced compared to that of the bare electrode with the ECL onset potential positively shifted from -1.2 V to -0.9 V. More interestingly, the ECL intensity of peroxydisulfate solution increased with the increase of DA concentration, based on which an ECL biosensor for DA determination was fabricated. The as-prepared solid-state ECL DA sensor showed a wide linear response of 0.02-40 μM with a detection limit of 6.7 nM (S/N = 3). Moreover, we expect this work would open up a new field in the application of peroxydisulfate solution ECL for highly sensitive bioassays.

Experimental Evidence for Space-Charge Effects between Ions of the Same Mass-to-Charge in Fourier-Transform Ion Cyclotron Resonance Mass Spectrometry

PubMed Central

Wong, Richard L.; Amster, I. Jonathan

2009-01-01

It is often stated that ions of the same mass-to-charge do not induce space-charge frequency shifts among themselves in an ion cyclotron resonance mass spectrometry measurement. Here, we demonstrate space-charge induced frequency shifts for ions of a single mass-to-charge. The monoisotopic atomic ion, Cs+, was used for this study. The measured frequency is observed to decrease linearly with an increase in the number of ions, as has been reported previously for space-charge effects between ions of different mass-to-charge. The frequency shift between ions of the same m/z value are compared to that induced between ions of different m/z value, and is found to be 7.5 times smaller. Control experiments were performed to ensure that the observed space-charge effects are not artifacts of the measurement or of experimental design. The results can be rationalized by recognizing that the electric forces between ions in a magnetic field conform to the weak form of the Newton's third law, where the action and reaction forces do not cancel exactly. PMID:19562102

Occupational heat strain in a hot underground metal mine.

PubMed

Lutz, Eric A; Reed, Rustin J; Turner, Dylan; Littau, Sally R

2014-04-01

In a hot underground metal mine, this study evaluated the relationship between job task, physical body type, work shift, and heat strain. Thirty-one miners were evaluated during 98 shifts while performing deep shaft-sinking tasks. Continuous core body temperature, heart rate, pre- and postshift urine specific gravity (USG), and body mass index were measured. Cutting and welding tasks were associated with significantly (P < 0.05) increased core body temperature, maximum heart rate, and increased postshift urine specific gravity. Miners in the obese level II and III body mass index categories, as well as those working night shift, had lower core body temperatures (P < 0.05). This study confirms that job task, body type, and shift are risk factors for heat strain.

Low molecular weight components in an aquatic humic substance as characterized by membrane dialysis and orbitrap mass spectrometry.

PubMed

Remucal, Christina K; Cory, Rose M; Sander, Michael; McNeill, Kristopher

2012-09-04

Suwannee River fulvic acid (SRFA) was dialyzed through a 100-500 molecular weight cutoff dialysis membrane, and the dialysate and retentate were analyzed by UV-visible absorption and high-resolution Orbitrap mass spectrometry (MS). A significant fraction (36% based on dissolved organic carbon) of SRFA passed through the dialysis membrane. The fraction of SRFA in the dialysate had a different UV-visible absorption spectrum and was enriched in low molecular weight molecules with a more aliphatic composition relative to the initial SRFA solution. Comparison of the SRFA spectra collected by Orbitrap MS and Fourier transform ion cyclotron resonance MS (FT-ICR MS) demonstrated that the mass accuracy of the Orbitrap MS is sufficient for determination of unique molecular formulas of compounds with masses <600 Da in a complex mixture, such as SRFA. The most intense masses detected by Orbitrap MS were found in the 100-200 Da mass range. Many of these low molecular masses corresponded to molecular formulas of previously identified compounds in organic matter, lignin, and plants, and the use of the standard addition method provided an upper concentration estimate of selected target compounds in SRFA. Collectively, these results provide evidence that SRFA contains low molecular weight components that are present individually or in loosely bound assemblies.

A Robust Two-Dimensional Separation for Top-Down Tandem Mass Spectrometry of the Low-Mass Proteome

PubMed Central

Lee, Ji Eun; Kellie, John F.; Tran, John C.; Tipton, Jeremiah D.; Catherman, Adam D.; Thomas, Haylee M.; Ahlf, Dorothy R.; Durbin, Kenneth R.; Vellaichamy, Adaikkalam; Ntai, Ioanna; Marshall, Alan G.; Kelleher, Neil L.

2010-01-01

For fractionation of intact proteins by molecular weight (MW), a sharply improved two-dimensional (2D) separation is presented to drive reproducible and robust fractionation before top-down mass spectrometry of complex mixtures. The “GELFrEE” (i.e., gel-eluted liquid fraction entrapment electrophoresis) approach is implemented by use of Tris-glycine and Tris-tricine gel systems applied to human cytosolic and nuclear extracts from HeLa S3 cells, to achieve a MW-based fractionation of proteins from 5 to >100 kDa in 1 h. For top-down tandem mass spectroscopy (MS/MS) of the low-mass proteome (5–25 kDa), between 5 and 8 gel-elution (GE) fractions are sampled by nanocapillary-LC-MS/MS with 12 or 14.5 tesla Fourier transform ion cyclotron resonance (FT-ICR) mass spectrometers. Single injections give about 40 detectable proteins, about half of which yield automated ProSight identifications. Reproducibility metrics of the system are presented, along with comparative analysis of protein targets in mitotic versus asynchronous cells. We forward this basic 2D approach to facilitate wider implementation of top-down mass spectrometry and a variety of other protein separation and/or characterization approaches. PMID:19747844

Using time-varying asymptotic length and body condition of top piscivores to indicate ecosystem regime shift in the main basin of Lake Huron: a Bayesian hierarchical modeling approach

USGS Publications Warehouse

He, Ji X.; Bence, James R.; Roseman, Edward F.; Fielder, David G.; Ebener, Mark P.

2015-01-01

We evaluated the ecosystem regime shift in the main basin of Lake Huron that was indicated by the 2003 collapse of alewives, and dramatic declines in Chinook salmon abundance thereafter. We found that the period of 1995-2002 should be considered as the early phase of the final regime shift. We developed two Bayesian hierarchical models to describe time-varying growth based on the von Bertalanffy growth function and the length-mass relationship. We used asymptotic length as an index of growth potential, and predicted body mass at a given length as an index of body condition. Modeling fits to length and body mass at age of lake trout, Chinook salmon, and walleye were excellent. Based on posterior distributions, we evaluated the shifts in among-year geometric means of the growth potential and body condition. For a given top piscivore, one of the two indices responded to the regime shift much earlier than the 2003 collapse of alewives, the other corresponded to the 2003 changes, and which index provided the early signal differed among the three top piscivores.

Interactions of green coffee bean phenolics with wheat bread matrix in a model of simulated in vitro digestion.

PubMed

Świeca, Michał; Gawlik-Dziki, Urszula; Sęczyk, Łukasz; Dziki, Dariusz; Sikora, Małgorzata

2018-08-30

Interactions of phenolics from green coffee bean flour (GCS) with the matrix of wheat bread have been studied employing direct (electrophoretic and chromatographic techniques) and indirect tests (nutrient digestibility). According to the chromatograms of digests, the antiradical activity of enriched bread was exhibited by free phenolics. An increase the area of chromatograms and some additional peaks observed for enriched bread may confirm some interactions of proteins with phenolics. The electrophoretic profile of these extracts showed that the band corresponding to a protein with molecular mass of 38 kDA had much higher intensity in enriched bread. Electrophoretic analysis of pellets remaining after digestion revealed GCS dose-dependent differences in bands corresponding to proteins with molecular masses of 52 kDa and 23 kDa. The relative digestibility of both starch and proteins was slightly decreased by addition of GCS; however, these changes did not exceed 10%, which justifies the use of this functional material. Copyright © 2018 Elsevier Ltd. All rights reserved.

A comparative study on seismic response of two unstable rock slopes within same tectonic setting but different activity level

NASA Astrophysics Data System (ADS)

Kleinbrod, Ulrike; Burjánek, Jan; Hugentobler, Marc; Amann, Florian; Fäh, Donat

2017-12-01

In this study, the seismic response of two slope instabilities is investigated with seismic ambient vibration analysis. Two similar sites have been chosen: an active deep-seated slope instability at Cuolm da Vi and the geologically, structurally and morphologically similar, but presently not moving Alp Caschlè slope. Both slopes are located at the upper Vorderrheintal (Canton Graubünden, Switzerland). Ambient vibrations were recorded on both slopes and processed by time-frequency polarization and site-to-reference spectral ratio analysis. The data interpretation shows correlations between degree of disintegration of the rock mass and amplification. However, the ambient vibration analysis conducted, does not allow retrieving a resonance frequency that can be related to the total depth of the instability of Cuolm da Vi. Even though seismic waves can be hardly traced in rock instabilities containing open fractures, it was possible to retrieve a dispersion curve and a velocity profile from the array measurement at Cuolm da Vi due to the high level of disintegration of the rock material down to a depth of about 100 m. From the similar amplification pattern at the two sites, we expect a similar structure, indicating that also the slope at Alp Caschlè was active in the past in a similar manner as Cuolm da Vi. However, a smoother increase of amplification with frequency is observed at Alp Caschlè, which might indicate less disintegration of the rock mass in a particular depth range at this site, when comparing to Cuolm da Vi where a high level of disintegration is observed, resulting from the high activity at the slope. From the frequency-dependent amplification, we can distinguish between two parts within both instabilities, one part showing decreasing disintegration of the rock mass with increasing depth, for the other parts less-fractured blocks are observed. Since the block structures are found in the lower part of the instabilities, they might contribute to the stability of the slopes. Using the velocity profiles, it was possible to estimate the depth of the two largest open fractures (i.e. tension cracks) at Cuolm da Vi.

Mass sensing based on a circuit cavity electromechanical system

NASA Astrophysics Data System (ADS)

Jiang, Cheng; Chen, Bin; Li, Jin-Jin; Zhu, Ka-Di

2011-10-01

We present a scheme for mass sensing based on a circuit cavity electromechanical system where a free-standing, flexible aluminium membrane is capacitively coupled to a superconducting microwave cavity. Integration with the microwave cavity enables capacitive readout of the mechanical resonance directly on the chip. A microwave pump field and a second probe field are simultaneously applied to the cavity. The accreted mass landing on the membrane can be measured conveniently by tracking the mechanical resonance frequency shifts due to mass changes in the probe transmission spectrum. The mass responsivity for the membrane is 0.72 Hz/ag and we demonstrate that frequency shifts induced by adsorption of one hundred 1587 bp DNA molecules can be well resolved in the probe transmission spectrum.

Identification of a 27.8 kDa protein from flounder gill cells involved in lymphocystis disease virus binding and infection.

PubMed

Wang, Mu; Sheng, Xiu-Zhen; Xing, Jing; Tang, Xiao-Qian; Zhan, Wen-Bin

2011-03-16

In vitro, lymphocystis disease virus (LCDV) infection of flounder gill (FG) cell cultures causes obvious cytopathic effect (CPE). We describe attempts to isolate and characterize the LCDV-binding molecule(s) on the plasma membrane of FG cells that were responsible for virus entry. The results showed that the co-immunoprecipitation assay detected a 27.8 kDa molecule from FG cells that bound to LCDV. In a blocking ELISA, pre-incubation of FG cell membrane proteins with the specific antiserum developed against the 27.8 kDa protein could block LCDV binding. Similarly, antiserum against 27.8 kDa protein could also inhibit LCDV infection of FG cells in vitro. Mass spectrometric analysis established that the 27.8 kDa protein and beta-actin had a strong association. These results strongly supported the possibility that the 27.8 kDa protein was the putative receptor specific for LCDV infection of FG cells.

Chemical and biological activities of a 64-kilodalton outer sheath protein from Treponema denticola strains.

PubMed Central

Weinberg, A; Holt, S C

1991-01-01

This study examined the distribution of the major outer sheath proteins (MOSP) in several Treponema denticola strains and reports the isolation of a 64-kDa protein from the outer sheath of human clinical isolate T. denticola GM-1. The outer sheath was isolated by freeze-thaw procedures, and the distribution of outer sheath proteins was examined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). T. denticola GM-1, MS25, SR-5, and three low-passage clinical isolates possessed an MOSP with a relative molecular mass of 60 to 64 kDa. This MOSP was absent in T. denticola ATCC 35404 (TD-4) and clinical isolate SR-4. The latter possessed an MOSP of 58 kDa. 125I labeling revealed both MOSP to be dissociated forms of higher-molecular-mass oligomeric units between 116 and 162 kDa. Two-dimensional SDS-PAGE confirmed the modifiability of these MOSP. Isoelectric focusing of the 64-kDa MOSP indicated a pI of 6.7. Immunoblots with antiserum to GM-1 whole cells revealed the 64-kDa protein to be immunogenic and not cross-reactive with the MOSP of TD-4 or SR-4, and monospecific antibody to the 64-kDa protein recognized common epitopes on the high-molecular-weight oligomeric protein. These antibodies did not react with any component of TD-4 whole cells in immunoblots or in immunogold electron microscopy. Fab fragments inhibited the adherence of T. denticola GM-1 to human gingival fibroblasts by 78% (1:1,600; 0.72 micrograms of protein per ml), while TD-4 adherence was not inhibited. Amino acid analysis revealed a slightly acidic protein, devoid of cysteine, with 36% hydrophobic residues. Cyanogen bromide fragmentation of the 64-kDa protein revealed that a 42-kDa fragment contained a T-L-D-L-A-L-D segment which was 100% homologous with an integrin alpha subunit of a human leukocyte adhesion glycoprotein p 150,95. Images FIG. 1 FIG. 2 FIG. 3 FIG. 4 FIG. 5 FIG. 6 FIG. 7 FIG. 8 FIG. 9 FIG. 10 FIG. 11 FIG. 12 FIG. 13 FIG. 14 PMID:1938897

Functional and Structural Characterization of a Thermostable Phospholipase A2 from a Sparidae Fish (Diplodus annularis).

PubMed

Smichi, Nabil; Othman, Houcemeddine; Achouri, Neila; Noiriel, Alexandre; Arondel, Vincent; Srairi-Abid, Najet; Abousalham, Abdelkarim; Gargouri, Youssef; Miled, Nabil; Fendri, Ahmed

2017-03-22

Novel phospholipase (PLA 2 ) genes from the Sparidae family were cloned. The sequenced PLA 2 revealed an identity with pancreatic PLA 2 group IB. To better understand the structure/function relationships of these enzymes and their evolution, the Diplodus annularis PLA 2 (DaPLA 2 ) was overexpressed in E. coli. The refolded enzyme was purified by Ni-affinity chromatography and has a molecular mass of 15 kDa as determined by MALDI-TOF spectrometry. Interestingly, unlike the pancreatic type, the DaPLA 2 was active and stable at higher temperatures, which suggests its great potential in biotechnological applications. The 3D structure of DaPLA 2 was constructed to gain insights into the functional properties of sparidae PLA 2 . Molecular docking and dynamic simulations were performed to explain the higher thermal stability and the substrate specificity of DaPLA 2 . Using the monolayer technique, the purified DaPLA 2 was found to be active on various phospholipids ranging from 10 to 20 mN·m -1 , which explained the absence of the hemolytic activity for DaPLA 2 .

70 DA White Dwarfs Identified in LAMOST Pilot Survey

NASA Astrophysics Data System (ADS)

Zhao, J. K.; Luo, A. L.; Oswalt, T. D.; Zhao, G.

2013-06-01

We present a spectroscopically identified catalog of 70 DA white dwarfs (WDs) from the LAMOST pilot survey. Thirty-five are found to be new identifications after cross-correlation with the Eisenstein et al. and Villanova catalogs. The effective temperature and gravity of these WDs are estimated by Balmer lines fitting. Most of them are hot WDs. The cooling times and masses of these WDs are estimated by interpolation in theoretical evolution tracks. The peak of the mass distribution is found to be ~0.6 M ⊙, which is consistent with prior work in the literature. The distances of these WDs are estimated using the method of synthetic spectral distances. All of these WDs are found to be in the Galactic disk from our analysis of space motions. Our sample supports the expectation that WDs with high mass are concentrated near the plane of the Galactic disk.

70 DA WHITE DWARFS IDENTIFIED IN LAMOST PILOT SURVEY

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

Zhao, J. K.; Luo, A. L.; Zhao, G.

2013-06-01

We present a spectroscopically identified catalog of 70 DA white dwarfs (WDs) from the LAMOST pilot survey. Thirty-five are found to be new identifications after cross-correlation with the Eisenstein et al. and Villanova catalogs. The effective temperature and gravity of these WDs are estimated by Balmer lines fitting. Most of them are hot WDs. The cooling times and masses of these WDs are estimated by interpolation in theoretical evolution tracks. The peak of the mass distribution is found to be {approx}0.6 M {sub Sun }, which is consistent with prior work in the literature. The distances of these WDs aremore » estimated using the method of synthetic spectral distances. All of these WDs are found to be in the Galactic disk from our analysis of space motions. Our sample supports the expectation that WDs with high mass are concentrated near the plane of the Galactic disk.« less

Biomarkers of Aspergillus spores

NASA Astrophysics Data System (ADS)

Sulc, Miroslav; Peslova, Katerina; Zabka, Martin; Hajduch, Marian; Havlicek, Vladimir

2009-02-01

We applied both matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectrometric and 1D sodium dodecylsulfate polyacrylamide gel electrophoretic (1D-PAGE) approaches for direct analysis of intact fungal spores of twenty four Aspergillus species. In parallel, we optimized various protocols for protein extraction from Aspergillus spores using acidic conditions, step organic gradient and variable sonication treatment. The MALDI-TOF mass spectra obtained from optimally prepared samples provided a reproducible fingerprint demonstrating the capability of the MALDI-TOF approach to type and characterize different fungal strains within the Aspergillus genus. Mass spectra of intact fungal spores provided signals mostly below 20 kDa. The minimum material amount represented 0.3 [mu]g (10,000 spores). Proteins with higher molecular weight were detected by 1D-PAGEE Eleven proteins were identified from three selected strains in the range 5-25 kDa by the proteomic approach. Hemolysin and hydrophobin have the highest relevance in host-pathogen interactions.

Identification of a low digestibility δ-Conglutin in yellow lupin (Lupinus luteus L.) seed meal for atlantic salmon (Salmo salar L.) by coupling 2D-PAGE and mass spectrometry.

PubMed

Ogura, Takahiro; Hernández, Adrián; Aizawa, Tomoko; Ogihara, Jun; Sunairi, Michio; Alcaino, Javier; Salvo-Garrido, Haroldo; Maureira-Butler, Iván J

2013-01-01

The need of quality protein in the aquaculture sector has forced the incorporation of alternative plant proteins into feeding diets. However, most plant proteins show lower digestibility levels than fish meal proteins, especially in carnivorous fishes. Manipulation of protein content by plant breeding can improve the digestibility rate of plant proteins in fish, but the identification of low digestibility proteins is essential. A reduction of low digestibility proteins will not only increase feed efficiency, but also reduce water pollution. Little is known about specific digestible protein profiles and/or molecular identification of more bioavailable plant proteins in fish diets. In this study, we identified low digestibility L. luteus seed proteins using Atlantic salmon (Salmo salar) crude digestive enzymes in an in vitro assay. Low digestibility proteins were identified by comparing SDS-PAGE banding profiles of digested and non-digested lupin seed proteins. Gel image analysis detected a major 12 kDa protein band in both lupin meal and protein isolate digested products. The 12 kDa was confirmed by 2D-PAGE gels and the extracted protein was analyzed with an ion trap mass spectrometer in tandem mass mode. The MS/MS data showed that the 12 kDa low digestibility protein was a large chain δconglutin, a common seed storage protein of yellow lupin. Comparison of the protein band profiles between lupin meal and protein isolates showed that the isolatation process did not affect the low digestibility of the 12 kDa protein.

Identification of a Low Digestibility δ-Conglutin in Yellow Lupin (Lupinus luteus L.) Seed Meal for Atlantic Salmon (Salmo salar L.) by Coupling 2D-PAGE and Mass Spectrometry

PubMed Central

Ogura, Takahiro; Hernández, Adrián; Aizawa, Tomoko; Ogihara, Jun; Sunairi, Michio; Alcaino, Javier; Salvo-Garrido, Haroldo; Maureira-Butler, Iván J.

2013-01-01

The need of quality protein in the aquaculture sector has forced the incorporation of alternative plant proteins into feeding diets. However, most plant proteins show lower digestibility levels than fish meal proteins, especially in carnivorous fishes. Manipulation of protein content by plant breeding can improve the digestibility rate of plant proteins in fish, but the identification of low digestibility proteins is essential. A reduction of low digestibility proteins will not only increase feed efficiency, but also reduce water pollution. Little is known about specific digestible protein profiles and/or molecular identification of more bioavailable plant proteins in fish diets. In this study, we identified low digestibility L. luteus seed proteins using Atlantic salmon (Salmo salar) crude digestive enzymes in an in vitro assay. Low digestibility proteins were identified by comparing SDS-PAGE banding profiles of digested and non-digested lupin seed proteins. Gel image analysis detected a major 12 kDa protein band in both lupin meal and protein isolate digested products. The 12 kDa was confirmed by 2D-PAGE gels and the extracted protein was analyzed with an ion trap mass spectrometer in tandem mass mode. The MS/MS data showed that the 12 kDa low digestibility protein was a large chain δconglutin, a common seed storage protein of yellow lupin. Comparison of the protein band profiles between lupin meal and protein isolates showed that the isolatation process did not affect the low digestibility of the 12 kDa protein. PMID:24278278

Comparative analysis of monoclonal antibody N-glycosylation using stable isotope labelling and UPLC-fluorescence-MS.

PubMed

Millán Martín, Silvia; Delporte, Cédric; Farrell, Amy; Navas Iglesias, Natalia; McLoughlin, Niaobh; Bones, Jonathan

2015-03-07

A twoplex method using (12)C6 and (13)C6 stable isotope analogues (Δmass = 6 Da) of 2-aminobenzoic acid (2-AA) is described for quantitative analysis of N-glycans present on monoclonal antibodies and other glycoproteins using ultra performance liquid chromatography with sequential fluorescence and accurate mass tandem quadrupole time of flight (QToF) mass spectrometric detection.

Role of the strange quark in the rho(770) meson

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

Molina Peralta, Raquel; Guo, Dehua; Hu, B.

2017-03-01

Recently, the GWU lattice group has evaluated high-precision phase-shift data formore » $$\\pi\\pi$$ scattering in the $I = 1$, $J = 1$ channel. Unitary Chiral Perturbation Theory describes these data well around the resonance region and for different pion masses. Moreover, it allows to extrapolate to the physical point and estimate the effect of the missing $$K\\bar{K}$$ channel in the two-flavor lattice calculation. The absence of the strange quark in the lattice data leads to a lower $$\\rho$$ mass, and the analysis with U$$\\chi$$PT shows that the $$K \\bar{K}$$ channel indeed pushes the $$\\pi\\pi$$-scattering phase shift upward, having a surprisingly large effect on the $$\\rho$$-mass. The inelasticity is shown to be compatible with the experimental data. The analysis is then extended to all available two-flavor lattice simulations and similar mass shifts are observed. Chiral extrapolations of $$N_f = 2 + 1$$ lattice simulations for the $$\\rho(770)$$ are also reported.« less

Analysis of amyloid fibrils in the cheetah (Acinonyx jubatus).

PubMed

Bergström, Joakim; Ueda, Mitsuharu; Une, Yumi; Sun, Xuguo; Misumi, Shogo; Shoji, Shozo; Ando, Yukio

2006-06-01

Recently, a high prevalence of amyloid A (AA) amyloidosis has been documented among captive cheetahs worldwide. Biochemical analysis of amyloid fibrils extracted from the liver of a Japanese captive cheetah unequivocally showed that protein AA was the main fibril constituent. Further characterization of the AA fibril components by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blot analysis revealed three main protein AA bands with approximate molecular weights of 8, 10 and 12 kDa. Mass spectrometry analysis of the 12-kDa component observed in SDS-PAGE and Western blotting confirmed the molecular weight of a 12,381-Da peak. Our finding of a 12-kDa protein AA component provides evidence that the cheetah SAA sequence is longer than the previously reported 90 amino acid residues (approximately 10 kDa), and hence SAA is part of the amyloid fibril.

Controlling the migration behaviors of vascular smooth muscle cells by methoxy poly(ethylene glycol) brushes of different molecular weight and density.

PubMed

Wu, Jindan; Mao, Zhengwei; Gao, Changyou

2012-01-01

Cell migration is an important biological activity. Regulating the migration of vascular smooth muscle cells (VSMCs) is critical in tissue engineering and therapy of cardiovascular disease. In this work, methoxy poly(ethylene glycol) (mPEG) brushes of different molecular weight (Mw 2 kDa, 5 kDa and 10 kDa) and grafting mass (0-859 ng/cm(2)) were prepared on aldehyde-activated glass slides, and were characterized by X-ray photoelectron spectrometer (XPS) and quartz crystal microbalance with dissipation (QCM-d). Adhesion and migration processes of VSMCs were studied as a function of different mPEG Mw and grafting density. We found that these events were mainly regulated by the grafting mass of mPEG regardless of mPEG Mw and grafting density. The VSMCs migrated on the surfaces randomly without a preferential direction. Their migration rates increased initially and then decreased along with the increase of mPEG grafting mass. The fastest rates (~24 μm/h) appeared on the mPEG brushes with grafting mass of 300-500 ng/cm(2) depending on the Mw. Cell adhesion strength, arrangement of cytoskeleton, and gene and protein expression levels of adhesion related proteins were studied to unveil the intrinsic mechanism. It was found that the cell-substrate interaction controlled the cell mobility, and the highest migration rate was achieved on the surfaces with appropriate adhesion force. Copyright © 2011 Elsevier Ltd. All rights reserved.

MALDI-TOF-MS with PLS Modeling Enables Strain Typing of the Bacterial Plant Pathogen Xanthomonas axonopodis

NASA Astrophysics Data System (ADS)

Sindt, Nathan M.; Robison, Faith; Brick, Mark A.; Schwartz, Howard F.; Heuberger, Adam L.; Prenni, Jessica E.

2018-02-01

Matrix-assisted desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS) is a fast and effective tool for microbial species identification. However, current approaches are limited to species-level identification even when genetic differences are known. Here, we present a novel workflow that applies the statistical method of partial least squares discriminant analysis (PLS-DA) to MALDI-TOF-MS protein fingerprint data of Xanthomonas axonopodis, an important bacterial plant pathogen of fruit and vegetable crops. Mass spectra of 32 X. axonopodis strains were used to create a mass spectral library and PLS-DA was employed to model the closely related strains. A robust workflow was designed to optimize the PLS-DA model by assessing the model performance over a range of signal-to-noise ratios (s/n) and mass filter (MF) thresholds. The optimized parameters were observed to be s/n = 3 and MF = 0.7. The model correctly classified 83% of spectra withheld from the model as a test set. A new decision rule was developed, termed the rolled-up Maximum Decision Rule (ruMDR), and this method improved identification rates to 92%. These results demonstrate that MALDI-TOF-MS protein fingerprints of bacterial isolates can be utilized to enable identification at the strain level. Furthermore, the open-source framework of this workflow allows for broad implementation across various instrument platforms as well as integration with alternative modeling and classification algorithms.

Paper spray mass spectrometry and PLS-DA improved by variable selection for the forensic discrimination of beers.

PubMed

Pereira, Hebert Vinicius; Amador, Victória Silva; Sena, Marcelo Martins; Augusti, Rodinei; Piccin, Evandro

2016-10-12

Paper spray mass spectrometry (PS-MS) combined with partial least squares discriminant analysis (PLS-DA) was applied for the first time in a forensic context to a fast and effective differentiation of beers. Eight different brands of American standard lager beers produced by four different breweries (141 samples from 55 batches) were studied with the aim at performing a differentiation according to their market prices. The three leader brands in the Brazilian beer market, which have been subject to fraud, were modeled as the higher-price class, while the five brands most used for counterfeiting were modeled as the lower-price class. Parameters affecting the paper spray ionization were examined and optimized. The best MS signal stability and intensity was obtained while using the positive ion mode, with PS(+) mass spectra characterized by intense pairs of signals corresponding to sodium and potassium adducts of malto-oligosaccharides. Discrimination was not apparent neither by using visual inspection nor principal component analysis (PCA). However, supervised classification models provided high rates of sensitivity and specificity. A PLS-DA model using full scan mass spectra were improved by variable selection with ordered predictors selection (OPS), providing 100% of reliability rate and reducing the number of variables from 1701 to 60. This model was interpreted by detecting fifteen variables as the most significant VIP (variable importance in projection) scores, which were therefore considered diagnostic ions for this type of beer counterfeit. Copyright © 2016 Elsevier B.V. All rights reserved.

Intervention in gene regulatory networks with maximal phenotype alteration.

PubMed

Yousefi, Mohammadmahdi R; Dougherty, Edward R

2013-07-15

A basic issue for translational genomics is to model gene interaction via gene regulatory networks (GRNs) and thereby provide an informatics environment to study the effects of intervention (say, via drugs) and to derive effective intervention strategies. Taking the view that the phenotype is characterized by the long-run behavior (steady-state distribution) of the network, we desire interventions to optimally move the probability mass from undesirable to desirable states Heretofore, two external control approaches have been taken to shift the steady-state mass of a GRN: (i) use a user-defined cost function for which desirable shift of the steady-state mass is a by-product and (ii) use heuristics to design a greedy algorithm. Neither approach provides an optimal control policy relative to long-run behavior. We use a linear programming approach to optimally shift the steady-state mass from undesirable to desirable states, i.e. optimization is directly based on the amount of shift and therefore must outperform previously proposed methods. Moreover, the same basic linear programming structure is used for both unconstrained and constrained optimization, where in the latter case, constraints on the optimization limit the amount of mass that may be shifted to 'ambiguous' states, these being states that are not directly undesirable relative to the pathology of interest but which bear some perceived risk. We apply the method to probabilistic Boolean networks, but the theory applies to any Markovian GRN. Supplementary materials, including the simulation results, MATLAB source code and description of suboptimal methods are available at http://gsp.tamu.edu/Publications/supplementary/yousefi13b. edward@ece.tamu.edu Supplementary data are available at Bioinformatics online.

Statistical analysis of fragmentation patterns of electron ionization mass spectra of enolized-trimethylsilylated anabolic androgenic steroids

NASA Astrophysics Data System (ADS)

Fragkaki, A. G.; Angelis, Y. S.; Tsantili-Kakoulidou, A.; Koupparis, M.; Georgakopoulos, C.

2009-08-01

Anabolic androgenic steroids (AAS) are included in the List of prohibited substances of the World Anti-Doping Agency (WADA) as substances abused to enhance athletic performance. Gas chromatography coupled to mass spectrometry (GC-MS) plays an important role in doping control analyses identifying AAS as their enolized-trimethylsilyl (TMS)-derivatives using the electron ionization (EI) mode. This paper explores the suitability of complementary GC-MS mass spectra and statistical analysis (principal component analysis, PCA and partial least squares-discriminant analysis, PLS-DA) to differentiate AAS as a function of their structural and conformational features expressed by their fragment ions. The results obtained showed that the application of PCA yielded a classification among the AAS molecules which became more apparent after applying PLS-DA to the dataset. The application of PLS-DA yielded a clear separation among the AAS molecules which were, thus, classified as: 1-ene-3-keto, 3-hydroxyl with saturated A-ring, 1-ene-3-hydroxyl, 4-ene-3-keto, 1,4-diene-3-keto and 3-keto with saturated A-ring anabolic steroids. The study of this paper also presents structurally diagnostic fragment ions and dissociation routes providing evidence for the presence of unknown AAS or chemically modified molecules known as designer steroids.

Experimental mixed infection of Leishmania (Leishmania) amazonensis and Leishmania (L.) infantum in hamsters (Mesocricetus auratus).

PubMed

DE Lima Celeste, Jordanna Luíza; Venuto Moura, Ana Paula; França-Silva, João Carlos; Matos DE Sousa, Gabriela; Oliveira Silva, Soraia; Norma Melo, Maria; Luiz Tafuri, Wagner; Carvalho Souza, Carolina; Monteiro DE Andrade, Hélida

2017-08-01

In South America, visceral leishmaniasis is frequently caused by Leishmania infantum and, at an unknown frequency, by Leishmania amazonensis. Therefore, mixed infections with these organisms are possible. Mixed infections might affect the clinical course, immune response, diagnosis, treatment and epidemiology of the disease. Here we describe the clinical course of mixed infections with L. amazonensis and L. infantum in a hamster model. We show that mixed infections are associated with more severe clinical disease than infection with L. amazonensis or L. infantum alone. In spleens with mixed infections, L. infantum outcompeted L. amazonensis in the tissue, but not in culture from tissue. We found increased levels of IgG in animals infected with L. infantum. Although more than 30 bands were revealed in a Western blot, the highest immunogenicity was observed with proteins having molecular masses of 95 and 90 kDa, whereas proteins with molecular masses of lower than 50 kDa were reactive frequently with serum from hamsters infected with L. amazonensis, and proteins with molecular masses of 80 and 70 kDa were reactive only with serum from hamsters infected with L. infantum. This finding has important implications regarding the biology of Leishmania and humoral immune responses to infections with these organisms.

Comparative proteomic analysis of male and female venoms from the Cuban scorpion Rhopalurus junceus.

PubMed

Rodríguez-Ravelo, Rodolfo; Batista, Cesar V F; Coronas, Fredy I V; Zamudio, Fernando Z; Hernández-Orihuela, Lorena; Espinosa-López, Georgina; Ruiz-Urquiola, Ariel; Possani, Lourival D

2015-12-01

A complete mass spectrometry analysis of venom components from male and female scorpions of the species Rhophalurus junceus of Cuba is reported. In the order of 200 individual molecular masses were identified in both venoms, from which 63 are identical in male and females genders. It means that a significant difference of venom components exists between individuals of different sexes, but the most abundant components are present in both sexes. The relative abundance of identical components is different among the genders. Three well defined groups of different peptides were separated and identified. The first group corresponds to peptides with molecular masses of 1000-2000 Da; the second to peptides with 3500-4500 Da molecular weight, and the third with 6500-8000 Da molecular weights. A total of 86 peptides rich in disulfide bridges were found in the venoms, 27 with three disulfide bridges and 59 with four disulfide bridges. LC-MS/MS analysis allowed the identification and amino acid sequence determination of 31 novel peptides in male venom. Two new putative K(+)-channel peptides were sequences by Edman degradation. They contain 37 amino acid residues, packed by three disulfide bridges and were assigned the systematic numbers: α-KTx 1.18 and α-KTx 2.15. Copyright © 2015 Elsevier Ltd. All rights reserved.

Purification and structure of human liver aspartylglucosaminidase.

PubMed Central

Rip, J W; Coulter-Mackie, M B; Rupar, C A; Gordon, B A

1992-01-01

We have recently diagnosed aspartylglucosaminuria (AGU) in four members of a Canadian family. AGU is a lysosomal storage disease in which asparagine-linked glycopeptides accumulate to particularly high concentrations in liver, spleen and thyroid of affected individuals. A lesser accumulation of these glycopeptides is seen in the kidney and brain, and they are also excreted in the urine. The altered metabolism in AGU results from a deficiency of the enzyme aspartylglucosaminidase (1-aspartamido-beta-N-acetylglucosamine amidohydrolase), which hydrolyses the asparagine to N-acetylglucosamine linkages of glycoproteins and glycopeptides. We have used human liver as a source of material for the purification of aspartylglucosaminidase. The enzyme has been purified to homogeneity by using heat treatment, (NH4)2SO4 fractionation, and chromatography on concanavalin A-Sepharose, DEAE-Sepharose, sulphopropyl-Sephadex, hydroxyapatite, DEAE-cellulose and Sephadex G-100. Enzyme activity was followed by measuring colorimetrically the N-acetylglucosamine released from aspartylglucosamine at 56 degrees C. The purified enzyme protein ran at a 'native' molecular mass of 56 kDa in SDS/12.5%-PAGE gels, and the enzyme activity could be quantitatively recovered at this molecular mass by using gel slices as enzyme source in the assay. After denaturation by boiling in SDS the 56 kDa protein was lost with the corresponding appearance of polypeptides alpha,beta and beta 1, lacking enzyme activity, at 24.6, 18.4 and 17.4 kDa respectively. Treatment of heat-denatured enzyme with N-glycosidase F resulted in the following decreases in molecular mass; 24.6 to 23 kDa and 18.4 and 17.4 to 15.8 kDa. These studies indicate that human liver aspartylglucosaminidase is composed of two non-identical polypeptides, each of which is glycosylated. The N-termini of alpha,beta and beta 1 were directly accessible for sequencing, and the first 21, 26 and 22 amino acids respectively were identified. Images Fig. 4. Fig. 7. Fig. 8. PMID:1281977

Frequency shift of a crystal quartz resonator in thickness-shear modes induced by an array of hemispherical material units.

PubMed

Yuantai Hu; Huiliang Hu; Bin Luo; Huan Xue; Jiemin Xie; Ji Wang

2013-08-01

A two-dimensional model was established to study the dynamic characteristics of a quartz crystal resonator with the upper surface covered by an array of hemispherical material units. A frequency-dependent equivalent mass ratio was proposed to simulate the effect of the covered units on frequency shift of the resonator system. It was found that the equivalent mass ratio alternately becomes positive or negative with change of shear modulus and radius of each material unit, which indicates that the equivalent mass ratio is strongly related to the vibration mode of the covered loadings. The further numerical results show the cyclical feature in the relationship of frequency shift and shear modulus/radius as expected. The solutions are useful in the analysis of frequency stability of quartz resonators and acoustic wave sensors.

Enhanced characterization of oil sands acid-extractable organics fractions using electrospray ionization-high-resolution mass spectrometry and synchronous fluorescence spectroscopy.

PubMed

Bauer, Anthony E; Frank, Richard A; Headley, John V; Peru, Kerry M; Hewitt, L Mark; Dixon, D George

2015-05-01

The open pit oil sands mining operations north of Fort McMurray, Alberta, Canada, are accumulating tailings waste at a rate approximately equal to 4.9 million m(3) /d. Naphthenic acids are among the most toxic components within tailings to aquatic life, but structural components have largely remained unidentified. In the present study, electrospray ionization high-resolution mass spectrometry (ESI-HRMS) and synchronous fluorescence spectroscopy (SFS) were used to characterize fractions derived from the distillation of an acid-extractable organics (AEO) mixture isolated from oil sands process-affected water (OSPW). Mean molecular weights of each fraction, and their relative proportions to the whole AEO extract, were as follows: fraction 1: 237 Da, 8.3%; fraction 2: 240 Da, 23.8%; fraction 3: 257 Da, 26.7%; fraction 4: 308 Da, 18.9%; fraction 5: 355 Da, 10.0%. With increasing mean molecular weight of the AEO fractions, a concurrent increase occurred in the relative abundance of nitrogen-, sulfur-, and oxygen-containing ions, double-bond equivalents, and degree of aromaticity. Structures present in the higher-molecular-weight fractions (fraction 4 and fraction 5) suggested the presence of heteroatoms, dicarboxyl and dihydroxy groups, and organic acid compounds with the potential to function as estrogens. Because organic acid compositions become dominated by more recalcitrant, higher-molecular-weight acids during natural degradation, these findings are important in the context of oil sands tailings pond water remediation. © 2015 SETAC.

The cell-wall glycoproteins of the green alga Scenedesmus obliquus. The predominant cell-wall polypeptide of Scenedesmus obliquus is related to the cell-wall glycoprotein gp3 of Chlamydomonas reinhardtii.

PubMed

Voigt, Jürgen; Stolarczyk, Adam; Zych, Maria; Malec, Przemysław; Burczyk, Jan

2014-02-01

The green alga Scenedesmus obliquus contains a multilayered cell wall, ultrastructurally similar to that of Chlamydomonas reinhardtii, although its proportion of hydroxyproline is considerably lower. Therefore, we have investigated the polypeptide composition of the insoluble and the chaotrope-soluble wall fractions of S. obliquus. The polypeptide pattern of the chaotrope-soluble wall fraction was strongly modified by chemical deglycosylation with anhydrous hydrogen fluoride (HF) in pyridine indicating that most of these polypeptides are glycosylated. Polypeptide constituents of the chaotrope-soluble cell-wall fraction with apparent molecular masses of 240, 270, 265, and 135 kDa cross-reacted with a polyclonal antibody raised against the 100 kDa deglycosylation product of the C. reinhardtii cell-wall glycoprotein GP3B. Chemical deglycosylation of the chaotrope-soluble wall fraction resulted in a 135 kDa major polypeptide and a 106 kDa minor component reacting with the same antibody. This antibody recognized specific peptide epitopes of GP3B. When the insoluble wall fraction of S. obliquus was treated with anhydrous HF/pyridine, three polypeptides with apparent molecular masses of 144, 135, and 65 kDa were solubilized, which also occured in the deglycosylated chaotrope-soluble wall fraction. These findings indicate that theses glycoproteins are cross-linked to the insoluble wall fraction via HF-sensitive bonds. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

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

Anheier, Norman C.; Cannon, Bret D.; Martinez, Alonzo

The International Atomic Energy Agency’s (IAEA’s) long-term research and development plan calls for more cost-effective and efficient safeguard methods to detect and deter misuse of gaseous centrifuge enrichment plants (GCEPs). The IAEA’s current safeguards approaches at GCEPs are based on a combination of routine and random inspections that include environmental sampling and destructive assay (DA) sample collection from UF6 in-process material and selected cylinders. Samples are then shipped offsite for subsequent laboratory analysis. In this paper, a new DA sample collection and onsite analysis approach that could help to meet challenges in transportation and chain of custody for UF6 DAmore » samples is introduced. This approach uses a handheld sampler concept and a Laser Ablation, Laser Absorbance Spectrometry (LAARS) analysis instrument, both currently under development at the Pacific Northwest National Laboratory. A LAARS analysis instrument could be temporarily or permanently deployed in the IAEA control room of the facility, in the IAEA data acquisition cabinet, for example. The handheld PNNL DA sampler design collects and stabilizes a much smaller DA sample mass compared to current sampling methods. The significantly lower uranium mass reduces the sample radioactivity and the stabilization approach diminishes the risk of uranium and hydrogen fluoride release. These attributes enable safe sample handling needed during onsite LAARS assay and may help ease shipping challenges for samples to be processed at the IAEA’s offsite laboratory. The LAARS and DA sampler implementation concepts will be described and preliminary technical viability results presented.« less

Mass Spectrometry Profiles Superoxide-Induced Intra-molecular Disulfide in the FMN-binding Subunit of Mitochondrial Complex I

PubMed Central

Zhang, Liwen; Xu, Hua; Chen, Chwen-Lih; Green-Church, Kari B.; Freitas, Michael A.; Chen, Yeong-Renn

2008-01-01

Protein thiols with regulatory functions play a critical role in maintaining the homeostasis of the redox state in mitochondria. One major host of regulatory cysteines in mitochondria is complex I, with the thiols primarily located on its 51 kDa FMN-binding subunit. In response to oxidative stress, these thiols are expected to form intra-molecular disulfide bridges as one of their oxidative post-translational modifications. Here, to test this hypothesis and gain insights into the molecular pattern of disulfide in complex I, the isolated bovine complex I was prepared. Superoxide (O2•−) is generated by complex I under the conditions of enzyme turnover. O2•−-induced intra-molecular disulfide formation at the 51 kDa subunit was determined by tandem mass spectrometry and database searching, with the latter accomplished by adaptation of the in-house developed database search engine, MassMatrix [Xu H., et. al J. Proteome Res. (2008) 7, 138–44]. LC/MS/MS analysis of tryptic/chymotryptic digests of the 51 kDa subunit from alkylated complex I revealed that four specific cysteines (C125, C142, C187, and C206) of the 51 kDa subunit were involved in the formation of mixed intra-molecular disulfide linkages. In all, three cysteine pairs were observed: C125/C142, C187/C206, and C142/C206. The formation of disulfide bond was subsequently inhibited by superoxide dismutase, indicating the involvement of O2•−. These results elucidated by mass spectrometry indicates that the residues of C125, C142, C187, and C206 are the specific regulatory cysteines of complex I, and they participate in the oxidative modification with disulfide formation under the physiological or pathophysiological conditions of oxidative stress. PMID:18789718

Nucleus accumbens neurotransmission and effort-related choice behavior in food motivation: effects of drugs acting on dopamine, adenosine, and muscarinic acetylcholine receptors.

PubMed

Nunes, Eric J; Randall, Patrick A; Podurgiel, Samantha; Correa, Mercè; Salamone, John D

2013-11-01

Mesolimbic dopamine (DA) is a critical component of the brain circuitry regulating behavioral activation and effort-related processes. Although nucleus accumbens (NAc) DA depletions or antagonism leave aspects of appetite and primary food motivation intact, rats with impaired DA transmission reallocate their instrumental behavior away from food-reinforced tasks with high response requirements, and instead select less effortful food-seeking behaviors. Previous work showed that adenosine A2A antagonists can reverse the effects of DA D2 antagonists on effort-related choice, and that stimulation of adenosine A2A receptors produces behavioral effects that are similar to those induced by DA antagonism. The present review summarizes the literature on the role of NAc DA and adenosine in effort-related processes, and also presents original data on the effects of local stimulation of muscarinic acetylcholine receptors in NAc core. Local injections of the muscarinic agonist pilocarpine directly into NAc core produces shifts in effort-related choice behavior similar to those induced by DA antagonism or A2A receptor stimulation, decreasing lever pressing but increasing chow intake in rats responding on a concurrent fixed ratio/chow feeding choice task. In contrast, injections into a neostriatal control site dorsal to the NAc were ineffective. The actions of pilocarpine on this task were attenuated by co-administration of the muscarinic antagonist scopolamine. Thus, drugs that act on DA, adenosine A2A, and muscarinic receptors regulate effort-related choice behavior, which may have implications for the treatment of psychiatric symptoms such as psychomotor slowing, fatigue or anergia that can be observed in depression and other disorders. Copyright © 2013 Elsevier Ltd. All rights reserved.

Purification of acetaldehyde dehydrogenase and alcohol dehydrogenases from Thermoanaerobacter ethanolicus 39E and characterization of the secondary-alcohol dehydrogenase (2 degrees Adh) as a bifunctional alcohol dehydrogenase--acetyl-CoA reductive thioesterase.

PubMed Central

Burdette, D; Zeikus, J G

1994-01-01

The purification and characterization of three enzymes involved in ethanol formation from acetyl-CoA in Thermoanaerobacter ethanolicus 39E (formerly Clostridium thermohydrosulfuricum 39E) is described. The secondary-alcohol dehydrogenase (2 degrees Adh) was determined to be a homotetramer of 40 kDa subunits (SDS/PAGE) with a molecular mass of 160 kDa. The 2 degrees Adh had a lower catalytic efficiency for the oxidation of 1 degree alcohols, including ethanol, than for the oxidation of secondary (2 degrees) alcohols or the reduction of ketones or aldehydes. This enzyme possesses a significant acetyl-CoA reductive thioesterase activity as determined by NADPH oxidation, thiol formation and ethanol production. The primary-alcohol dehydrogenase (1 degree Adh) was determined to be a homotetramer of 41.5 kDa (SDS/PAGE) subunits with a molecular mass of 170 kDa. The 1 degree Adh used both NAD(H) and NADP(H) and displayed higher catalytic efficiencies for NADP(+)-dependent ethanol oxidation and NADH-dependent acetaldehyde (identical to ethanal) reduction than for NADPH-dependent acetaldehyde reduction or NAD(+)-dependent ethanol oxidation. The NAD(H)-linked acetaldehyde dehydrogenase was a homotetramer (360 kDa) of identical subunits (100 kDa) that readily catalysed thioester cleavage and condensation. The 1 degree Adh was expressed at 5-20% of the level of the 2 degrees Adh throughout the growth cycle on glucose. The results suggest that the 2 degrees Adh primarily functions in ethanol production from acetyl-CoA and acetaldehyde, whereas the 1 degree Adh functions in ethanol consumption for nicotinamide-cofactor recycling. Images Figure 1 PMID:8068002

Relations between Mass Change and Frequency Shift of a QCM Sensor in Contact with Viscoelastic Medium

NASA Astrophysics Data System (ADS)

Tan, Feng; Huang, Xian-He

2013-05-01

We investigate frequency shift of a quartz crystal microbalance (QCM) sensor introduced by mass effect, and properties of material of its coated viscoelastic film are discussed. The validity of the Sauerbrey relation cannot be held if the viscoelastic properties of the contacting medium are considered. When the QCM sensor with a viscoelastic film works in the gas phase, the viscoelastic properties will introduce an extra mass effect. While in the liquid phase, the missing mass effect can be observed. The experimental results demonstrate that the QCM sensor is sensitive to the viscoelastic properties of the coating film. Properties of the viscoelastic contacting medium should be considered.

Innovation Policies of Brazil

DTIC Science & Technology

2013-09-01

technology priorities and coordinates policies Ministry of Science, Technology and Innovation (Ministério da Ciência, Tecnologia e Inovação, MCTI... Tecnologia e Inovação, MCTI) initiated the Greater IT policy to build and enhance the country’s information, communications, and technology...Technology Institute) MCMM Ministry of Communications and Mass Media MCTI Ministério da Ciência, Tecnologia e Inovação (Ministry of Science, Technology

A dominant sulfhydryl-containing protein in the outer membrane of Neisseria gonorrhoeae.

PubMed Central

Norrod, E P; Browne, S L; Feldweg, A; Leonard, J

1993-01-01

By using a method that labels sulfhydryl-containing proteins in situ, we have detected a major outer membrane protein of Neisseria gonorrhoeae at 41 kDa. A protein of this molecular mass has not previously been shown to be a major outer membrane protein in gonococci. In addition, a minor protein rich in cysteinyl residues was detected at 31.5 kDa. Images PMID:8432710

Tropomyosin and Actin Identified as Major Allergens of the Carpet Clam (Paphia textile) and the Effect of Cooking on Their Allergenicity

PubMed Central

Mohamad Yadzir, Zailatul Hani; Misnan, Rosmilah; Bakhtiar, Faizal; Abdullah, Noormalin; Murad, Shahnaz

2015-01-01

Objectives. To identify the major allergenic proteins of clam (Paphia textile) and to investigate the effect of different cooking methods on the allergenicity of these identified proteins. Methods. Clam protein extracts were separated by denaturing polyacrylamide gel electrophoresis. IgE reactive proteins were then analyzed by immunoblotting with sera from patients with positive skin prick tests (SPT) to the raw clam extract. Mass spectrometry was used to identify the major allergenic proteins of this clam. Results. Raw extract showed 12 protein bands (18–150 kDa). In contrast, fewer protein bands were seen in the boiled extract; those ranging from 40 to 150 kDa were denatured. The protein profiles were similarly altered by frying or roasting. The immunoblots of raw and boiled extracts yielded 10 and 2 IgE-binding proteins, respectively. The fried and roasted extracts showed only a single IgE-binding protein at 37 kDa. Mass spectrometry analysis of the 37 and 42 kDa major allergens indicated that these spots were tropomyosin and actin, respectively. Conclusion. The two major allergens of Paphia textile were identified as the thermostable tropomyosin and a new thermolabile allergen actin. PMID:26413512

A comparison of an ATPase from the archaebacterium Halobacterium saccharovorum with the F1 moiety from the Escherichia coli ATP Synthase

NASA Technical Reports Server (NTRS)

Stan-Lotter, Helga; Hochstein, Lawrence I.

1989-01-01

A purified ATPase associated with membranes from Halobacterium saccharovorum was compared with the F sub 1 moiety from the Escherichia coli ATP Synthase. The halobacterial enzyme was composed of two major (I and II) and two minor subunits (III and IV), whose molecular masses were 87 kDa, 60 kDa, 29 kDa, and 20 kDa, respectively. The isoelectric points of these subunits ranged from 4.1 to 4.8, which in the case of the subunits I and II was consistent with the presence of an excess of acidic amino acids (20 to 22 Mol percent). Peptide mapping of sodium dodecylsulfate-denatured subunits I and II showed no relationship between the primary structures of the individual halobacterial subunits or similarities to the subunits of the F sub 1 ATPase (EC 3.6.1.34) from E. coli. Trypsin inactivation of the halobacterial ATPase was accompanied by the partial degradation of the major subunits. This observation, taken in conjunction with molecular masses of the subunits and the native enzyme, was consistent with the previously proposed stoichiometry of 2:2:1:1. These results suggest that H. saccharovorum, and possibly, Halobacteria in general, possess an ATPase which is unlike the ubiquitous F sub o F sub 1 - ATP Synthase.

Biochemical characterization of the 49 kDa penicillin-binding protein of Mycobacterium smegmatis.

PubMed Central

Mukherjee, T; Basu, D; Mahapatra, S; Goffin, C; van Beeumen, J; Basu, J

1996-01-01

The 49 kDa penicillin-binding protein (PBP) of Mycobacterium smegmatis catalyses the hydrolysis of the peptide or S-ester bond of carbonyl donors R1-CONH-CHR2-COX-CHR2-COO- (where X is NH or S). In the presence of a suitable amino acceptor, the reaction partitions between the transpeptidation and hydrolysis pathways, with the amino acceptor, behaving as a simple alternative nucleophile at the level of the acyl-enzyme. By virtue of its N-terminal sequence similarity, the 49 kDa PBP represents one of the class of monofunctional low-molecular-mass PBPs. An immunologically related protein of M(r) 52,000 is present in M. tuberculosis. The 49 kDa PBP is sensitive towards amoxycillin, imipenem, flomoxef and cefoxitin. PMID:8947487

Reversibility between glass and melting transitions of poly(oxyethylene)

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

Qui, Wulin; Pyda, Marek; Nowak-Pyda, Elisabieta

2005-01-01

The heat capacities, C{sub p}, of poly(oxyethylene), POE, with molar masses from 1500 to 900,000 Da, were analyzed by differential scanning calorimetry (DSC), quasi-isothermal, temperature-modulated DSC (TMDSC), and wide-angle X-ray diffraction (WAXD). There is no change in crystal structure before melting, but the lattice parameters increase rapidly in the melting region. Perfected extended-chain and once- or twice-folded crystals of the oligomers with a molar mass above 1100 Da melt practically fully irreversibly and permit direct measurement of the thermodynamic C{sub p}. The folded-chain crystals of high molar mass show some locally reversible melting. The reversing, apparent C{sub p} depends onmore » molar mass and amplitude and frequency of modulation. After separation from the latent heat effects, the reversible, thermodynamic C{sub p} depends on the melting temperature for low molar masses and increases beyond the vibrational C{sub p} due to conformational motion. Molar masses of 8000-20,000 have almost the same C{sub p}. These observations permit a quantitative discussion of the thermodynamic C{sub p} and the locally reversible melting of the globally metastable POE in the melting range. The increase in C{sub p} between 250 K and the melting temperature is interpreted as a glass transition within the crystal.« less

PARABIOTIC INTOXICATION. II. THE DISTRIBUTION AND SURVIVAL OF Cr$sup 51$- LABELED RED BLOOD CELLS

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

Tokuda, S.; MacGillivray, M.H.

1962-04-01

The anemia and polycythemia in parent-to-F/sub 1/ parabiotic intoxication in mice were studied using Cr/sup 51/- and Fe/sup 59/ labeled red blood cells. It was observed that the anemia and polycythemia result from a shift in red cell mass from the hybrid into its parent strain partner. Because crosscirculation was observed between the parabionts during the anemia and polycythemia, the shift is attributed to unequal cross transfusion between the parabionts. There was neither preferential selection nor preferential destruction of either parental or hybrid red cells during the shift in red cell mass. (auth)

Mechanical and statistical evidence of the causality of human-made mass shifts on the Earth's upper crust and the occurrence of earthquakes

NASA Astrophysics Data System (ADS)

Klose, Christian D.

2013-01-01

A global catalog of small- to large-sized earthquakes was systematically analyzed to identify causality and correlatives between human-made mass shifts in the upper Earth's crust and the occurrence of earthquakes. The mass shifts, ranging between 1 kt and 1 Tt, result from large-scale geoengineering operations, including mining, water reservoirs, hydrocarbon production, fluid injection/extractions, deep geothermal energy production and coastal management. This article shows evidence that geomechanical relationships exist with statistical significance between (a) seismic moment magnitudes M of observed earthquakes, (b) lateral distances of the earthquake hypocenters to the geoengineering "operation points" and (c) mass removals or accumulations on the Earth's crust. Statistical findings depend on uncertainties, in particular, of source parameter estimations of seismic events before instrumental recoding. Statistical observations, however, indicate that every second, seismic event tends to occur after a decade. The chance of an earthquake to nucleate after 2 or 20 years near an area with a significant mass shift is 25 or 75 %, respectively. Moreover, causative effects of seismic activities highly depend on the tectonic stress regime in which the operations take place (i.e., extensive, transverse or compressive). Results are summarized as follows: First, seismic moment magnitudes increase the more mass is locally shifted on the Earth's crust. Second, seismic moment magnitudes increase the larger the area in the crust is geomechanically polluted. Third, reverse faults tend to be more trigger-sensitive than normal faults due to a stronger alteration of the minimum vertical principal stress component. Pure strike-slip faults seem to rupture randomly and independently from the magnitude of the mass changes. Finally, mainly due to high estimation uncertainties of source parameters and, in particular, of shallow seismic events (<10 km), it remains still very difficult to discriminate between induced and triggered earthquakes with respect to the data catalog of this study. However, first analyses indicate that small- to medium-sized earthquakes (M6) seem to be triggered. The rupture propagation of triggered events might be dominated by pre-existing tectonic stress conditions.

The inverse problem of sensing the mass and force induced by an adsorbate on a beam nanomechanical resonator

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

Liu, Yun; Zhang, Yin

2016-06-08

The mass sensing superiority of a micro/nanomechanical resonator sensor over conventional mass spectrometry has been, or at least, is being firmly established. Because the sensing mechanism of a mechanical resonator sensor is the shifts of resonant frequencies, how to link the shifts of resonant frequencies with the material properties of an analyte formulates an inverse problem. Besides the analyte/adsorbate mass, many other factors such as position and axial force can also cause the shifts of resonant frequencies. The in-situ measurement of the adsorbate position and axial force is extremely difficult if not impossible, especially when an adsorbate is as smallmore » as a molecule or an atom. Extra instruments are also required. In this study, an inverse problem of using three resonant frequencies to determine the mass, position and axial force is formulated and solved. The accuracy of the inverse problem solving method is demonstrated and how the method can be used in the real application of a nanomechanical resonator is also discussed. Solving the inverse problem is helpful to the development and application of mechanical resonator sensor on two things: reducing extra experimental equipments and achieving better mass sensing by considering more factors.« less

Comparison of different source calculations in two-nucleon channel at large quark mass

NASA Astrophysics Data System (ADS)

Yamazaki, Takeshi; Ishikawa, Ken-ichi; Kuramashi, Yoshinobu

2018-03-01

We investigate a systematic error coming from higher excited state contributions in the energy shift of light nucleus in the two-nucleon channel by comparing two different source calculations with the exponential and wall sources. Since it is hard to obtain a clear signal of the wall source correlation function in a plateau region, we employ a large quark mass as the pion mass is 0.8 GeV in quenched QCD. We discuss the systematic error in the spin-triplet channel of the two-nucleon system, and the volume dependence of the energy shift.

Diffusive transfer to membranes as an effective interface between gel electrophoresis and mass spectrometry

NASA Astrophysics Data System (ADS)

Ogorzalek Loo, Rachel R.; Mitchell, Charles; Stevenson, Tracy I.; Loo, Joseph A.; Andrews, Philip C.

1997-12-01

Diffusive transfer was examined as a blotting method to transfer proteins from polyacrylamide gels to membranes for ultraviolet matrix-assisted laser desorption ionization (MALDI) mass spectrometry. The method is well-suited for transfers from isoelectric focusing (IEF) gels. Spectra have been obtained for 11 pmol of 66 kDa albumin loaded onto an IEF gel and subsequently blotted to polyethylene. Similarly, masses of intact carbonic anhydrase and hemoglobin were obtained from 14 and 20 pmol loadings. This methodology is also compatible with blotting high molecular weight proteins, as seen for 6 pmol of the 150 kDa monoclonal antibody anti-[beta]-galactosidase transferred to Goretex. Polypropylene, Teflon, Nafion and polyvinylidene difluoride (PVDF) also produced good spectra following diffusive transfer. Only analysis from PVDF required that the membrane be kept wet prior to application of matrix. Considerations in mass accuracy for analysis from large-area membranes with continuous extraction and delayed extraction were explored, as were remedies for surface charging. Vapor phase CNBr cleavage was applied to membrane-bound samples for peptide mapping.

2MASS Photometry of the Hot DA White Dwarf Stars in the Palomar Green Survey

NASA Astrophysics Data System (ADS)

Holberg, J. B.; Magargal, K.

2003-12-01

The Palomar Green (PG) Survey is a complete, magnitude limited survey of UV excess objects that continues to provide well-defined sample populations for many types of objects, in particular hot white dwarf stars. The 2MASS All-Sky Survey limiting JHK magnitudes are reasonably well matched to the B magnitude limits of the PG survey. The 2MASS survey, therefore, constitutes an excellent source of uniform, high-quality of photometry, that can be used in conjunction with the PG Survey. The 2MASS Point Source Catalog in the All-Sky Data Release was searched for over 340 hot DA white dwarfs in the PG sample. The resulting JHK colors and apparent magnitudes are used to determine photometric distances for these stars and to place limits on the existence of possible cool binary companions. This publication makes use of data products from the Two Micron All Sky Survey, which is a joint project of the University of Massachusetts and the Infrared Processing and Analysis Center/California Institute of Technology, funded by the National Aeronautics and Space Administration and the National Science Foundation.

Discrimination Between Peptide O-Sulfo- and O-Phosphotyrosine Residues by Negative Ion Mode Electrospray Tandem Mass Spectrometry

NASA Astrophysics Data System (ADS)

Edelson-Averbukh, Marina; Shevchenko, Andrej; Pipkorn, Rüdiger; Lehmann, Wolf D.

2011-12-01

Unambiguous differentiation between isobaric sulfated and phosphorylated tyrosine residues (sTyr and pTyr) of proteins by mass spectrometry is challenging, even using high resolution mass spectrometers. Here we show that upon negative ion mode collision-induced dissociation (CID), pTyr- and sTyr-containing peptides exhibit entirely different modification-specific fragmentation patterns leading to a rapid discrimination between the isobaric covalent modifications using the tandem mass spectral data. This study reveals that the ratio between the relative abundances of [M-H-80]- and [M-H-98]- fragment ions in ion-trap CID and higher energy collision dissociation (HCD) spectra of singly deprotonated +80 Da Tyr-peptides can be used as a reliable indication of the Tyr modification group nature. For multiply deprotonated +80 Da Tyr-peptides, CID spectra of sTyr- and pTyr-containing sequences can be readily distinguished based on the presence/absence of the [M-nH-79](n-1)- and [M-nH-79-NL]( n-1)- ( n = 2, 3) fragment ions (NL = neutral loss).

Lipids and proteins in the Rathke's gland secretions of the North American mud turtle (Kinosternon subrubrum)

USGS Publications Warehouse

Seifert, W.E.; Gotte, S.W.; Leto, T.L.; Weldon, P.J.

1994-01-01

Lipids and proteins in the Rathke's gland secretions of the North American mud turtle (Kinosternon subrubrum, Kinosternidae) were analyzed by gas chromatography-mass spectrometry (GC-MS) and SDS-polyacrylamide gel electrophoresis (SDS-PAGE), respectively. Analysis by GC-MS indicates 2,3-dihydroxypropanal and C3–C24 free or esterified fatty acids. Analysis by SDS-PAGE indicates a major protein component with an approximate molecular mass of 60 kDa and minor components ranging from ca. 23 to 34 kDa. The major component of K. subrubrum glandular secretions exhibits a mobility that matches that of the Kemp's ridley sea turtle (Lepidochelys kempi, Cheloniidae), suggesting that these proteins are evolutionarily conserved.

Surface-Induced Dissociation of Protein Complexes in a Hybrid Fourier Transform Ion Cyclotron Resonance Mass Spectrometer

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

Yan, Jing; Zhou, Mowei; Gilbert, Joshua D.

Mass spectrometry continues to develop as a valuable tool in the analysis of proteins and protein complexes. In protein complex mass spectrometry studies, surface-induced dissociation (SID) has been successfully applied in quadrupole time-of-flight (Q-TOF) instruments. SID provides structural information on non-covalent protein complexes that is complementary to other techniques. However, the mass resolution of Q-TOF instruments can limit the information that can be obtained for protein complexes by SID. Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) provides ultrahigh resolution and ultrahigh mass accuracy measurements. In this study, an SID device was designed and successfully installed in a hybridmore » FT-ICR instrument in place of the standard gas collision cell. The SID-FT-ICR platform has been tested with several protein complex systems (homooligomers, a heterooligomer, and a protein-ligand complex, ranging from 53 kDa to 85 kDa), and the results are consistent with data previously acquired on Q-TOF platforms, matching predictions from known protein interface information. SID fragments with the same m/z but different charge states are well-resolved based on distinct spacing between adjacent isotope peaks, and the addition of metal cations and ligands can also be isotopically resolved with the ultrahigh mass resolution available in FT-ICR.« less

Proteolytic processing and activation of Clostridium perfringens epsilon toxin by caprine small intestinal contents.

PubMed

Freedman, John C; Li, Jihong; Uzal, Francisco A; McClane, Bruce A

2014-10-21

Epsilon toxin (ETX), a pore-forming toxin produced by type B and D strains of Clostridium perfringens, mediates severe enterotoxemia in livestock and possibly plays a role in human disease. During enterotoxemia, the nearly inactive ETX prototoxin is produced in the intestines but then must be activated by proteolytic processing. The current study sought to examine ETX prototoxin processing and activation ex vivo using the intestinal contents of a goat, a natural host species for ETX-mediated disease. First, this study showed that the prototoxin has a KEIS N-terminal sequence with a molecular mass of 33,054 Da. When the activation of ETX prototoxin ex vivo by goat small intestinal contents was assessed by SDS-PAGE, the prototoxin was processed in a stepwise fashion into an ~27-kDa band or higher-molecular-mass material that could be toxin oligomers. Purified ETX corresponding to the ~27-kDa band was cytotoxic. When it was biochemically characterized by mass spectrometry, the copresence of three ETX species, each with different C-terminal residues, was identified in the purified ~27-kDa ETX preparation. Cytotoxicity of each of the three ETX species was then demonstrated using recombinant DNA approaches. Serine protease inhibitors blocked the initial proteotoxin processing, while carboxypeptidase inhibitors blocked further processing events. Taken together, this study provides important new insights indicating that, in the intestinal lumen, serine protease (including trypsin and possibly chymotrypsin) initiates the processing of the prototoxin but other proteases, including carboxypeptidases, then process the prototoxin into multiple active and stable species. Importance: Processing and activation by intestinal proteases is a prerequisite for ETX-induced toxicity. Previous studies had characterized the activation of ETX using only arbitrarily chosen amounts of purified trypsin and/or chymotrypsin. Therefore, the current study examined ETX activation ex vivo by natural host intestinal contents. These analyses demonstrated that (i) ETX processing in host intestinal contents occurs in an ordered, stepwise fashion, (ii) processing of prototoxin by host intestinal contents results in higher-molecular-mass material and 3 distinct ~27-kDa ETX species, and (iii) serine proteases, such as trypsin, chymotrypsin, and other proteases, including carboxypeptidases, play a role in the activation of ETX by intestinal contents. These studies provide new insights into the activation and processing of ETX and demonstrate that this process is more complicated than previously appreciated. Copyright © 2014 Freedman et al.

Flexible Stoichiometry and Asymmetry of the PIDDosome Core Complex by Heteronuclear NMR Spectroscopy and Mass Spectrometry

PubMed Central

Nematollahi, Lily A.; Garza-Garcia, Acely; Bechara, Chérine; Esposito, Diego; Morgner, Nina; Robinson, Carol V.; Driscoll, Paul C.

2015-01-01

Homotypic death domain (DD)–DD interactions are important in the assembly of oligomeric signaling complexes such as the PIDDosome that acts as a platform for activation of caspase-2-dependent apoptotic signaling. The structure of the PIDDosome core complex exhibits an asymmetric three-layered arrangement containing five PIDD-DDs in one layer, five RAIDD-DDs in a second layer and an additional two RAIDD-DDs. We addressed complex formation between PIDD-DD and RAIDD-DD in solution using heteronuclear nuclear magnetic resonance (NMR) spectroscopy, nanoflow electrospray ionization mass spectrometry and size-exclusion chromatography with multi-angle light scattering. The DDs assemble into complexes displaying molecular masses in the range 130–158 kDa and RAIDD-DD:PIDD-DD stoichiometries of 5:5, 6:5 and 7:5. These data suggest that the crystal structure is representative of only the heaviest species in solution and that two RAIDD-DDs are loosely attached to the 5:5 core. Two-dimensional 1H,15N-NMR experiments exhibited signal loss upon complexation consistent with the formation of high-molecular-weight species. 13C-Methyl-transverse relaxation optimized spectroscopy measurements of the PIDDosome core exhibit signs of differential line broadening, cross-peak splitting and chemical shift heterogeneity that reflect the presence of non-equivalent sites at interfaces within an asymmetric complex. Experiments using a mutant RAIDD-DD that forms a monodisperse 5:5 complex with PIDD-DD show that the spectroscopic signature derives from the quasi- but non-exact equivalent environments of each DD. Since this characteristic was previously demonstrated for the complex between the DDs of CD95 and FADD, the NMR data for this system are consistent with the formation of a structure homologous to the PIDDosome core. PMID:25528640

Profiling and relative quantification of phosphatidylethanolamine based on acetone stable isotope derivatization.

PubMed

Wang, Xiang; Wei, Fang; Xu, Ji-Qu; Lv, Xin; Dong, Xu-Yan; Han, Xianlin; Quek, Siew-Young; Huang, Feng-Hong; Chen, Hong

2016-01-01

Phosphatidylethanolamine (PE) is considered to be one of the pivotal lipids for normal cellular function as well as disease initiation and progression. In this study, a simple, efficient, reliable, and inexpensive method for the qualitative analysis and relative quantification of PE, based on acetone stable isotope derivatization combined with double neutral loss scan-shotgun electrospray ionization tandem-quadrupole mass spectrometry analysis (ASID-DNLS-Shotgun ESI-MS/MS), was developed. The ASID method led to alkylation of the primary amino groups of PE with an isopropyl moiety. The use of acetone (d0-acetone) and deuterium-labeled acetone (d6-acetone) introduced a 6 Da mass shift that was ideally suited for relative quantitative analysis, and enhanced sensitivity for mass analysis. The DNLS model was introduced to simultaneously analyze the differential derivatized PEs by shotgun ESI-MS/MS with high selectivity and accuracy. The reaction specificity, labeling efficiency, and linearity of the ASID method were thoroughly evaluated in this study. Its excellent applicability was validated by qualitative and relative quantitative analysis of PE species presented in liver samples from rats fed different diets. Using the ASID-DNLS-Shotgun ESI-MS/MS method, 45 PE species from rat livers have been identified and quantified in an efficient manner. The level of total PEs tended to decrease in the livers of rats on high fat diets compared with controls. The levels of PE 32:1, 34:3, 34:2, 36:3, 36:2, 42:10, plasmalogen PE 36:1 and lyso PE 22:6 were significantly reduced, while levels of PE 36:1 and lyso PE 16:0 increased. Copyright © 2015 Elsevier B.V. All rights reserved.

Purification and partial characterization of peroxidase from human term placenta of non-smokers: metabolism of benzo(a)pyrene-7, 8-dihydrodiol.

PubMed

Madhavan, N D; Naidu, K A

2000-01-01

Peroxidase (Donor: H(2)O(2)oxidoreductase EC 1.11.1.7) from human term placentae of non-smokers was purified to homogeneity by a combination of NH(4)Cl extraction, affinity chromatography, (NH(4))(2)SO(4)precipitation, ion-exchange and gel filtration chromatography. The homogeneity of purified human placental peroxidase (HTPP) was confirmed by gel filtration, reverse phase high performance liquid chromatography (HPLC) and SDS-PAGE. Peroxidase was found to be a membrane bound enzyme. A high concentration of NH(4)Cl (1.2 m) was needed to extract and solublize the enzyme. Removal of the salt resulted in irreversible precipitation of the enzyme. The protein exhibited a molecular mass of 126 000 kDa according to gel filtration and approximately 60 000 kDa by SDS-PAGE, indicating that the peroxidase is a homodimer. The purified peroxidase showed an optimum pH range of 7 to 8.5 and the K(m)for H(2)O(2)and guaiacol were found to be 0.08 m m and 10.0 m m, respectively. The purified peroxidase oxidized several substrates, namely potassium iodide, tetramethyl benzidine, guaiacol, ortho dianisidne and tyrosine. The enzyme was resistant to thermal denaturation up to 70 degrees C and also to chaotropic agents, guanidinium chloride and urea. Spectral properties indicated the presence of Soret band at 433 which shifted to 451 nm on complexation with cyanide. The circular dichroism studies showed that HTPP has a predominantly helical secondary structure. The enzyme showed similarities to the myeloperoxidase with regard to spectral and catalytical properties but differed significantly in amino acid composition, the R(z)value and molecular mass. Purified HTPP differed from eosinophil peroxidase in all physico-chemical properties indicating that it is not of eosinophil origin, but may represent a distinct, constitutive peroxidase in human placenta. Further, purified peroxidase catalyzed oxidation of benzo(a)pyrene-7, 8-dihydrodiol in presence of tyrosine and hydrogen peroxide to BP-tetrols, the hydrolytic products of BP-diol-epoxides, demonstrating the ability of peroxidase in bioactivation of benzo(a)pyrene in human placenta. Copyright 2000 Harcourt Publishers Ltd.

The 60 kDa heat shock proteins in the hyperthermophilic archaeon Sulfolobus shibatae.

PubMed

Kagawa, H K; Osipiuk, J; Maltsev, N; Overbeek, R; Quaite-Randall, E; Joachimiak, A; Trent, J D

1995-11-10

One of the most abundant proteins in the hyperthermophilic archaeon Sulfolobus shibatae is the 59 kDa heat shock protein (TF55) that is believed to form a homo-oligomeric double ring complex structurally similar to the bacterial chaperonins. We discovered a second protein subunit in the S. shibatae ring complex (referred to as alpha) that is stoichiometric with TF55 (renamed beta). The gene and flanking regions of alpha were cloned and sequenced and its inferred amino acid sequence has 54.4% identity and 74.4% similarity to beta. Transcription start sites for both alpha and beta were mapped and three potential transcription regulatory regions were identified. Northern analyses of cultures shifted from normal growth temperatures (70 to 75 degrees C) to heat shock temperatures (85 to 90 degrees C) indicated that the levels of alpha and beta mRNAs increased during heat shock, but at all temperatures their relative proportions remained constant. Monitoring protein synthesis by autoradiography of total proteins from cultures pulse labeled with L(-)[35S]methionine at normal and heat shock temperatures indicated significant increases in alpha and beta synthesis during heat shock. Under extreme heat shock conditions (> or = 90 degrees C) alpha and beta appeared to be the only two proteins synthesized. The purified alpha and beta subunits combined to form high molecular mass complexes with similar mobilities on native polyacrylamide gels to the complexes isolated directly from cells. Equal proportions of the two subunits gave the greatest yield of the complex, which we refer to as a "rosettasome". It is argued that the rosettasome consists of two homo-oligomeric rings; one of alpha and the other of beta. Polyclonal antibodies against alpha and beta from S. shibatae cross-reacted with proteins of similar molecular mass in 10 out of the 17 archaeal species tested, suggesting that the two rosettasome proteins are highly conserved among the archaea. The archaeal sequences were aligned with bacterial and eukaryotic chaperonins to generate a phylogenetic tree. The tree reveals the close relationship between the archaeal rosettasomes and the eukaryotic TCP1 protein family and the distant relationship to the bacterial GroEL/HSP60 proteins.

[Amanitine determination as an example of peptide analysis in the biological samples with HPLC-MS technique].

PubMed

Janus, Tomasz; Jasionowicz, Ewa; Potocka-Banaś, Barbara; Borowiak, Krzysztof

Routine toxicological analysis is mostly focused on the identification of non-organic and organic, chemically different compounds, but generally with low mass, usually not greater than 500–600 Da. Peptide compounds with atomic mass higher than 900 Da are a specific analytical group. Several dozen of them are highly-toxic substances well known in toxicological practice, for example mushroom toxin and animal venoms. In the paper the authors present an example of alpha-amanitin to explain the analytical problems and different original solutions in identifying peptides in urine samples with the use of the universal LC MS/MS procedure. The analyzed material was urine samples collected from patients with potential mushroom intoxication, routinely diagnosed for amanitin determination. Ultra filtration with centrifuge filter tubes (limited mass cutoff 3 kDa) was used. Filtrate fluid was directly injected on the chromatographic column and analyzed with a mass detector (MS/MS). The separation of peptides as organic, amphoteric compounds from biological material with the use of the SPE technique is well known but requires dedicated, specific columns. The presented paper proved that with the fast and simple ultra filtration technique amanitin can be effectively isolated from urine, and the procedure offers satisfactory sensitivity of detection and eliminates the influence of the biological matrix on analytical results. Another problem which had to be solved was the non-characteristic fragmentation of peptides in the MS/MS procedure providing non-selective chromatograms. It is possible to use higher collision energies in the analytical procedure, which results in more characteristic mass spectres, although it offers lower sensitivity. The ultra filtration technique as a procedure of sample preparation is effective for the isolation of amanitin from the biological matrix. The monitoring of selected mass corresponding to transition with the loss of water molecule offers satisfactory sensitivity of determination.

Casein kinase II protein kinase is bound to lamina-matrix and phosphorylates lamin-like protein in isolated pea nuclei

NASA Technical Reports Server (NTRS)

Li, H.; Roux, S. J.

1992-01-01

A casein kinase II (CK II)-like protein kinase was identified and partially isolated from a purified envelope-matrix fraction of pea (Pisum sativum L.) nuclei. When [gamma-32P]ATP was directly added to the envelope-matrix preparation, the three most heavily labeled protein bands had molecular masses near 71, 48, and 46 kDa. Protein kinases were removed from the preparation by sequential extraction with Triton X-100, EGTA, 0.3 M NaCl, and a pH 10.5 buffer, but an active kinase still remained bound to the remaining lamina-matrix fraction after these treatments. This kinase had properties resembling CK II kinases previously characterized from animal and plant sources: it preferred casein as an artificial substrate, could use GTP as efficiently as ATP as the phosphoryl donor, was stimulated by spermine, was calcium independent, and had a catalytic subunit of 36 kDa. Some animal and plant CK II kinases have regulatory subunits near 29 kDa, and a lamina-matrix-bound protein of this molecular mass was recognized on immunoblot by anti-Drosophila CK II polyclonal antibodies. Also found associated with the envelope-matrix fraction of pea nuclei were p34cdc2-like and Ca(2+)-dependent protein kinases, but their properties could not account for the protein kinase activity bound to the lamina. The 71-kDa substrate of the CK II-like kinase was lamin A-like, both in its molecular mass and in its cross-reactivity with anti-intermediate filament antibodies. Lamin phosphorylation is considered a crucial early step in the entry of cells into mitosis, so lamina-bound CK II kinases may be important control points for cellular proliferation.

Identification of pheromone-carrying protein in the preorbital gland post in the endangered Indian male Blackbuck Antelope cervicapra L.

PubMed

Rajagopal, T; Rajkumar, R; Ponmanickam, P; Achiraman, S; Padmanabhan, P; Archunan, G

2015-12-01

In mammals, a low molecular mass protein (17-20 KDa) reported from the pheromone sources such as urine, saliva, glandular secretion, etc., as ligand-carrier (pheromone carrier) has been associated with chemo-communication. Since the preorbital gland post is one of the major pheromone sources in Indian Blackbuck, an endangered species, we assumed that it possibly contains low molecular mass protein for chemical communication. Hence, we investigated the preorbital gland post in territorial and non-territorial male blackbucks for such low molecular mass proteins adopting SDS-PAGE and LC-MS/MS analysis. The total content of protein was higher in the post of territorial males than non-territorial males of adult and sub-adult. In fact, the protein profiles such as 17, 21, 25, 42 and 61 kDa were noted in the gland secretion of territorial and non-territorial males. The intensity of the 17 kDa protein band was higher in territorial males than non-territorial males. In-gel trypsin digestion of the 17 kDa band was processed and subjected to LC-MS/MS and SEQUEST analyses. The results of LC-MS/MS and SEQUEST search showed the presence of α(2u)-globulin in the 17 kDa band. In addition, the identified α(2u)-globulin sequence possessed GDW residues, which are the characteristic signature for lipocalin family. Since the α(2u)-globulin has been reported from the pheromone-carrying proteins in some mammals, this protein may carry the volatiles (pheromone compounds) in male Blackbucks preorbital gland to evoke the scent marking for maintaining territoriality (home range) and attraction towards female, through the secretion of glandular protein.

Serine Protease PrtA from Streptococcus pneumoniae Plays a Role in the Killing of S. pneumoniae by Apolactoferrin ▿

PubMed Central

Mirza, Shaper; Wilson, Landon; Benjamin, William H.; Novak, Jan; Barnes, Stephen; Hollingshead, Susan K.; Briles, David E.

2011-01-01

It is known that apolactoferrin, the iron-free form of human lactoferrin, can kill many species of bacteria, including Streptococcus pneumoniae. Lactoferricin, an N-terminal peptide of apolactoferrin, and fragments of it are even more bactericidal than apolactoferrin. In this study we found that apolactoferrin must be cleaved by a serine protease in order for it to kill pneumococci. The serine protease inhibitors were able to block killing by apolactoferrin but did not block killing by a lactoferrin-derived peptide. Thus, the killing of pneumococci by apolactoferrin appears to require a protease to release a lactoferricin-like peptide(s). Incubation of apolactoferrin with growing pneumococci resulted in a 12-kDa reduction in its molecular mass, of which about 7 to 8 kDa of the reduction was protease dependent. Capsular type 2 and 19F strains with mutations in the gene encoding the major cell wall-associated serine protease, prtA, lost much of their ability to degrade apolactoferrin and were relatively resistant to killing by apolactoferrin (P < 0.001). Recombinant PrtA was also able to cleave apolactoferrin, reducing its mass by about 8 kDa, and greatly enhance the killing activity of the solution containing the apolactoferrin and its cleavage products. Mass spectroscopy revealed that PrtA makes a major cut between amino acids 78 and 79 of human lactoferrin, removing the N-terminal end of the molecule (about 8.6 kDa). The simplest interpretation of these data is that the mechanism by which apolactoferrin kills Streptococcus pneumoniae requires the release of a lactoferricin-like peptide(s) and that it is this peptide(s), and not the intact apolactoferrin, which kills pneumococci. PMID:21422179

Sequencing Larger Intact Proteins (30-70 kDa) with Activated Ion Electron Transfer Dissociation

NASA Astrophysics Data System (ADS)

Riley, Nicholas M.; Westphall, Michael S.; Coon, Joshua J.

2018-01-01

The analysis of intact proteins via mass spectrometry can offer several benefits to proteome characterization, although the majority of top-down experiments focus on proteoforms in a relatively low mass range (<30 kDa). Recent studies have focused on improving the analysis of larger intact proteins (up to 75 kDa), but they have also highlighted several challenges to be addressed. One major hurdle is the efficient dissociation of larger protein ions, which often to do not yield extensive fragmentation via conventional tandem MS methods. Here we describe the first application of activated ion electron transfer dissociation (AI-ETD) to proteins in the 30-70 kDa range. AI-ETD leverages infrared photo-activation concurrent to ETD reactions to improve sequence-informative product ion generation. This method generates more product ions and greater sequence coverage than conventional ETD, higher-energy collisional dissociation (HCD), and ETD combined with supplemental HCD activation (EThcD). Importantly, AI-ETD provides the most thorough protein characterization for every precursor ion charge state investigated in this study, making it suitable as a universal fragmentation method in top-down experiments. Additionally, we highlight several acquisition strategies that can benefit characterization of larger proteins with AI-ETD, including combination of spectra from multiple ETD reaction times for a given precursor ion, multiple spectral acquisitions of the same precursor ion, and combination of spectra from two different dissociation methods (e.g., AI-ETD and HCD). In all, AI-ETD shows great promise as a method for dissociating larger intact protein ions as top-down proteomics continues to advance into larger mass ranges. [Figure not available: see fulltext.

Light propagation in Swiss-cheese models of random close-packed Szekeres structures: Effects of anisotropy and comparisons with perturbative results

NASA Astrophysics Data System (ADS)

Koksbang, S. M.

2017-03-01

Light propagation in two Swiss-cheese models based on anisotropic Szekeres structures is studied and compared with light propagation in Swiss-cheese models based on the Szekeres models' underlying Lemaitre-Tolman-Bondi models. The study shows that the anisotropy of the Szekeres models has only a small effect on quantities such as redshift-distance relations, projected shear and expansion rate along individual light rays. The average angular diameter distance to the last scattering surface is computed for each model. Contrary to earlier studies, the results obtained here are (mostly) in agreement with perturbative results. In particular, a small negative shift, δ DA≔D/A-DA ,b g DA ,b g , in the angular diameter distance is obtained upon line-of-sight averaging in three of the four models. The results are, however, not statistically significant. In the fourth model, there is a small positive shift which has an especially small statistical significance. The line-of-sight averaged inverse magnification at z =1100 is consistent with 1 to a high level of confidence for all models, indicating that the area of the surface corresponding to z =1100 is close to that of the background.

Plant mitochondrial pyruvate dehydrogenase complex: purification and identification of catalytic components in potato.

PubMed Central

Millar, A H; Knorpp, C; Leaver, C J; Hill, S A

1998-01-01

The pyruvate dehydrogenase complex (mPDC) from potato (Solanum tuberosum cv. Romano) tuber mitochondria was purified 40-fold to a specific activity of 5.60 micromol/min per mg of protein. The activity of the complex depended on pyruvate, divalent cations, NAD+ and CoA and was competitively inhibited by both NADH and acetyl-CoA. SDS/PAGE revealed the complex consisted of seven polypeptide bands with apparent molecular masses of 78, 60, 58, 55, 43, 41 and 37 kDa. N-terminal sequencing revealed that the 78 kDa protein was dihydrolipoamide transacetylase (E2), the 58 kDa protein was dihydrolipoamide dehydrogenase (E3), the 43 and 41 kDa proteins were alpha subunits of pyruvate dehydrogenase, and the 37 kDa protein was the beta subunit of pyruvate dehydrogenase. N-terminal sequencing of the 55 kDa protein band yielded two protein sequences: one was another E3; the other was similar to the sequence of E2 from plant and yeast sources but was distinctly different from the sequence of the 78 kDa protein. Incubation of the mPDC with [2-14C]pyruvate resulted in the acetylation of both the 78 and 55 kDa proteins. PMID:9729464

Vocal-fold collision mass as a differentiator between registers in the low-pitch range.

PubMed

Vilkman, E; Alku, P; Laukkanen, A M

1995-03-01

Register shift between the chest and falsetto register is generally studied in the higher-than-speaking pitch range. However, a similar difference can also be produced at speaking pitch level. The shift from breathy "falsetto" phonation to normal chest voice phonation was studied in normal female (pitch range 170-180 Hz) and male (pitch range 94-110 Hz) subjects. The phonations gliding from falsetto to normal chest voice were analyzed using iterative adaptive inverse filtering and electroglottography. Both trained and untrained, as well as female and male subjects, were able to produce an abrupt register shift from soft falsetto to soft chest register phonation. The differences between male and female speakers in the glottal flow waveforms were smaller than expected. The register shift is interpreted in terms of a "critical mass" concept of chest register phonation.

Isolation, purification and characterization of collagenase from hepatopancreas of the land snail Achatina fulica.

PubMed

Indra, D; Ramalingam, K; Babu, Mary

2005-09-01

Collagenase (matrix metalloproteinase-1, EC:3.4.24.7) was isolated from the hepatopancreas of Achatina fulica and characterized for its enzymatic activity and immunological properties. Procollagenase was isolated using ammonium sulphate precipitation and gel filtration, followed by purification by reverse-phase high performance liquid chromatography in the presence of trifluoroacetic acid and by dialysis in neutral buffer. In the presence of SDS and beta-mercaptoethanol, the procollagenase resolved into two subunits with molecular masses of 63 and 28 kDa, respectively. The 63 kDa fragment retained its ability to bind and degrade gelatin, but the 28 kDa was inactive. Analysis by 2D gel electrophoresis revealed that the 63 kDa fragment was basic (pIs 7.6, 7.8 and 8.15), while the 28 kDa fragment was acidic (pI 4.7 and 5.1). Western blot analysis confirmed the identity of collagenase, as only matrix metalloproteinase-1 rabbit antibodies against human matrix metalloproteinase-1 (N-terminal region) recognized both the isolated procollagenase and the 63 kDa fragment.

The proline-rich region of 18.5 kDa myelin basic protein binds to the SH3-domain of Fyn tyrosine kinase with the aid of an upstream segment to form a dynamic complex in vitro

PubMed Central

De Avila, Miguel; Vassall, Kenrick A.; Smith, Graham S. T.; Bamm, Vladimir V.; Harauz, George

2014-01-01

The intrinsically disordered 18.5 kDa classic isoform of MBP (myelin basic protein) interacts with Fyn kinase during oligodendrocyte development and myelination. It does so primarily via a central proline-rich SH3 (Src homology 3) ligand (T92–R104, murine 18.5 kDa MBP sequence numbering) that is part of a molecular switch due to its high degree of conservation and modification by MAP (mitogen-activated protein) and other kinases, especially at residues T92 and T95. Here, we show using co-transfection experiments of an early developmental oligodendroglial cell line (N19) that an MBP segment upstream of the primary ligand is involved in MBP–Fyn–SH3 association in cellula. Using solution NMR spectroscopy in vitro, we define this segment to comprise MBP residues (T62–L68), and demonstrate further that residues (V83–P93) are the predominant SH3-target, assessed by the degree of chemical shift change upon titration. We show by chemical shift index analysis that there is no formation of local poly-proline type II structure in the proline-rich segment upon binding, and by NOE (nuclear Overhauser effect) and relaxation measurements that MBP remains dynamic even while complexed with Fyn–SH3. The association is a new example first of a non-canonical SH3-domain interaction and second of a fuzzy MBP complex. PMID:25343306

The proline-rich region of 18.5 kDa myelin basic protein binds to the SH3-domain of Fyn tyrosine kinase with the aid of an upstream segment to form a dynamic complex in vitro.

PubMed

De Avila, Miguel; Vassall, Kenrick A; Smith, Graham S T; Bamm, Vladimir V; Harauz, George

2014-12-08

The intrinsically disordered 18.5 kDa classic isoform of MBP (myelin basic protein) interacts with Fyn kinase during oligodendrocyte development and myelination. It does so primarily via a central proline-rich SH3 (Src homology 3) ligand (T92-R104, murine 18.5 kDa MBP sequence numbering) that is part of a molecular switch due to its high degree of conservation and modification by MAP (mitogen-activated protein) and other kinases, especially at residues T92 and T95. Here, we show using co-transfection experiments of an early developmental oligodendroglial cell line (N19) that an MBP segment upstream of the primary ligand is involved in MBP-Fyn-SH3 association in cellula. Using solution NMR spectroscopy in vitro, we define this segment to comprise MBP residues (T62-L68), and demonstrate further that residues (V83-P93) are the predominant SH3-target, assessed by the degree of chemical shift change upon titration. We show by chemical shift index analysis that there is no formation of local poly-proline type II structure in the proline-rich segment upon binding, and by NOE (nuclear Overhauser effect) and relaxation measurements that MBP remains dynamic even while complexed with Fyn-SH3. The association is a new example first of a non-canonical SH3-domain interaction and second of a fuzzy MBP complex.

Diet quality and sleep quality among day and night shift nurses.

PubMed

Beebe, Deborah; Chang, Jen Jen; Kress, Kathleen; Mattfeldt-Beman, Mildred

2017-10-01

To determine whether night shift workers have a poorer diet quality and sleep quality when compared with day shift nurses. There is a dearth of research investigating the association between diet quality and sleep quality of day and night shift nurses. Data on nurses (n = 103) working either a day or night shift from two Midwestern hospitals were obtained from August 2015 to February 2016. The instruments used were the Diet History Questionnaire and the Pittsburg Sleep Quality Index. Independent samples t-tests were used to examine differences in diet and sleep quality by work shift schedule. There were no statistically significant differences between nurses working day or night shift and sleep quality (P = 0.0684), as well as diet quality (P = 0.6499). There was a significant difference between both body mass index (P = 0.0014) and exercise (P = 0.0020) with regard to diet quality. Body mass index and sleep quality were also significantly associated (P = 0.0032). Our study found no differences between day and night shift with regard to sleep and diet quality among nurses. Deliberate health initiatives and wellness programmes specifically targeting nurses are needed to increase knowledge about maintaining a healthy lifestyle while working as a nurse, whether it is day or night shift. © 2017 John Wiley & Sons Ltd.

Feature Based Retention Time Alignment for Improved HDX MS Analysis

NASA Astrophysics Data System (ADS)

Venable, John D.; Scuba, William; Brock, Ansgar

2013-04-01

An algorithm for retention time alignment of mass shifted hydrogen-deuterium exchange (HDX) data based on an iterative distance minimization procedure is described. The algorithm performs pairwise comparisons in an iterative fashion between a list of features from a reference file and a file to be time aligned to calculate a retention time mapping function. Features are characterized by their charge, retention time and mass of the monoisotopic peak. The algorithm is able to align datasets with mass shifted features, which is a prerequisite for aligning hydrogen-deuterium exchange mass spectrometry datasets. Confidence assignments from the fully automated processing of a commercial HDX software package are shown to benefit significantly from retention time alignment prior to extraction of deuterium incorporation values.

Quantitative LC-MS of polymers: determining accurate molecular weight distributions by combined size exclusion chromatography and electrospray mass spectrometry with maximum entropy data processing.

PubMed

Gruendling, Till; Guilhaus, Michael; Barner-Kowollik, Christopher

2008-09-15

We report on the successful application of size exclusion chromatography (SEC) combined with electrospray ionization mass spectrometry (ESI-MS) and refractive index (RI) detection for the determination of accurate molecular weight distributions of synthetic polymers, corrected for chromatographic band broadening. The presented method makes use of the ability of ESI-MS to accurately depict the peak profiles and retention volumes of individual oligomers eluting from the SEC column, whereas quantitative information on the absolute concentration of oligomers is obtained from the RI-detector only. A sophisticated computational algorithm based on the maximum entropy principle is used to process the data gained by both detectors, yielding an accurate molecular weight distribution, corrected for chromatographic band broadening. Poly(methyl methacrylate) standards with molecular weights up to 10 kDa serve as model compounds. Molecular weight distributions (MWDs) obtained by the maximum entropy procedure are compared to MWDs, which were calculated by a conventional calibration of the SEC-retention time axis with peak retention data obtained from the mass spectrometer. Comparison showed that for the employed chromatographic system, distributions below 7 kDa were only weakly influenced by chromatographic band broadening. However, the maximum entropy algorithm could successfully correct the MWD of a 10 kDa standard for band broadening effects. Molecular weight averages were between 5 and 14% lower than the manufacturer stated data obtained by classical means of calibration. The presented method demonstrates a consistent approach for analyzing data obtained by coupling mass spectrometric detectors and concentration sensitive detectors to polymer liquid chromatography.

Development of a simultaneous PET/microdialysis method to identify the optimal dose of 11C-raclopride for small animal imaging

PubMed Central

Schiffer, Wynne K.; Alexoff, David L.; Shea, Colleen; Logan, Jean; Dewey, Stephen L.

2005-01-01

In the field of small animal positron emission tomography (PET), the assumptions underlying human and primate kinetic models may not be sustained in rodents. That is, the threshold dose at which a pharmacologic response occurs may be lower in small animals. In order to define this relationship, we combined microPET imaging using 11C-raclopride with microdialysis measures of extracellular fluid (ECF) dopamine (DA). In addition, we performed a series of studies in which a known mass of raclopride was microinfused into one striatum prior to a high specific activity (SA) systemic injection of 11C-raclopride. This single-injection approach provided a high and low SA region of radiotracer binding in the same animal during the same scanning session. Our data demonstrate that the binding potential (BP) declines above 3.5 pmol/ml (0.35 μg), with an ED50 of 8.55 ± 5.62 pmol/ml. These data also provide evidence that BP may be compromised by masses of raclopride below 2.0 pmol/ml (0.326 μg). Increases in ECF DA were produced by mass doses of raclopride over 3.9 pmol/ml (0.329 μg) with an ED50 of 8.53 ± 2.48 pmol/ml. Taken together, it appears that an optimal range of raclopride mass exists between 2.0 and 3.5 pmol/ml, around which the measured BP can be compromised by system sensitivity, endogenous DA, or excessive competition with unlabeled compound. PMID:15848236

Mass sensor based on split-nanobeam optomechanical oscillator

NASA Astrophysics Data System (ADS)

Zhang, Yeping; Ai, Jie; Xiang, Yanjun; He, Qinghua; Li, Tao; Ma, Jingfang

2016-03-01

Mass sensing based on monitoring the frequency shifts induced by added mass in oscillators is a well-known and widely used technique. The optomechanical crystal cavity has strong interaction between optical mode and mechanical mode. Radiation pressure driven optomechanical crystal cavity are excellent candidates for mass detection due to their simplicity, sensitivity and all optical operation. In an optomechanical crystal cavity, a high quality factor optical mode simultaneously serves as an efficient actuator and a sensitive probe for precise monitoring the mechanical frequency change of the cavity structure. Here, a split-nanobeam optomechanical crystal cavity is proposed, the sensing resolution as small as 0.33ag (1ag=10-21kg) and the frequency shift is more than 30MHz. This is important and promising for achieve ultimate-precision mass sensing including proteins and other molecules.

Atmospheric pressure chemical ionisation mass spectrometry analysis linked with chemometrics for food classification - a case study: geographical provenance and cultivar classification of monovarietal clarified apple juices.

PubMed

Gan, Heng-Hui; Soukoulis, Christos; Fisk, Ian

2014-03-01

In the present work, we have evaluated for first time the feasibility of APCI-MS volatile compound fingerprinting in conjunction with chemometrics (PLS-DA) as a new strategy for rapid and non-destructive food classification. For this purpose 202 clarified monovarietal juices extracted from apples differing in their botanical and geographical origin were used for evaluation of the performance of APCI-MS as a classification tool. For an independent test set PLS-DA analyses of pre-treated spectral data gave 100% and 94.2% correct classification rate for the classification by cultivar and geographical origin, respectively. Moreover, PLS-DA analysis of APCI-MS in conjunction with GC-MS data revealed that masses within the spectral ACPI-MS data set were related with parent ions or fragments of alkyesters, carbonyl compounds (hexanal, trans-2-hexenal) and alcohols (1-hexanol, 1-butanol, cis-3-hexenol) and had significant discriminating power both in terms of cultivar and geographical origin. Copyright © 2013 The Authors. Published by Elsevier Ltd.. All rights reserved.

Endotoxin induction of an inhibitor of plasminogen activator in bovine pulmonary artery endothelial cells

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

Not Available

The effects of bacterial lipopolysaccharide (endotoxin) on the fibrinolytic activity of bovine pulmonary artery endothelial cells were examined. Endotoxin suppressed the net fibrinolytic activity of cell extracts and conditioned media in a dose-dependent manner. The effects of endotoxin required at least 6 h for expression. Cell extracts and conditioned media contained a 44-kDa urokinase-like plasminogen activator. Media also contained multiple plasminogen activators with molecular masses of 65-75 and 80-100 kDa. Plasminogen activators in extracts and media were unchanged by treatment of cells with endotoxin. Diisopropyl fluorophosphate (DFP)-abolished fibrinolytic activity of extracts and conditioned media. DFP-treated samples from endotoxin-treated but notmore » untreated cells inhibited urokinase and tissue plasminogen activator, but not plasmin. Inhibitory activity was lost by incubation at pH 3 or heating to 56/sup 0/C for 10 min. These treatments did not affect inhibitory activity of fetal bovine serum. Incubation of /sup 125/I-urokinase with DFP-treated medium from endotoxin-treated cells produced an inactive complex with an apparent molecular mass of 80-85 kDa.« less

NMR resonance assignments of the lantibiotic immunity protein NisI from Lactococcus lactis.

PubMed

Hacker, Carolin; Christ, Nina Alexandra; Duchardt-Ferner, Elke; Korn, Sophie; Berninger, Lucija; Kötter, Peter; Entian, Karl-Dieter; Wöhnert, Jens

2015-10-01

The lantibiotic nisin is a small antimicrobial peptide which acts against a wide range of Gram-positive bacteria. Nisin-producing Lactococcus lactis strains express four genes for self-protection against their own antimicrobial compound. This immunity system consists of the lipoprotein NisI and the ABC transporter NisFEG. NisI is attached to the outside of the cytoplasmic membrane via a covalently linked diacylglycerol anchor. Both the lipoprotein and the ABC transporter are needed for full immunity but the exact immunity mechanism is still unclear. To gain insights into the highly specific immunity mechanism of nisin producing strains on a structural level we present here the backbone resonance assignment of NisI (25.8 kDa) as well as the virtually complete (1)H,(15)N,(13)C chemical shift assignments for the isolated 12.7 kDa N-terminal and 14.6 kDa C-terminal domains of NisI.

Purification and chemical characterisation of a cell wall-associated β-galactosidase from mature sweet cherry (Prunus avium L.) fruit.

PubMed

Gerardi, Carmela; Blando, Federica; Santino, Angelo

2012-12-01

Using four different chromatographic steps, β-galactosidase was purified from the ripe fruit of sweet cherry to apparent electrophoretic homogeneity with approximately 131-fold purification. The Prunus avium β-galactosidase showed an apparent molecular mass of about 100 kDa and consisted of four different active polypeptides with pIs of about 7.9, 7.4, 6.9 and 6.4 as estimated by native IEF and β-galactosidase-activity staining. The active polypeptides were individually excised from the gel and subjected to SDS-PAGE. Each of the four native enzymes showing β-galactosidase activity was composed of two polypeptides with an estimated mass of 54 and 33 kDa. Both of these polypeptides were subjected to N-terminal amino acid sequence analysis. The 54 kDa polypeptide of sweet cherry β-galactosidase showed a 43% identity with the 44 kDa subunit of persimmon and apple β-galactosidases and the 48 kDa subunit of carambola galactosidase I. The sweet cherry β-galactosidase exhibited a strict specificity towards p-nitrophenyl β-D-galactopyranoside, a pH optimum of 4.0 and K(m) and V(max) values of 0.42 mM and 4.12 mmol min(-1) mg(-1) of protein respectively with this substrate. The enzyme was also active towards complex glycans. Taken together the results of this study prompted a role for this class of enzymes on sweet cherry fruit ripening and softening. Copyright © 2012 Elsevier Masson SAS. All rights reserved.

Detection of prosecretory mitogen lacritin in nonprimate tears primarily as a C-terminal-like fragment.

PubMed

Laurie, Diane E; Splan, Rebecca K; Green, Kari; Still, Katherine M; McKown, Robert L; Laurie, Gordon W

2012-09-12

Lacritin is a human tear glycoprotein that promotes basal tear protein secretion in cultured rat lacrimal acinar cells and proliferation of subconfluent human corneal epithelial cells. When topically added to rabbit eyes, lacritin promotes basal tearing. Despite these activities on several species, lacritin's presence in nonprimate tears or other tissues has not been explored. Here we probed for lacritin in normal horse tears. Sequences were collected from the Ensembl genomic alignment of human LACRT gene with high-quality draft horse genome (EquCab2.0) and analyzed. Normal horse tears were collected and assayed by Western blotting, ELISA, and mass spectrometry. Newly generated rabbit antibodies, respectively, against N- and C-terminal regions of human lacritin were employed. Identity was 75% and 45%, respectively, at nucleotide and protein levels. Structural features were conserved, including a C-terminal amphipathic α-helix. Anti-C-terminal antibodies strongly detected a ∼13 kDa band in horse tears that was validated by mass spectrometry. In human tears, the same antibody detected uncleaved lacritin (∼24 kDa) strongly and C-terminal fragments of ∼13 and ∼11 kDa weakly. Anti-N-terminal antibodies were slightly reactive with a ∼24 kDa horse antigen and showed no reaction with the anti-C-terminal-reactive ∼13 kDa species. Similar respective levels of horse C-terminal versus N-terminal immunoreactivity were apparent by ELISA. Lacritin is present in horse tears, largely as a C-terminal fragment homologous to the mitogenic and bactericidal region in human lacritin, suggesting potential benefit in corneal wound repair.

Plant Ecological Strategies Shift Across the Cretaceous–Paleogene Boundary

PubMed Central

Blonder, Benjamin; Royer, Dana L.; Johnson, Kirk R.; Miller, Ian; Enquist, Brian J.

2014-01-01

The Chicxulub bolide impact caused the end-Cretaceous mass extinction of plants, but the associated selectivity and ecological effects are poorly known. Using a unique set of North Dakota leaf fossil assemblages spanning 2.2 Myr across the event, we show among angiosperms a reduction of ecological strategies and selection for fast-growth strategies consistent with a hypothesized recovery from an impact winter. Leaf mass per area (carbon investment) decreased in both mean and variance, while vein density (carbon assimilation rate) increased in mean, consistent with a shift towards “fast” growth strategies. Plant extinction from the bolide impact resulted in a shift in functional trait space that likely had broad consequences for ecosystem functioning. PMID:25225914

Rapid identif ication and comparative analysis of chemical constituents in herbal medicine Fufang decoction by ultra-high-pressure liquid chromatography coupled with a hybrid linear ion trap-high-resolution mass spectrometry.

PubMed

Cao, Gang; Chen, Xiaocheng; Wu, Xin; Li, Qinglin; Zhang, Hongyan

2015-05-01

This study was conducted to reveal the relation between herbal medicine Fufang decoction and a single drug in terms of material base. Da-Cheng-Qi decoction (DCQD) was used as a model. Ultrahigh-pressure liquid chromatography coupled with a hybrid linear ion trap-high-resolution mass spectrometry (UHPLC-LTQ-Orbitrap) was applied to detect and identify the main chemical compounds. This technique was also employed to determine the different chemical components. Under optimized liquid chromatography and mass spectrometry conditions, 64 components, including iridoids, flavonoids, anthraquinones and coumarins, were separated and tentatively characterized in Da-Cheng-Qi decoction. After decoction, the contents of 18 compounds were markedly changed, and two components were no longer detected in Fufang decoction compared with single-medicine decoction. The established method provided a good example for the rapid identification of complicated polar constituents in herbal medicine prescriptions. Copyright © 2014 John Wiley & Sons, Ltd.

Identification of a major yolk protein as an allergen in sea urchin roe.

PubMed

Yamasaki, Ayako; Higaki, Hiromi; Nakashima, Keiko; Yamamoto, Osamu; Hein, Kyaw Zaw; Takahashi, Hitoshi; Chinuki, Yuko; Morita, Eishin

2010-05-01

Anaphylaxis after eating sea urchin roe has been reported. However, its major allergens have not yet been identified. The aim of this study was to identify the major allergens of sea urchin roe. Proteins of sea urchin roe were separated by sodium dodecyl sulphate-polyacrylamide gel electrophoresis and two-dimensional electrophoresis (2-DE). An immunoglobulin (Ig)E-binding protein was detected by immunoblotting using the patient's serum. An allergen isolated from 2DE-gel was identified by peptide mass fingerprinting using matrix-assisted laser desorption/ionization-time of flight-mass spectrometry. Immunoblot analysis of sea urchin extracts showed that a 160-kDa protein at pI 6-7 was recognized by the patient's IgE. Peptide mass fingerprint analysis revealed that the protein was the major yolk protein (152 kDa, pI 6.9) of sea urchins. The results show that a major allergen of sea urchin roe is the major yolk protein.

An asteroseismic constraint on the mass of the axion from the period drift of the pulsating DA white dwarf star L19-2

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

Córsico, Alejandro H.; Althaus, Leandro G.; Bertolami, Marcelo M. Miller

We employ an asteroseismic model of L19-2, a relatively massive ( M {sub *} ∼ 0.75 M {sub ⊙}) and hot ( T {sub eff} ∼ 12 100 K) pulsating DA (H-rich atmosphere) white dwarf star (DAV or ZZ Ceti variable), and use the observed values of the temporal rates of period change of its dominant pulsation modes (Π ∼ 113 s and Π ∼ 192 s), to derive a new constraint on the mass of the axion, the hypothetical non-barionic particle considered as a possible component of the dark matter of the Universe. If the asteroseismic model employed ismore » an accurate representation of L19-2, then our results indicate hints of extra cooling in this star, compatible with emission of axions of mass m {sub a} cos{sup 2}β ∼< 25 meV or an axion-electron coupling constant of g {sub ae} ∼< 7 × 10{sup −13}.« less

Electrospray-assisted laser desorption/ionization and tandem mass spectrometry of peptides and proteins.

PubMed

Peng, Ivory X; Shiea, Jentaie; Ogorzalek Loo, Rachel R; Loo, Joseph A

2007-01-01

We have constructed an electrospray-assisted laser desorption/ionization (ELDI) source which utilizes a nitrogen laser pulse to desorb intact molecules from matrix-containing sample solution droplets, followed by electrospray ionization (ESI) post-ionization. The ELDI source is coupled to a quadrupole ion trap mass spectrometer and allows sampling under ambient conditions. Preliminary data showed that ELDI produces ESI-like multiply charged peptides and proteins up to 29 kDa carbonic anhydrase and 66 kDa bovine albumin from single-protein solutions, as well as from complex digest mixtures. The generated multiply charged polypeptides enable efficient tandem mass spectrometric (MS/MS)-based peptide sequencing. ELDI-MS/MS of protein digests and small intact proteins was performed both by collisionally activated dissociation (CAD) and by nozzle-skimmer dissociation (NSD). ELDI-MS/MS may be a useful tool for protein sequencing analysis and top-down proteomics study, and may complement matrix-assisted laser desorption/ionization (MALDI)-based measurements. Copyright (c) 2007 John Wiley & Sons, Ltd.

A NMR experiment for simultaneous correlations of valine and leucine/isoleucine methyls with carbonyl chemical shifts in proteins.

PubMed

Tugarinov, Vitali; Venditti, Vincenzo; Marius Clore, G

2014-01-01

A methyl-detected 'out-and-back' NMR experiment for obtaining simultaneous correlations of methyl resonances of valine and isoleucine/leucine residues with backbone carbonyl chemical shifts, SIM-HMCM(CGCBCA)CO, is described. The developed pulse-scheme serves the purpose of convenience in recording a single data set for all Ile(δ1), Leu(δ) and Val(γ) (ILV) methyl positions instead of acquiring two separate spectra selective for valine or leucine/isoleucine residues. The SIM-HMCM(CGCBCA)CO experiment can be used for ILV methyl assignments in moderately sized protein systems (up to ~100 kDa) where the backbone chemical shifts of (13)C(α), (13)Cβ and (13)CO are known from prior NMR studies and where some losses in sensitivity can be tolerated for the sake of an overall reduction in NMR acquisition time.

Characterization of a novel Y2K-type dehydrin VrDhn1 from Vigna radiata.

PubMed

Lin, Chia-Hui; Peng, Po-Hsin; Ko, Chia-Yun; Markhart, Albert H; Lin, Tsai-Yun

2012-05-01

A novel dehydrin gene (VrDhn1) was isolated from an embryo cDNA library of Vigna radiata (L.) Wilczek (mungbean) variety VC1973A. The intronless VrDhn1 gene encodes a protein belonging to the Y(2)K-type dehydrin family. VrDhn1 protein accumulated in embryos and cotyledons during seed maturation and disappeared 2 days after seed imbibition (DAI). The expression of VrDhn1 mRNA and accumulation of VrDhn1 protein were at high levels in mature seeds, but neither mRNA nor protein was detected in mungbean vegetative tissues under normal growth conditions. The VrDhn1 mRNA level was extremely high in mature seeds and decreased to ∼30% at 1 DAI, and was not detectable at ~7 DAI. Tissue dehydration, salinity and exogenous ABA markedly induced VrDhn1 transcripts in plants as measured by quantitative real-time reverse transcription-PCR (qRT-PCR). VrDhn1 protein was not detected using immunoblots in seedlings under stress treatments. In mature seeds or 1 DAI seedlings, VrDhn1 proteins were immunolocalized in the nucleus and cytoplasm. VrDhn1 exhibited low affinity for non-specific interaction with DNA using electrophoretic mobility shift assays (EMSAs), and the exogenous addition of Zn(2+) or Ni(2+) stimulated interaction. The His-tagged VrDhn1 (30.17 kDa) protein showed a molecular mass of 63.1 kDa on gel filtration, suggesting a dimer form. This is the first report showing that a Y(2)K-type VrDhn1 enters the nucleus and interacts with DNA during seed maturation.

Increase in local protein concentration by field-inversion gel electrophoresis.

PubMed

Tsai, Henghang; Low, Teck Yew; Freeby, Steve; Paulus, Aran; Ramnarayanan, Kalpana; Cheng, Chung-Pui Paul; Leung, Hon-Chiu Eastwood

2007-09-26

Proteins that migrate through cross-linked polyacrylamide gels (PAGs) under the influence of a constant electric field experience negative factors, such as diffusion and non-specific trapping in the gel matrix. These negative factors reduce protein concentrations within a defined gel volume with increasing migration distance and, therefore, decrease protein separation efficiency. Enhancement of protein separation efficiency was investigated by implementing pulsed field-inversion gel electrophoresis (FIGE). Separation of model protein species and large protein complexes was compared between FIGE and constant field electrophoresis (CFE) in different percentages of PAGs. Band intensities of proteins in FIGE with appropriate ratios of forward and backward pulse times were superior to CFE despite longer running times. These results revealed an increase in band intensity per defined gel volume. A biphasic protein relative mobility shift was observed in percentages of PAGs up to 14%. However, the effect of FIGE on protein separation was stochastic at higher PAG percentage. Rat liver lysates subjected to FIGE in the second-dimension separation of two-dimensional polyarcylamide gel electrophoresis (2D PAGE) showed a 20% increase in the number of discernible spots compared with CFE. Nine common spots from both FIGE and CFE were selected for peptide sequencing by mass spectrometry (MS), which revealed higher final ion scores of all nine protein spots from FIGE. Native protein complexes ranging from 800 kDa to larger than 2000 kDa became apparent using FIGE compared with CFE. The present investigation suggests that FIGE under appropriate conditions improves protein separation efficiency during PAGE as a result of increased local protein concentration. FIGE can be implemented with minimal additional instrumentation in any laboratory setting. Despite the tradeoff of longer running times, FIGE can be a powerful protein separation tool.

Osmotic Compounds Enhance Antibiotic Efficacy against Acinetobacter baumannii Biofilm Communities

PubMed Central

Falghoush, Azeza; Beyenal, Haluk; Besser, Thomas E.; Omsland, Anders

2017-01-01

ABSTRACT Biofilm-associated infections are a clinical challenge, in part because a hydrated matrix protects the bacterial community from antibiotics. Herein, we evaluated how different osmotic compounds (maltodextrin, sucrose, and polyethylene glycol [PEG]) enhance antibiotic efficacy against Acinetobacter baumannii biofilm communities. Established (24-h) test tube biofilms (strain ATCC 17978) were treated with osmotic compounds in the presence or absence of 10× the MIC of different antibiotics (50 μg/ml tobramycin, 20 μg/ml ciprofloxacin, 300 μg/ml chloramphenicol, 30 μg/ml nalidixic acid, or 100 μg/ml erythromycin). Combining antibiotics with hypertonic concentrations of the osmotic compounds for 24 h reduced the number of biofilm bacteria by 5 to 7 log (P < 0.05). Increasing concentrations of osmotic compounds improved the effect, but there was a trade-off with increasing solution viscosity, whereby low-molecular-mass compounds (sucrose, 400-Da PEG) worked better than higher-mass compounds (maltodextrin, 3,350-Da PEG). Ten other A. baumannii strains were similarly treated with 400-Da PEG and tobramycin, resulting in a mean 2.7-log reduction in recoverable bacteria compared with tobramycin treatment alone. Multivariate regression models with data from different osmotic compounds and nine antibiotics demonstrated that the benefit from combining hypertonic treatments with antibiotics is a function of antibiotic mass and lipophilicity (r2 > 0.82; P < 0.002), and the relationship was generalizable for biofilms formed by A. baumannii and Escherichia coli K-12. Augmenting topical antibiotic therapies with a low-mass hypertonic treatment may enhance the efficacy of antibiotics against wound biofilms, particularly when using low-mass hydrophilic antibiotics. IMPORTANCE Biofilms form a barrier that protects bacteria from environmental insults, including exposure to antibiotics. We demonstrated that multiple osmotic compounds can enhance antibiotic efficacy against Acinetobacter baumannii biofilm communities, but viscosity is a limiting factor, and the most effective compounds have lower molecular mass. The synergism between osmotic compounds and antibiotics is also dependent on the hydrophobicity and mass of the antibiotics. The statistical models presented herein provide a basis for predicting the optimal combination of osmotic compounds and antibiotics against surface biofilms communities. PMID:28733283

Peculiarities of hemoglobin interaction with serum proteins of mice with Ehrlich carcinoma.

PubMed

Sitdikova, S M; Amandzholov, B S; Serebryakova, M V; Zhdanovich, M Yu; Kiselevskii, M V; Donenko, F V

2006-05-01

In male C57Bl/6 mice with transplanted Ehrlich carcinoma, hemoglobin forms a complex with serum proteins characterized by a molecular weight of about 300 kDa. The complex incorporates proteins weighing 100, 68, 65, and 15 kDa identified by MALDI-TOF mass spectrometry as haptoglobin, serum albumin, gi/26341396 nameless protein Mus musculus, and alpha-hemoglobin, respectively. This complex can possess biological activity and contribute to the control of tumor growth.

A meiotic DNA polymerase from a mushroom, Agaricus bisporus.

PubMed Central

Takami, K; Matsuda, S; Sono, A; Sakaguchi, K

1994-01-01

A meiotic DNA polymerase [DNA nucleotidyltransferase (DNA-directed), EC 2.7.7.7], which likely has a role in meiotic DNA repair, was isolated from a mushroom, Agaricus bisporus. The purified fraction displays three bands in SDS/PAGE, at molecular masses of 72 kDa, 65 kDa and 36 kDa. Optimal activity is at pH 7.0-8.0 in the presence of 5 mM Mg2+ and 50 mM KCl and at 28-30 degrees C, which is the temperature for meiosis. This enzyme is resistant to N-ethylmaleimide and sensitive to 2',3'-dideoxythymidine 5'-triphosphate, suggesting that it is a beta-like DNA polymerase. These characteristics are similar to those of Coprinus DNA polymerase beta [Sakaguchi and Lu (1982) Mol. Cell. Biol. 2, 752-757]. In Western-blot analysis, the antiserum against the Coprinus polymerase reacts only with the 65 kDa band, which coincides with the molecular mass of the Coprinus polymerase. Western-blot analysis also showed that the antiserum could react with crude extracts not only from the Agaricales family, to which Agaricus and Coprinus belong, but also from different mushroom families and Saccharomyces. The Agaricus polymerase activity can be found only in the meiotic-cell-rich fraction, but the enzyme is also present in the somatic cells in an inactive state. Images Figure 2 Figure 5 Figure 6 PMID:8172591

Up-Regulation of Antioxidant Proteins in the Plasma Proteome during Saturation Diving: Unique Coincidence under Hypobaric Hypoxia.

PubMed

Domoto, Hideharu; Iwaya, Keiichi; Ikomi, Fumitaka; Matsuo, Hirotaka; Tadano, Yutaka; Fujii, Shigenori; Tachi, Kazuyoshi; Itoh, Yoshiyuki; Sato, Michiya; Inoue, Kimitoshi; Shinomiya, Nariyoshi

2016-01-01

Saturation diving (SD) is one of the safest techniques for tolerating hyperbaric conditions for long durations. However, the changes in the human plasma protein profile that occur during SD are unknown. To identify differential protein expression during or after SD, 65 blood samples from 15 healthy Japanese men trained in SD were analyzed by two-dimensional fluorescence difference gel electrophoresis. The expression of two proteins, one 32.4 kDa with an isoelectric point (pI) of 5.8 and the other 44.8 kDa with pI 4.0, were elevated during SD to 60, 100, and 200 meters sea water (msw). The expression of these proteins returned to pre-diving level when the SD training was completed. The two proteins were identified using in-gel digestion and mass spectrometric analysis; the 32.4 kDa protein was transthyretin and the 44.8 kDa protein was alpha-1-acid glycoprotein 1. Oxidation was detected at methionine 13 of transthyretin and at methionine 129 of alpha-1-acid glycoprotein 1 by tandem mass spectrometry. Moreover, haptoglobin was up-regulated during the decompression phase of 200 msw. These plasma proteins up-regulated during SD have a common function as anti-oxidants. This suggests that by coordinating their biological effects, these proteins activate a defense mechanism to counteract the effects of hyperbaric-hyperoxic conditions during SD.

Up-Regulation of Antioxidant Proteins in the Plasma Proteome during Saturation Diving: Unique Coincidence under Hypobaric Hypoxia

PubMed Central

Domoto, Hideharu; Iwaya, Keiichi; Ikomi, Fumitaka; Matsuo, Hirotaka; Tadano, Yutaka; Fujii, Shigenori; Tachi, Kazuyoshi; Itoh, Yoshiyuki; Sato, Michiya; Inoue, Kimitoshi; Shinomiya, Nariyoshi

2016-01-01

Saturation diving (SD) is one of the safest techniques for tolerating hyperbaric conditions for long durations. However, the changes in the human plasma protein profile that occur during SD are unknown. To identify differential protein expression during or after SD, 65 blood samples from 15 healthy Japanese men trained in SD were analyzed by two-dimensional fluorescence difference gel electrophoresis. The expression of two proteins, one 32.4 kDa with an isoelectric point (pI) of 5.8 and the other 44.8 kDa with pI 4.0, were elevated during SD to 60, 100, and 200 meters sea water (msw). The expression of these proteins returned to pre-diving level when the SD training was completed. The two proteins were identified using in-gel digestion and mass spectrometric analysis; the 32.4 kDa protein was transthyretin and the 44.8 kDa protein was alpha-1-acid glycoprotein 1. Oxidation was detected at methionine 13 of transthyretin and at methionine 129 of alpha-1-acid glycoprotein 1 by tandem mass spectrometry. Moreover, haptoglobin was up-regulated during the decompression phase of 200 msw. These plasma proteins up-regulated during SD have a common function as anti-oxidants. This suggests that by coordinating their biological effects, these proteins activate a defense mechanism to counteract the effects of hyperbaric-hyperoxic conditions during SD. PMID:27741252

The belonging of gpMuc, a glycoprotein from Mucuna pruriens seeds, to the Kunitz-type trypsin inhibitor family explains its direct anti-snake venom activity.

PubMed

Scirè, Andrea; Tanfani, Fabio; Bertoli, Enrico; Furlani, Emiliano; Nadozie, Hope-Onyekwere N; Cerutti, Helena; Cortelazzo, Alessio; Bini, Luca; Guerranti, Roberto

2011-07-15

In Nigeria, Mucuna pruriens seeds are locally prescribed as an oral prophylactic for snake bite and it is claimed that when two seeds are swallowed they protect the individual for a year against snake bites. In order to understand the Mucuna pruriens antisnake properties, the proteins from the acqueous extract of seeds were purified by three chromatographic steps: ConA affinity chromatography, tandem anionic-cationic exchange and gel filtration, obtaining a fraction conventionally called gpMucB. This purified fraction was analysed by SDS-PAGE obtaining 3 bands with apparent masses ranging from 20 to 24 kDa, and by MALDI-TOF which showed two main peaks of 21 and 23 kDa and another small peak of 19 kDa. On the other hand, gel filtration analysis of the native protein indicated a molecular mass of about 70 kDa suggesting that in its native form, gpMucB is most likely an oligomeric multiform protein. Infrared spectroscopy of gpMucB indicated that the protein is particularly thermostable both at neutral and acidic pHs and that it is an all beta protein. All data suggest that gpMucB belongs to the Kunitz-type trypsin inhibitor family explaining the direct anti-snake venom activity of Mucuna pruriens seeds. Copyright © 2011 Elsevier GmbH. All rights reserved.

Fourier Transform-Ion Cyclotron Resonance Mass Spectrometry as a Platform for Characterizing Multimeric Membrane Protein Complexes

NASA Astrophysics Data System (ADS)

Lippens, Jennifer L.; Nshanian, Michael; Spahr, Chris; Egea, Pascal F.; Loo, Joseph A.; Campuzano, Iain D. G.

2018-01-01

Membrane protein characterization is consistently hampered by challenges with expression, purification, and solubilization. Among several biophysical techniques employed for their characterization, native-mass spectrometry (MS) has emerged as a powerful tool for the analysis of membrane proteins and complexes. Here, two MS platforms, the FT-ICR and Q-ToF, have been explored to analyze the homotetrameric water channel protein, AquaporinZ (AqpZ), under non-denaturing conditions. This 97 kDa membrane protein complex can be readily liberated from the octylglucoside (OG) detergent micelle under a range of instrument conditions on both MS platforms. Increasing the applied collision energy of the FT-ICR collision cell yielded varying degrees of tetramer (97 kDa) liberation from the OG micelles, as well as dissociation into the trimeric (72 kDa) and monomeric (24 kDa) substituents. Tandem-MS on the Q-ToF yielded higher intensity tetramer signal and, depending on the m/z region selected, the observed monomer signal varied in intensity. Precursor ion selection of an m/z range above the expected protein signal distribution, followed by mild collisional activation, is able to efficiently liberate AqpZ with a high S/N ratio. The tetrameric charge state distribution obtained on both instruments demonstrated superpositioning of multiple proteoforms due to varying degrees of N-terminal formylation. [Figure not available: see fulltext.

HDA1 and HDA3 are components of a yeast histone deacetylase (HDA) complex.

PubMed

Carmen, A A; Rundlett, S E; Grunstein, M

1996-06-28

Histone acetylation is maintained through the action of histone acetyltransferases and deacetylases and has been correlated with increased gene activity. To investigate the functional role of these enzymes in the regulation of transcription, we have purified from Saccharomyces cerevisiae two histone deacetylase activities, HDA and HDB, with molecular masses of approximately 350 and 600 kDa, respectively. In vitro, the HDA activity deacetylates all four core histones, has a preference for histone H3, and is strongly inhibited by trichostatin A (a specific inhibitor of histone deacetylases). HDB is considerably less sensitive to trichostatin A. We report the extensive purification of the HDA activity and the identification of peptides (p75, p73, p72, and p71) whose presence correlates with deacetylase activity on native polyacrylamide gels. An antibody to p75 immunoprecipitates peptides with molecular masses similar to those in the 350-kDa complex. Additionally, antibodies to p75 and p71 specifically precipitate histone deacetylase activity and co-immunoprecipitate each other. Gene disruptions of p75 (HDA1) or p71 (HDA3) cause the loss of the 350-kDa (but not the 600-kDa) activity from our chromatography profiles. These data argue strongly that HDA1 and HDA3 are subunits of the HDA complex, which is structurally distinct from the second, HDB complex.

Kinetics and mechanism investigation on the destruction of oxytetracycline by UV-254nm activation of persulfate.

PubMed

Liu, Yiqing; He, Xuexiang; Fu, Yongsheng; Dionysiou, Dionysios D

2016-03-15

Oxytetracycline (OTC), an important broad-spectrum antibiotic, has been detected extensively in various environmental systems, which may have a detrimental impact on ecosystem and human health through the development of drug resistant bacteria and pathogens. In this study, the degradation of OTC was evaluated by UV-254nm activated persulfate (PS). The observed UV fluence based pseudo first-order rate constant (kobs) was found to be the highest at near neutral pH conditions (pH 5.5-8.5). Presence of various natural water constituents had different effects on OTC degradation, with a significant enhancement in the presence of bicarbonate or Cu(2+). Limited elimination of total organic carbon (TOC) and PS was observed during the mineralization of OTC. Transformation byproducts in the presence and absence of hydroxyl radical scavenging agent tert-butanol (t-BuOH) were identified using ultra-high definition accurate-mass quadrupole time-of-flight liquid chromatography/mass spectrometer (LC-QTOF/MS). Potential OTC degradation mechanism was subsequently proposed revealing four different reaction pathways by SO4(-) reaction including hydroxylation (+16Da), demethylation (-14Da), decarbonylation (-28Da) and dehydration (-18Da). This study suggests that UV-254nm/PS is a promising treatment technology for the control of water pollution caused by emerging contaminants such as OTC. Copyright © 2015 Elsevier B.V. All rights reserved.

Bacteriocins from Gram-Negative Bacteria: A Classification?

NASA Astrophysics Data System (ADS)

Rebuffat, Sylvie

Bacteria produce an arsenal of toxic peptides and proteins, which are termed bacteriocins and play a role in mediating the dynamics of microbial populations and communities. Bacteriocins from Gram-negative bacteria arise mainly from Enterobacteriaceae. They assemble into two main families: high molecular mass modular proteins (30-80 kDa) termed colicins and low molecular mass peptides (between 1 and 10 kDa) termed microcins. The production of colicins is mediated by the SOS response regulon, which plays a role in the response of many bacteria to DNA damages. Microcins are highly stable hydrophobic peptides that are produced under stress conditions, particularly nutrient depletion. Colicins and microcins are found essentially in Escherichia coli, but several other Gram-negative species also produce bacteriocin-like substances. This chapter presents the basis of a classification of colicins and microcins.

[Comparative study of sleep patterns in nurses working day and night shifts].

PubMed

De Martino, Milva Maria Figueiredo

2002-08-01

To compare sleep patterns in nurses working day and night shifts in a hospital in Campinas (SP), Brazil. Fifty-nine nurses between 23 and 59 years of age participated in the study. For day shift workers, the pattern of nocturnal sleep was examined; for night shift workers, nocturnal and diurnal sleep patterns were examined. During 1 week, participants filled out a sleep diary right after waking up. The following items were assessed: time going to bed, falling asleep, and waking up; sleep latency; duration in hours of nocturnal and diurnal sleep; naps; quality of sleep; mode of waking up; and comparison between the sleep recorded in the diary with the usual sleep. Personal and professional information was also collected. Day shift workers went to bed at 23h36min, and night workers at 23h52min (P > 0.05). The nurses working a day schedule woke up earlier (7h3min) than those working a night schedule when they slept at night (8h30min) (P

Protein denaturation improves enzymatic digestion efficiency for direct tissue analysis using mass spectrometry

NASA Astrophysics Data System (ADS)

Setou, M.; Hayasaka, T.; Shimma, S.; Sugiura, Y.; Matsumoto, M.

2008-12-01

Molecular identification using high-sensitivity tandem mass spectrometry is essential for protein analysis on the tissue surface. Here we report an improved digestion protocol for protein identification directly on the tissue surface using mass spectrometry. By denaturation process and the use of detergent-supplemented trypsin solution, we could successfully detect and identify many molecules such as tubulin, neurofilament, and synaptosomal-associated 25 kDa protein directly from a mouse cerebellum section.

The antigens contributing to the serological cross-reactions of Proteus antisera with Klebsiella representatives.

PubMed

Palusiak, Agata

2015-03-01

Proteus sp. and Klebsiella sp. mainly cause infections of the urinary and respiratory tracts or wounds in humans. The representatives of both genera produce virulence factors like lipopolysaccharide (LPS) or outer membrane proteins (OMPs) having much in common in the structures and/or functions. To check how far this similarity is revealed in the serological cross-reactivity, the bacterial masses of 24 tested Klebsiella sp. strains were tested in ELISA with polyclonal rabbit antisera specific to the representatives of 79 Proteus O serogroups. The strongest reacting systems were selected to Western blot, where the majority of Klebsiella masses reacted in a way characteristic for electrophoretic patterns of proteins. The strongest reactions were obtained for proteins of near 67 and 40 kDa and 12.5 kDa. Mass spectrometry analysis of the proteins samples of one Proteus sp. and one Klebsiella sp. strain showed the GroEL like protein of a sequence GI number 2980926 to be similar for both strains. In Western blot some Klebsiella sp. masses reacted similarly to the homologous Proteus LPSs. The LPS contribution in the observed reactions of the high molecular-mass LPS species was confirmed for Klebsiella oxytoca 0.062. Copyright © 2014 Elsevier Ltd. All rights reserved.

Mapping the primary structure of copper/topaquinone-containing methylamine oxidase from Aspergillus niger.

PubMed

Lenobel, R; Sebela, M; Frébort, I

2005-01-01

The amino acid sequence of methylamine oxidase (MeAO) from the fungus Aspergillus niger was analyzed using mass spectrometry (MS). First, MeAO was characterized by an accurate molar mass of 72.4 kDa of the monomer measured using MALDI-TOF-MS and by a pI value of 5.8 determined by isoelectric focusing. MALDI-TOF-MS revealed a clear peptide mass fingerprint after tryptic digestion, which did not provide any relevant hit when searched against a nonredundant protein database and was different from that of A. niger amine oxidase AO-I. Tandem mass spectrometry with electrospray ionization coupled to liquid chromatography allowed unambiguous reading of six peptide sequences (11-19 amino acids) and seven sequence tags (4-15 amino acids), which were used for MS BLAST homology searching. MeAO was found to be largely homologous to a hypothetical protein AN7641.2 (EMBL/GenBank protein-accession code EAA61827) from Aspergillus nidulans FGSC A4 with a theoretical molar mass of 76.46 kDa and pI 6.14, which belongs to the superfamily of copper amine oxidases. The protein AN7641.2 is only little homologous to the amine oxidase AO-I (32% identity, 49 % similarity).

Using He I λ10830 to Diagnose Mass Flows Around Herbig Ae/Be Stars

NASA Astrophysics Data System (ADS)

Cauley, Paul W.; Johns-Krull, Christopher M.

2015-01-01

The pre-main sequence Herbig Ae/Be stars (HAEBES) are the intermediate mass cousins of the low mass T Tauri stars (TTSs). However, it is not clear that the same accretion and mass outflow mechanisms operate identically in both mass regimes. Classical TTSs (CTTSs) accrete material from their disks along stellar magnetic field lines in a scenario called magnetospheric accretion. Magnetospheric accretion requires a strong stellar dipole field in order to truncate the inner gas disk. These fields are either absent or very weak on a large majority of HAEBES, challenging the view that magnetospheric accretion is the dominant accretion mechanism. If magnetospheric accretion does not operate similarly around HAEBES as it does around CTTSs, then strong magnetocentrifugal outflows, which are directly linked to accretion and are ubiquitous around CTTSs, may be driven less efficiently from HAEBE systems. Here we present high resolution spectroscopic observations of the He I λ10830 line in a sample of 48 HAEBES. He I λ10830 is an excellent tracer of both mass infall and outflow which is directly manifested as red and blue-shifted absorption in the profile morphologies. These features, among others, are common in our sample. The occurrence of both red and blue-shifted absorption profiles is less frequent, however, than is found in CTTSs. Statistical contingency tests confirm this difference at a significant level. In addition, we find strong evidence for smaller disk truncation radii in the objects displaying red-shifted absorption profiles. This is expected for HAEBES experiencing magnetospheric accretion based on their large rotation rates and weak magnetic field strengths. Finally, the low incidence of blue-shifted absorption in our sample compared to CTTSs and the complete lack of simultaneous red and blue-shifted absorption features suggests that magnetospheric accretion in HAEBES is less efficient at driving strong outflows. The stellar wind-like outflows that are observed are likely driven, at least in part, by boundary layer accretion. The smaller (or absent) disk truncation radii in HAEBES may have consequences for the frequency of planets in close orbits around main sequence B and A stars.

ASSESSMENT OF BODY FAT IN OBESE PATIENTS PREOPERATIVELY FOR BARIATRIC SURGERY.

PubMed

Fernandez, Mônica; Toimil, Rosana Farah; Rasslan, Zied; Ilias, Elias Jirjoss; Gradinar, Ana Lúcia Torloni; Malheiros, Carlos Alberto

The study of body composition in patient candidates for bariatric surgery is directly related to the increase and distribution of body fat in the development of cardiovascular disease. To correlate anthropometric indicators and bioelectrical impedance in the assessment of body fat in female candidates for bariatric surgery. Cross-sectional, observational study of 88 women. The weight, height, body mass index and waist circumference data were evaluated in the anthropometric analysis. The body fat was determinate by bioelectrical impedance conducted according to the manufacturer´s recommended technique with a specific severe obesity formula. The patients were divided into two subgroups according to the average waist circumference and body mass index for better analysis of the results. The group had a mean age of 39.7 years (±7.2), average weight of 125.6 kg (±16.2), mean body mass index of 48.7 kg/m2 (±6.4) and the mean waist circumference 137.6 cm (±12.4). Negative and significant relationship between BMI values waist circumference and resistance obtained by bioelectrical impedance ​​were found. By analyzing the two groups the mean BMI and waist circumference, a significant relationship was observed, ie, the higher the degree of obesity less resistance was obtained by bioelectrical impedance. The higher is the obesity the lower is value found for resistance. The increase of anthropometric indicators (BMI and waist circumference) determined reduction in resistance and reactance obtained by bioelectrical impedance analysis in obese women candidates to bariatric surgery. O estudo da composição corporal em pacientes candidatas à cirurgia bariátrica tem relação direta com o aumento e distribuição da gordura corporal e no desenvolvimento de doenças cardiovasculares. Estudar a correlação entre indicadores antropométricos e da bioimpedância elétrica na avaliação da gordura corporal em mulheres candidatas à cirurgia bariátrica. Estudo transversal, observacional com 88 pacientes. Na análise antropométrica foram avaliados os dados de peso, altura, IMC e circunferência da cintura. Para bioimpedância elétrica utilizou-se a técnica recomendada pelo fabricante do aparelho e fórmula específica para obeso grave na determinação da gordura corporal. Para melhor análise dos resultados as pacientes foram divididas em dois subgrupos de acordo com a média de circunferência da cintura e IMC. O grupo apresentou média de idade 39,7 anos (±7,2), média de peso 125,6 kg (±16,2), média de IMC 48,7 kg/m2 (±6,4) e média de circunferência da cintura 137,6 cm (±12,4). Foram encontradas relações negativas e significativas entre os valores de IMC, circunferência da cintura e a resistência obtida através da bioimpedância elétrica. Ao analisar os dois subgrupos de valores médios de IMC e circunferência da cintura, observou-se relação significativa, ou seja, quanto maior o grau de obesidade menor é a resistência obtida através da bioimpedância elétrica. O aumento dos indicadores antropométricos (IMC e circunferência da cintura) determinaram redução nas variáveis resistência e reatância obtidas pela bioimpedância elétrica em mulheres candidatas à cirurgia bariátrica.

Night-shift work is associated with poorer glycaemic control in patients with type 2 diabetes.

PubMed

Manodpitipong, Areesa; Saetung, Sunee; Nimitphong, Hataikarn; Siwasaranond, Nantaporn; Wongphan, Thanawat; Sornsiriwong, Chotima; Luckanajantachote, Pranee; Mangjit, Prasitchai; Keesukphan, Prasit; Crowley, Stephanie J; Hood, Megan M; Reutrakul, Sirimon

2017-12-01

The circadian system plays a role in regulating metabolism. Night-shift work, a form of circadian misalignment, is associated with increased type 2 diabetes risk. This study aimed to determine if night-shift workers with type 2 diabetes experience poorer glycaemic control than non-shift workers. Patients with type 2 diabetes (104 unemployed, 85 day workers and 60 night-shift workers) participated. Sleep duration, sleep quality, morningness-eveningness preference, depressive symptoms and dietary intake were assessed using standardized questionnaires. Haemoglobin A1c levels were measured. Night-shift workers had significantly higher haemoglobin A1c levels compared with others, while there were no differences between day workers and unemployed participants (median 7.86% versus 7.24% versus 7.09%, respectively). Additionally, night-shift workers were younger, had a higher body mass index, and consumed more daily calories than others. Among night-shift workers, there were no significant differences in haemoglobin A1c levels between those performing rotating versus non-rotating shifts (P = 0.856), or those with clockwise versus counterclockwise shift rotation (P = 0.833). After adjusting for age, body mass index, insulin use, sleep duration, morningness-eveningness preference and percentage of daily intake from carbohydrates, night-shift work, compared with day work, was associated with significantly higher haemoglobin A1c (B = 0.059, P = 0.044), while there were no differences between unemployed participants and day workers (B = 0.016, P = 0.572). In summary, night-shift work is associated with poorer glycaemic control in patients with type 2 diabetes. © The Authors. Journal of Sleep Research published by John Wiley & Sons Ltd on behalf of European Sleep Research Society.

How changing the particle structure can speed up protein mass transfer kinetics in liquid chromatography.

PubMed

Gritti, Fabrice; Horvath, Krisztian; Guiochon, Georges

2012-11-09

The mass transfer kinetics of a few compounds (uracil, 112 Da), insulin (5.5 kDa), lysozyme (13.4 kDa), and bovine serum albumin (BSA, 67 kDa) in columns packed with several types of spherical particles was investigated under non-retained conditions, in order to eliminate the poorly known contribution of surface diffusion to overall sample diffusivity across the porous particles in RPLC. Diffusivity across particles is then minimum. Based on the porosity of the particles accessible to analytes, it was accurately estimated from the elution times, the internal obstruction factor (using Pismen correlation), and the hindrance diffusion factor (using Renkin correlation). The columns used were packed with fully porous particles 2.5 μm Luna-C(18) 100 Å, core-shell particles 2.6 μm Kinetex-C(18) 100 Å, 3.6 μm Aeris Widepore-C(18) 200 Å, and prototype 2.7 μm core-shell particles (made of two concentric porous shells with 100 and 300 Å average pore size, respectively), and with 3.3 μm non-porous silica particles. The results demonstrate that the porous particle structure and the solid-liquid mass transfer resistance have practically no effect on the column efficiency for small molecules. For them, the column performance depends principally on eddy dispersion (packing homogeneity), to a lesser degree on longitudinal diffusion (effective sample diffusivity along the packed bed), and only slightly on the solid-liquid mass transfer resistance (sample diffusivity across the particle). In contrast, for proteins, this third HETP contribution, hence the porous particle structure, together with eddy dispersion govern the kinetic performance of columns. Mass transfer kinetics of proteins was observed to be fastest for columns packed with core-shell particles having either a large core-to-particle ratio or having a second, external, shell made of a thin porous layer with large mesopores (200-300 Å) and a high porosity (~/=0.5-0.7). The structure of this external shell seems to speed up the penetration of proteins into the particles. A stochastic model of the penetration of bulky proteins driven by a concentration gradient across an infinitely thin membrane of known porosity and pore size is suggested to explain this mechanism. Yet, under retained conditions, surface diffusion speeds up the mass transfer into the mesopores and levels the kinetic performance of particles built with either one or two porous shells. Copyright © 2012 Elsevier B.V. All rights reserved.

Resummation of Goldstone infrared divergences: A proof to all orders

NASA Astrophysics Data System (ADS)

Espinosa, J. R.; Konstandin, T.

2018-03-01

The perturbative effective potential calculated in Landau gauge suffers from infrared problems due to Goldstone boson loops. These divergences are spurious and can be removed by a resummation procedure that amounts to a shift of the mass of soft Goldstones. We prove this to all loops using an effective theory approach, providing a compact recipe for the shift of the Goldstone mass that relies on the use of the method of regions to split soft and hard Goldstone contributions.

Identification of ubiquitin/ubiquitin-like protein modification from tandem mass spectra with various PTMs

PubMed Central

2011-01-01

Background Various solutions have been introduced for the identification of post-translational modification (PTM) from tandem mass spectrometry (MS/MS) in proteomics field but the identification of peptide modifiers, such as Ubiquitin (Ub) and ubiquitin-like proteins (Ubls), is still a challenge. The fragmentation of peptide modifier produce complex shifted ion mass patterns in combination with other PTMs, which makes it difficult to identify and locate the PTMs on a protein sequence. Currently, most PTM identification methods do not consider the complex fragmentation of peptide modifier or deals it separately from the other PTMs. Results We developed an advanced PTM identification method that inspects possible ion patterns of the most known peptide modifiers as well as other known biological and chemical PTMs to make more comprehensive and accurate conclusion. The proposed method searches all detectable mass differences of measured peaks from their theoretical values and the mass differences within mass tolerance range are grouped as mass shift classes. The most possible locations of multiple PTMs including peptide modifiers can be determined by evaluating all possible scenarios generated by the combination of the qualified mass shift classes.The proposed method showed excellent performance in the test with simulated spectra having various PTMs including peptide modifiers and in the comparison with recently developed methods such as QuickMod and SUMmOn. In the analysis of HUPO Brain Proteome Project (BPP) datasets, the proposed method could find the ubiquitin modification sites that were not identified by other conventional methods. Conclusions This work presents a novel method for identifying bothpeptide modifiers that generate complex fragmentation patternsand PTMs that are not fragmented during fragmentation processfrom tandem mass spectra. PMID:22373085

Petroleomics: the next grand challenge for chemical analysis.

PubMed

Marshall, Alan G; Rodgers, Ryan P

2004-01-01

Ultrahigh-resolution Fourier transform ion cyclotron resonance mass spectrometry has recently revealed that petroleum crude oil contains heteroatom-containing (N,O,S) organic components having more than 20,000 distinct elemental compositions (C(c)H(h)N(n)O(o)S(s)). It is therefore now possible to contemplate the ultimate characterization of all of the chemical constituents of petroleum, along with their interactions and reactivity, a concept we denote as "petroleomics". Such knowledge has already proved capable of distinguishing petroleum and its distillates according to their geochemical origin and maturity, distillation cut, extraction method, catalytic processing, etc. The key features that have opened up this new field have been (a) ultrahigh-resolution FT-ICR mass analysis, specifically, the capability to resolve species differing in elemental composition by C(3) vs SH(4) (i.e., 0.0034 Da); (b) higher magnetic field to cover the whole mass range at once; (c) dynamic range extension by external mass filtering; and (d) plots of Kendrick mass defect vs nominal Kendrick mass as a means for sorting different compound "classes" (i.e., numbers of N, O, and S atoms), "types" (rings plus double bonds), and alkylation ((-CH(2))(n)) distributions, thereby extending to >900 Da the upper limit for unique assignment of elemental composition based on accurate mass measurement. The same methods are also being applied successfully to analysis of humic and fulvic acids, coals, and other complex natural mixtures, often without prior or on-line chromatographic separation.

Prolonged Permian Triassic ecological crisis recorded by molluscan dominance in Late Permian offshore assemblages.

PubMed

Clapham, Matthew E; Bottjer, David J

2007-08-07

The end-Permian mass extinction was the largest biotic crisis in the history of animal life, eliminating as many as 95% of all species and dramatically altering the ecological structure of marine communities. Although the causes of this pronounced ecosystem shift have been widely debated, the broad consensus based on inferences from global taxonomic diversity patterns suggests that the shift from abundant brachiopods to dominant molluscs was abrupt and largely driven by the catastrophic effects of the end-Permian mass extinction. Here we analyze relative abundance counts of >33,000 fossil individuals from 24 silicified Middle and Late Permian paleocommunities, documenting a substantial ecological shift to numerical dominance by molluscs in the Late Permian, before the major taxonomic shift at the end-Permian mass extinction. This ecological change was coincident with the development of fluctuating anoxic conditions in deep marine basins, suggesting that numerical dominance by more tolerant molluscs may have been driven by variably stressful environmental conditions. Recognition of substantial ecological deterioration in the Late Permian also implies that the end-Permian extinction was the climax of a protracted environmental crisis. Although the Late Permian shift to molluscan dominance was a pronounced ecological change, quantitative counts of 847 Carboniferous-Cretaceous collections from the Paleobiology Database indicate that it was only the first stage in a stepwise transition that culminated with the final shift to molluscan dominance in the Late Jurassic. Therefore, the ecological transition from brachiopods to bivalves was more protracted and complex than their simple Permian-Triassic switch in diversity.

Prolonged Permian–Triassic ecological crisis recorded by molluscan dominance in Late Permian offshore assemblages

PubMed Central

Clapham, Matthew E.; Bottjer, David J.

2007-01-01

The end-Permian mass extinction was the largest biotic crisis in the history of animal life, eliminating as many as 95% of all species and dramatically altering the ecological structure of marine communities. Although the causes of this pronounced ecosystem shift have been widely debated, the broad consensus based on inferences from global taxonomic diversity patterns suggests that the shift from abundant brachiopods to dominant molluscs was abrupt and largely driven by the catastrophic effects of the end-Permian mass extinction. Here we analyze relative abundance counts of >33,000 fossil individuals from 24 silicified Middle and Late Permian paleocommunities, documenting a substantial ecological shift to numerical dominance by molluscs in the Late Permian, before the major taxonomic shift at the end-Permian mass extinction. This ecological change was coincident with the development of fluctuating anoxic conditions in deep marine basins, suggesting that numerical dominance by more tolerant molluscs may have been driven by variably stressful environmental conditions. Recognition of substantial ecological deterioration in the Late Permian also implies that the end-Permian extinction was the climax of a protracted environmental crisis. Although the Late Permian shift to molluscan dominance was a pronounced ecological change, quantitative counts of 847 Carboniferous–Cretaceous collections from the Paleobiology Database indicate that it was only the first stage in a stepwise transition that culminated with the final shift to molluscan dominance in the Late Jurassic. Therefore, the ecological transition from brachiopods to bivalves was more protracted and complex than their simple Permian–Triassic switch in diversity. PMID:17664426

Mechanisms of oxygen sensing: a key to therapy of pulmonary hypertension and patent ductus arteriosus

PubMed Central

Weir, E K; Obreztchikova, M; Vargese, A; Cabrera, J A; Peterson, D A; Hong, Z

2008-01-01

Specialized tissues that sense acute changes in the local oxygen tension include type 1 cells of the carotid body, neuroepithelial bodies in the lungs, and smooth muscle cells of the resistance pulmonary arteries and the ductus arteriosus (DA). Hypoxia inhibits outward potassium current in carotid body type 1 cells, leading to depolarization and calcium entry through L-type calcium channels. Increased intracellular calcium concentration ([Ca++]i) leads to exocytosis of neurotransmitters, thus stimulating the carotid sinus nerve and respiration. The same K+ channel inhibition occurs with hypoxia in pulmonary artery smooth muscle cells (PASMCs), causing contraction and providing part of the mechanism of hypoxic pulmonary vasoconstriction (HPV). In the SMCs of the DA, the mechanism works in reverse. It is the shift from hypoxia to normoxia that inhibits K+ channels and causes normoxic ductal contraction. In both PA and DA, the contraction is augmented by release of Ca++ from the sarcoplasmic reticulum, entry of Ca++ through store-operated channels (SOC) and by Ca++ sensitization. The same three ‘executive' mechanisms are partly responsible for idiopathic pulmonary arterial hypertension (IPAH). While vasoconstrictor mediators constrict both PA and DA and vasodilators dilate both vessels, only redox changes mimic oxygen by having directly opposite effects on the K+ channels, membrane potential, [Ca++]i and tone in the PA and DA. There are several different hypotheses as to how redox might alter tone, which remain to be resolved. However, understanding the mechanism will facilitate drug development for pulmonary hypertension and patent DA. PMID:18641675

Partial dopaminergic denervation-induced impairment in stimulus discrimination acquisition in parkinsonian rats: a model for early Parkinson's disease.

PubMed

Eagle, Andrew L; Olumolade, Oluyemi O; Otani, Hajime

2015-03-01

Parkinson's disease (PD) produces progressive nigrostriatal dopamine (DA) denervation resulting in cognitive and motor impairment. However, it is unknown whether cognitive impairments, such as instrumental learning deficits, are associated with the early stage PD-induced mild DA denervation. The current study sought to model early PD-induced instrumental learning impairments by assessing the effects of low dose (5.5μg), bilateral 6OHDA-induced striatal DA denervation on acquisition of instrumental stimulus discrimination in rats. 6OHDA (n=20) or sham (n=10) lesioned rats were tested for stimulus discrimination acquisition either 1 or 2 weeks post surgical lesion. Stimulus discrimination acquisition across 10 daily sessions was used to assess discriminative accuracy, or a probability measure of the shift toward reinforced responding under one stimulus condition (Sd) away from extinction, when reinforcement was withheld, under another (S(d) phase). Striatal DA denervation was assayed by tyrosine hydroxylase (TH) staining intensity. Results indicated that 6OHDA lesions produced significant loss of dorsal striatal TH staining intensity and marked impairment in discrimination acquisition, without inducing akinetic motor deficits. Rather 6OHDA-induced impairment was associated with perseveration during extinction (S(Δ) phase). These findings suggest that partial, bilateral striatal DA denervation produces instrumental learning deficits, prior to the onset of gross motor impairment, and suggest that the current model is useful for investigating mild nigrostriatal DA denervation associated with early stage clinical PD. Copyright © 2014 Elsevier Ireland Ltd and the Japan Neuroscience Society. All rights reserved.

Didehydrophenylalanine, an abundant modification in the beta subunit of plant polygalacturonases.

PubMed

Sergeant, Kjell; Printz, Bruno; Gutsch, Annelie; Behr, Marc; Renaut, Jenny; Hausman, Jean-Francois

2017-01-01

The structure and the activity of proteins are often regulated by transient or stable post- translational modifications (PTM). Different from well-known, abundant modifications such as phosphorylation and glycosylation some modifications are limited to one or a few proteins across a broad range of related species. Although few examples of the latter type are known, the evolutionary conservation of these modifications and the enzymes responsible for their synthesis suggest an important physiological role. Here, the first observation of a new, fold-directing PTM is described. During the analysis of alfalfa cell wall proteins a -2Da mass shift was observed on phenylalanine residues in the repeated tetrapeptide FxxY of the beta-subunit of polygalacturonase. This modular protein is known to be involved in developmental and stress-responsive processes. The presence of this modification was confirmed using in-house and external datasets acquired by different commonly used techniques in proteome studies. Based on these analyses it was found that all identified phenylalanine residues in the sequence FxxY of this protein were modified to α,β-didehydro-Phe (ΔPhe). Besides showing the reproducible identification of ΔPhe in different species arguments that substantiate the fold-determining role of ΔPhe are given.

Cloning, functional expression, and characterization of a PKA-activated gastric Cl- channel.

PubMed

Malinowska, D H; Kupert, E Y; Bahinski, A; Sherry, A M; Cuppoletti, J

1995-01-01

cDNA encoding a Cl- channel was isolated from a rabbit gastric library, sequenced, and expressed in Xenopus oocytes. The predicted protein (898 amino acids, relative molecular mass 98,433 Da) was overall 93% similar to the rat brain ClC-2 Cl- channel. However, a 151-amino acid stretch toward the COOH-terminus was 74% similar to ClC-2 with six amino acids deleted. Two new potential protein kinase A (PKA) phosphorylation sites (also protein kinase C phosphorylation sites) were introduced. cRNA-injected Xenopus oocytes expressed a Cl- channel that was active at pHtrans 3 and had a linear current-voltage (I-V) curve and a slope conductance of 29 +/- 1 pS at 800 mM CsCl. A fivefold Cl- gradient caused a rightward shift in the I-V curve with a reversal potential of +30 +/- 3 mV, indicating anion selectivity. The selectivity was I- > Cl- > NO3-. The native and recombinant Cl- channel were both activated in vitro by PKA catalytic subunit and ATP. The electrophysiological and regulatory properties of the cloned and the native channel were similar. The cloned protein may be the Cl- channel involved in gastric HCl secretion.

Z-DNA binding protein from chicken blood nuclei

NASA Technical Reports Server (NTRS)

Herbert, A. G.; Spitzner, J. R.; Lowenhaupt, K.; Rich, A.

1993-01-01

A protein (Z alpha) that appears to be highly specific for the left-handed Z-DNA conformer has been identified in chicken blood nuclear extracts. Z alpha activity is measured in a band-shift assay by using a radioactive probe consisting of a (dC-dG)35 oligomer that has 50% of the deoxycytosines replaced with 5-bromodeoxycytosine. In the presence of 10 mM Mg2+, the probe converts to the Z-DNA conformation and is bound by Z alpha. The binding of Z alpha to the radioactive probe is specifically blocked by competition with linear poly(dC-dG) stabilized in the Z-DNA form by chemical bromination but not by B-form poly(dC-dG) or boiled salmon-sperm DNA. In addition, the binding activity of Z alpha is competitively blocked by supercoiled plasmids containing a Z-DNA insert but not by either the linearized plasmid or by an equivalent amount of the parental supercoiled plasmid without the Z-DNA-forming insert. Z alpha can be crosslinked to the 32P-labeled brominated probe with UV light, allowing us to estimate that the minimal molecular mass of Z alpha is 39 kDa.

Oxygenation decreases elastin secretion from rat ductus arteriosus smooth muscle cells.

PubMed

Kawakami, Shoji; Minamisawa, Susumu

2015-08-01

The ductus arteriosus (DA), a fetal arterial connection between the main pulmonary artery and the descending aorta, normally closes immediately after birth. The oxygen concentration in the blood rises after birth, and in the DA this increase in oxygen concentration causes functional closure, which is induced by smooth muscle contraction. Previous studies have demonstrated that hypoxia and/or oxygenation affect vascular remodeling of various vessels. Therefore, we hypothesized that the rise in oxygen concentration would affect the vascular structure of the DA due to production of proteins secreted from DA smooth muscle cells (SMC). Liquid chromatography-tandem mass spectrometry was used to comprehensively investigate the secreted proteins in the supernatant of rat DA SMC harvested under hypoxic conditions (1% oxygen) or under normoxic conditions (21% oxygen). We found that the rise in oxygen concentration reduced the secretion of elastin from DA SMC. On reverse transcription-polymerase chain reaction, the expression of elastin mRNA was not significantly changed in DA SMC from hypoxic to normoxic conditions. Given that elastin forms internal elastic lamina and elastic fibers in the vascular muscle layers, and that a rise in oxygen concentration reduced the secretion of elastin, this suggests that the rise in blood oxygen concentration after birth reduces the secretion of elastin, and therefore may play a role in DA structural remodeling after birth. © 2015 Japan Pediatric Society.

Characterization of a major 31-kilodalton peptidoglycan-bound protein of Legionella pneumophila

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

Butler, C.A.; Hoffman, P.S.

1990-05-01

A 31-kilodalton (kDa) protein was solubilized from the peptidoglycan (PG) fraction of Legionella pneumophila after treatment with either N-acetylmuramidase from the fungus Chalaropsis sp. or with mutanolysin from Streptomyces globisporus. The protein exhibited a ladderlike banding pattern by autoradiography when radiolabeled ((35S)cysteine or (35S)methionine) PG material was extensively treated with hen lysozyme. The banding patterns ranging between 31 and 45 kDa and between 55 and 60 kDa resolved as a single 31-kDa protein when the material was subsequently treated with N-acetylmuramidase. Analysis of the purified 31-kDa protein for diaminopimelic acid by gas chromatography revealed 1 mol of diaminopimelic acid permore » mol of protein. When outer membrane PG material containing the major outer membrane porin protein was treated with N-acetylmuramidase or mutanolysin, both the 28.5-kDa major outer membrane protein and the 31-kDa protein were solubilized from the PG material under reducing conditions. In the absence of 2-mercaptoethanol, a high-molecular-mass complex (100 kDa) was resolved. The results of this study indicate that a 31-kDa PG-bound protein is a major component of the cell wall of L. pneumophila whose function may be to anchor the major outer membrane protein to PG. Finally, a survey of other Legionella species and other serogroups of L. pneumophila suggested that PG-bound proteins may be a common feature of this genus.« less

The molar hydrodynamic volume changes of factor VIIa due to GlycoPEGylation.

PubMed

Plesner, Bitten; Westh, Peter; Hvidt, Søren; Nielsen, Anders D

2011-06-01

The effects of GlycoPEGylation on the molar hydrodynamic volume of recombinant human rFVIIa were investigated using rFVIIa and two GlycoPEGylated recombinant human FVIIa derivatives, a linear 10kDa PEG and a branched 40kDa PEG, respectively. Molar hydrodynamic volumes were determined by capillary viscometry and mass spectrometry. The intrinsic viscosities of rFVIIa, its two GlycoPEGylated compounds, and of linear 8kDa, 10kDa, 20kDa and branched 40kDa PEG polymers were determined. The measured intrinsic viscosity of rFVIIa is 6.0mL/g, while the intrinsic viscosities of 10kDa PEG-rFVIIa and 40kDa PEG-rFVIIa are 29.5mL/g and 79.0mL/g, respectively. The intrinsic viscosities of the linear PEG polymers are 20, 22.6 and 41.4mL/g for 8, 10, and 20kDa, respectively, and 61.1mL/g for the branched 40kDa PEG. From the results of the intrinsic viscosity and MALDI-TOF measurements it is evident, that the molar hydrodynamic volume of the conjugated protein is not just an addition of the molar hydrodynamic volume of the PEG and the protein. The molar hydrodynamic volume of the GlycoPEGylated protein is larger than the volume of its composites. These results suggest that both the linear and the branched PEG are not wrapped around the surface of rFVIIa but are chains that are significantly stretched out when attached to the protein. Copyright © 2011 Elsevier B.V. All rights reserved.

Morphing Downwind-Aligned Rotor Concept Based on a 13-MW Wind Turbine

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

Ichter, Brian; Steele, Adam; Loth, Eric

To alleviate the mass-scaling issues associated with conventional upwind rotors of extreme-scale wind turbines (>/=10 MW), a morphing downwind-aligned rotor (MoDaR) concept is proposed herein. The concept employs a downwind rotor with blades whose elements are stiff (no intentional flexibility) but with hub-joints that can be unlocked to allow for moment-free downwind alignment. Aligning the combination of gravitational, centrifugal and thrust forces along the blade path reduces downwind cantilever loads, resulting in primarily tensile loading. For control simplicity, the blade curvature can be fixed with a single morphing degree of freedom using a near-hub joint for coning angle: 22 degreesmore » at rated conditions. The conventional baseline was set as the 13.2-MW Sandia 100-m all glass blade in a three-bladed upwind configuration. To quantify potential mass savings, a downwind load-aligning, two-bladed rotor was designed. Because of the reduced number of blades, the MoDaR concept had a favorable 33% mass reduction. The blade reduction and coning led to a reduction in rated power, but morphing increased energy capture at lower speeds such that both the MoDaR and conventional rotors have the same average power: 5.4 MW. A finite element analysis showed that quasi-steady structural stresses could be reduced, over a range of operating wind speeds and azimuthal angles, despite the increases in loading per blade. However, the concept feasibility requires additional investigation of the mass, cost and complexity of the morphing hinge, the impact of unsteady aeroelastic influence because of turbulence and off-design conditions, along with system-level Levelized Cost of Energy analysis.« less

Evaluation of the LTQ-Orbitrap mass spectrometer for the analysis of polymerase chain reaction products.

PubMed

Manduzio, Hélène; Ezan, Eric; Fenaille, François

2010-12-30

We have investigated the potential and robustness of the off-line coupling of polymerase chain reaction (PCR) with electrospray ionization mass spectrometry (ESI-MS), for further applications in the screening of single-nucleotide polymorphisms (SNPs). This was based on recently reported data demonstrating that anion-exchange solid-phase extraction was the most efficient technique for efficiently desalting PCR products, with a recovery of ∼70%. Results showed that this purification approach efficiently removes almost all the chemicals commonly added to PCR buffers. ESI-MS analysis of a model 114-bp PCR product performed on the LTQ-Orbitrap instrument demonstrated that detection limits in the nM range along with an average mass measurement uncertainty of 9.15 ± 7.11 ppm can be routinely obtained using an external calibration. The PCR/ESI-MS platform was able to detect just a few copies of a targeted oligonucleotide. However, it was shown that if two PCR products are present in a mixture in a ratio higher than 10 to 1, the lower abundance one might not be reproducibly detected. Applications to SNPs demonstrated that an LTQ-Orbitrap with a resolution of 30 000 (at m/z 400) easily identified a single (A ↔ G) switch, i.e. a 16 Da difference, in binary mixtures of ∼ 35 kDa PCR products. Complementary experiments also showed that the combination of endonucleases and ESI-MS could be used to confirm base composition and sequence, and thus to screen for unknown polymorphisms in specific sequences. For example, a single (T ↔ A) switch (9 Da mass difference) was successfully identified in a 114-bp PCR product. Copyright © 2010 John Wiley & Sons, Ltd.

Mass spectrometric characterization of tamoxifene metabolites in human urine utilizing different scan parameters on liquid chromatography/tandem mass spectrometry.

PubMed

Mazzarino, Monica; de la Torre, Xavier; Di Santo, Roberto; Fiacco, Ilaria; Rosi, Federica; Botrè, Francesco

2010-03-01

Different liquid chromatographic/tandem mass spectrometric (LC/MS/MS) scanning techniques were considered for the characterization of tamoxifene metabolites in human urine for anti-doping purpose. Five different LC/MS/MS scanning methods based on precursor ion scan (precursor ion scan of m/z 166, 152 and 129) and neutral loss scan (neutral loss of 72 Da and 58 Da) in positive ion mode were assessed to recognize common ions or common losses of tamoxifene metabolites. The applicability of these methods was checked first by infusion and then by the injection of solution of a mixture of reference standards of four tamoxifene metabolites available in our laboratory. The data obtained by the analyses of the mixture of the reference standards showed that the five methods used exhibited satisfactory results for all tamoxifene metabolites considered at a concentration level of 100 ng/mL, whereas the analysis of blank urine samples spiked with the same tamoxifene metabolites at the same concentration showed that the neutral loss scan of 58 Da lacked sufficient specificity and sensitivity. The limit of detection in urine of the compounds studied was in the concentration range 10-100 ng/mL, depending on the compound structure and on the selected product ion. The suitability of these approaches was checked by the analysis of urine samples collected after the administration of a single dose of 20 mg of tamoxifene. Six metabolites were detected: 4-hydroxytamoxifene, 3,4-dihydroxytamoxifene, 3-hydroxy-4-methoxytamoxifene, N-demethyl-4-hydroxytamoxifene, tamoxifene-N-oxide and N-demethyl-3-hydroxy-4-methoxytamoxifene, which is in conformity to our previous work using a time-of-flight (TOF) mass spectrometer in full scan acquisition mode. Copyright (c) 2010 John Wiley & Sons, Ltd.

Rapid and High-Throughput pan-Orthopoxvirus Detection and Identification using PCR and Mass Spectrometry

DTIC Science & Technology

2009-07-01

used in this manuscript. Several of the co- authors are or were Ibis employees. * E -mail: meshoo@ibisbio.com Introduction The orthopoxviruses (family...its Eradication. Geneva: World Health Organization. 4. Henderson DA, Inglesby TV, Bartlett JG, Ascher MS, Eitzen E , et al. (1999) Smallpox as a...of Rio de Janeiro: preliminary report. Mem Inst Oswaldo Cruz 95: 625–627. 15. Trindade GS, da Fonseca FG, Marques JT, Diniz S, Leite JA, et al. (2004

Two-dimensional polyacrylamide gel electrophoresis of equine seminal plasma proteins and their relation with semen freezability.

PubMed

Jobim, M I M; Trein, C; Zirkler, H; Gregory, R M; Sieme, H; Mattos, R C

2011-09-01

The objective was to evaluate protein profiles of equine seminal plasma using two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and to determine whether any of these proteins were related to semen freezability. Seminal plasma was collected from 10 stallions, of high and low semen freezability, housed at the State Stud of Lower Saxony, and routinely used in AI programs. Twenty-five protein spots were identified from the two-dimensional gel (12%), seven of which were present in all samples (all proteins were identified by MALDI-MS). Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) has been used to generate ion images of samples in one or more mass-to-charge (m/z) values, providing the capability of mapping specific molecules to two-dimensional coordinates of the original sample. Of the 25 proteins identified, two spots had greater relative content (P < 0.05) in seminal plasma samples collected from stallions with high semen freezability: spot 5 (80-85 kDa, isoelectric point [pI] 7.54), identified as CRISP-3; and spot 45 (18.2 kDa, pI 5.0-5.2), identified as HSP-2. Conversely, protein content was greater (P < 0.05) in seminal plasma samples from stallions with low semen freezability: spot 7 (75.4 kDa, pI 6.9-7.4), identified as lactoferrin; spot 15 (26.7 kDa, pI 5.51), identified as kallikrein; spot 25 (25 kDa, pI 7.54), identified as CRISP-3; and spot 35 (13.9 kDa, pI 3.8-4.2), identified as HSP-1. In conclusion, there were differences in the seminal plasma protein profile from stallions with high and low semen freezability. Furthermore, CRISP-3 and HSP-2 were potential seminal plasma markers of high semen freezability. Copyright © 2011 Elsevier Inc. All rights reserved.

Ovulation-inducing factor: a protein component of llama seminal plasma

PubMed Central

2010-01-01

Background Previously, we documented the presence of ovulation-inducing factor (OIF) in the seminal plasma of llamas and alpacas. The purpose of the study was to define the biochemical characteristics of the molecule(s) in seminal plasma responsible for inducing ovulation. Methods In Experiment 1, llama seminal plasma was centrifuged using filtration devices with nominal molecular mass cut-offs of 30, 10 and 5 kDa. Female llamas (n = 9 per group) were treated i.m. with whole seminal plasma (positive control), phosphate-buffered saline (negative control), or the fraction of seminal plasma equal or higher than 30 kDa, 10 to 30 kDa, 5 to 10 kDa, or < 5 kDa. In Experiment 2, female llamas (n = 7 per group) were given an i.m. dose of seminal plasma treated previously by: 1) enzymatic digestion with proteinase-K, 2) incubation with charcoal-dextran, 3) heating to 65°C, or 4) untreated (control). In Experiment 3, female llamas (n = 10 per group) were given an i.m. dose of pronase-treated or non-treated (control) seminal plasma. In all experiments, llamas were examined by transrectal ultrasonography to detect ovulation and CL formation. Ovulation rate was compared among groups by Fisher's exact test and follicle and CL diameters were compared among groups by analyses of variance or student's t-tests. Results In Experiment 1, all llamas in the equal or higher than 30 kDa and positive control groups ovulated (9/9 in each), but none ovulated in the other groups (P < 0.001). In Experiment 2, ovulations were detected in all llamas in each treatment group; i.e., respective treatments of seminal plasma failed to inactivate the ovulation-inducing factor. In Experiment 3, ovulations were detected in 0/10 llamas given pronase-treated seminal plasma and in 9/10 controls (P < 0.01). Conclusions We conclude that ovulation-inducing factor (OIF) in llama seminal plasma is a protein molecule that is resistant to heat and enzymatic digestion with proteinase K, and has a molecular mass of approximately equal or higher than 30 kDa. PMID:20462434

Deep proton tunneling in the electronically adiabatic and non-adiabatic limits: Comparison of the quantum and classical treatment of donor-acceptor motion in a protein environment

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

Benabbas, Abdelkrim; Salna, Bridget; Sage, J. Timothy

2015-03-21

Analytical models describing the temperature dependence of the deep tunneling rate, useful for proton, hydrogen, or hydride transfer in proteins, are developed and compared. Electronically adiabatic and non-adiabatic expressions are presented where the donor-acceptor (D-A) motion is treated either as a quantized vibration or as a classical “gating” distribution. We stress the importance of fitting experimental data on an absolute scale in the electronically adiabatic limit, which normally applies to these reactions, and find that vibrationally enhanced deep tunneling takes place on sub-ns timescales at room temperature for typical H-bonding distances. As noted previously, a small room temperature kinetic isotopemore » effect (KIE) does not eliminate deep tunneling as a major transport channel. The quantum approach focuses on the vibrational sub-space composed of the D-A and hydrogen atom motions, where hydrogen bonding and protein restoring forces quantize the D-A vibration. A Duschinsky rotation is mandated between the normal modes of the reactant and product states and the rotation angle depends on the tunneling particle mass. This tunnel-mass dependent rotation contributes substantially to the KIE and its temperature dependence. The effect of the Duschinsky rotation is solved exactly to find the rate in the electronically non-adiabatic limit and compared to the Born-Oppenheimer (B-O) approximation approach. The B-O approximation is employed to find the rate in the electronically adiabatic limit, where we explore both harmonic and quartic double-well potentials for the hydrogen atom bound states. Both the electronically adiabatic and non-adiabatic rates are found to diverge at high temperature unless the proton coupling includes the often neglected quadratic term in the D-A displacement from equilibrium. A new expression is presented for the electronically adiabatic tunnel rate in the classical limit for D-A motion that should be useful to experimentalists working near room temperature. This expression also holds when a broad protein conformational distribution of D-A equilibrium distances dominates the spread of the D-A vibrational wavefunction.« less

Characterization of hallucinogenic phenethylamines using high-resolution mass spectrometry for non-targeted screening purposes.

PubMed

Pasin, Daniel; Cawley, Adam; Bidny, Sergei; Fu, Shanlin

2017-10-01

Hallucinogenic phenethylamines such as 2,5-dimethoxyphenethylamines (2C-X) and their N-(2-methoxybenzyl) derivatives (25X-NBOMe) have seen an increase in novel analogues in recent years. These rapidly changing analogues make it difficult for laboratories to rely on traditional targeted screening methods to detect unknown new psychoactive substances (NPS). In this study, twelve 2C-X, six 2,5-dimethoxyamphetamines (DOX), and fourteen 25X-NBOMe derivatives, including two deuterated derivatives (2C-B-d 6 and 25I-NBOMe-d 9 ), were analyzed using ultra-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS). Collision-induced dissociation (CID) experiments were performed using collision energies set at 10, 20, and 40 eV. For 2C-X and DOX derivatives, common losses were observed including neutral and radical losses such as NH 3 (17.0265 Da), •CH 6 N (32.0500 Da), C 2 H 7 N (45.0578 Da) and C 2 H 9 N (47.0735 Da). 2C-X derivatives displayed common product ions at m/z 164.0837 ([C 10 H 12 O 2 ] +• ), 149.0603 ([C 9 H 9 O 2 ] + ), and 134.0732 ([C 9 H 10 O] +• ) while DOX derivatives had common product ions at m/z 178.0994 ([C 11 H 14 O 2 ] +• ), 163.0754 ([C 10 H 11 O 2 ] + ), 147.0804 ([C 10 H 11 O] + ), and 135.0810 ([C 9 H 11 O] + ). 25X-NBOMe had characteristic product ions at m/z 121.0654 ([C 8 H 9 O] + ) and 91.0548 ([C 7 H 7 ] + ) with minor common losses corresponding to 2-methylanisole (C 8 H 10 O, 122.0732 Da), 2-methoxybenzylamine (C 8 H 11 NO, 137.0847 Da), and •C 9 H 14 NO (152.1074 Da). Novel analogues of the selected classes can be detected by applying neutral loss filters (NLFs) and extracting the common product ions. Copyright © 2017 John Wiley & Sons, Ltd. Copyright © 2017 John Wiley & Sons, Ltd.

Deep proton tunneling in the electronically adiabatic and non-adiabatic limits: comparison of the quantum and classical treatment of donor-acceptor motion in a protein environment.

PubMed

Benabbas, Abdelkrim; Salna, Bridget; Sage, J Timothy; Champion, Paul M

2015-03-21

Analytical models describing the temperature dependence of the deep tunneling rate, useful for proton, hydrogen, or hydride transfer in proteins, are developed and compared. Electronically adiabatic and non-adiabatic expressions are presented where the donor-acceptor (D-A) motion is treated either as a quantized vibration or as a classical "gating" distribution. We stress the importance of fitting experimental data on an absolute scale in the electronically adiabatic limit, which normally applies to these reactions, and find that vibrationally enhanced deep tunneling takes place on sub-ns timescales at room temperature for typical H-bonding distances. As noted previously, a small room temperature kinetic isotope effect (KIE) does not eliminate deep tunneling as a major transport channel. The quantum approach focuses on the vibrational sub-space composed of the D-A and hydrogen atom motions, where hydrogen bonding and protein restoring forces quantize the D-A vibration. A Duschinsky rotation is mandated between the normal modes of the reactant and product states and the rotation angle depends on the tunneling particle mass. This tunnel-mass dependent rotation contributes substantially to the KIE and its temperature dependence. The effect of the Duschinsky rotation is solved exactly to find the rate in the electronically non-adiabatic limit and compared to the Born-Oppenheimer (B-O) approximation approach. The B-O approximation is employed to find the rate in the electronically adiabatic limit, where we explore both harmonic and quartic double-well potentials for the hydrogen atom bound states. Both the electronically adiabatic and non-adiabatic rates are found to diverge at high temperature unless the proton coupling includes the often neglected quadratic term in the D-A displacement from equilibrium. A new expression is presented for the electronically adiabatic tunnel rate in the classical limit for D-A motion that should be useful to experimentalists working near room temperature. This expression also holds when a broad protein conformational distribution of D-A equilibrium distances dominates the spread of the D-A vibrational wavefunction.

A study on 2, 4, 6-trinitrotolurene (TNT) vapor detection by using a quartz crystal microbalance with 18-crown-6 ether film

NASA Astrophysics Data System (ADS)

Phetchakul, Toempong; Phuvanatai, Pavaris

2017-07-01

The application of 18-crown-6 ether film for 2, 4, 6-trinitrotolurene (TNT) vapor detection by using quartz crystal microbalance (QCM) is studied. The film is coated on the quartz electrodes as sensitive material for capture TNT molecule. The parameters that are studied are concentration and mass or thickness of film. When the explosive adheres to surface of the crystal oscillator, the weight is changed and the resonance frequency of the crystal oscillator is shifted lower. The frequency shift (Δf) relates to concentration and mass or thickness. The high concentration and mass/thickness of film enhance the TNT detection.

D mesons in a magnetic field

DOE PAGES

Gubler, Philipp; Hattori, Koichi; Lee, Su Houng; ...

2016-03-15

In this paper, we investigate the mass spectra of open heavy flavor mesons in an external constant magnetic field within QCD sum rules. Spectral Ansatze on the phenomenological side are proposed in order to properly take into account mixing effects between the pseudoscalar and vector channels, and the Landau levels of charged mesons. The operator product expansion is implemented up to dimension-5 operators. As a result, we find for neutral D mesons a significant positive mass shift that goes beyond simple mixing effects. In contrast, charged D mesons are further subject to Landau level effects, which together with the mixingmore » effects almost completely saturate the mass shifts obtained in our sum rule analysis.« less

Recent Earth oblateness variations: unraveling climate and postglacial rebound effects.

PubMed

Dickey, Jean O; Marcus, Steven L; de Viron, Olivier; Fukumori, Ichiro

2002-12-06

Earth's dynamic oblateness (J2) has been decreasing due to postglacial rebound (PGR). However, J2 began to increase in 1997, indicating a pronounced global-scale mass redistribution within Earth's system. We have determined that the observed increases in J2 are caused primarily by a recent surge in subpolar glacial melting and by mass shifts in the Southern, Pacific, and Indian oceans. When these effects are removed, the residual trend in J2 (-2.9 x 10(-11) year-1) becomes consistent with previous estimates of PGR from satellite and eclipse data. The climatic significance of these rapid shifts in glacial and oceanic mass, however, remains to be investigated.

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

Barrett, Christopher A.; Martinez, Alonzo; McNamara, Bruce K.

International Atom Energy Agency (IAEA) safeguard verification measures in gaseous centrifuge enrichment plants (GCEPs) rely on environmental sampling, non-destructive assay (NDA), and destructive assay (DA) sampling and analysis to determine uranium enrichment. UF6 bias defect measurements are made by DA sampling and analysis to assure that enrichment is consistent with declarations. DA samples are collected from a limited number of cylinders for high precision, offsite mass spectrometer analysis. Samples are typically drawn from a sampling tap into a UF6 sample bottle, then packaged, sealed, and shipped under IAEA chain of custody to an offsite analytical laboratory. Future DA safeguard measuresmore » may require improvements in efficiency and effectiveness as GCEP capacities increase and UF6 shipping regulations become increasingly more restrictive. The Pacific Northwest National Laboratory (PNNL) DA sampler concept and Laser Ablation Absorption Ratio Spectrometry (LAARS) assay method are under development to potentially provide DA safeguard tools that increase inspection effectiveness and reduce sample shipping constraints. The PNNL DA sampler concept uses a handheld sampler to collect DA samples for either onsite LAARS assay or offsite laboratory analysis. The DA sampler design will use a small sampling planchet that is coated with an adsorptive film to collect controlled quantities of UF6 gas directly from a cylinder or process sampling tap. Development efforts are currently underway at PNNL to enhance LAARS assay performance to allow high-precision onsite bias defect measurements. In this paper, we report on the experimental investigation to develop adsorptive films for the PNNL DA sampler concept. These films are intended to efficiently capture UF6 and then stabilize the collected DA sample prior to onsite LAARS or offsite laboratory analysis. Several porous material composite films were investigated, including a film designed to maximize the chemical adsorption and binding of gaseous UF6 onto the sampling planchet.« less

Isolation, purification and characterisation of selenium-containing polysaccharides and proteins in selenium-enriched Radix puerariae.

PubMed

Zou, Ye; Zhao, Ting; Mao, Guanghua; Zhang, Min; Zheng, Daheng; Feng, Weiwei; Wang, Wei; Wu, Xiangyang; Yang, Liuqing

2014-01-30

Selenium (Se) is an essential dietary mineral and Radix puerariae (RP) (the dried root of Pueraria lobata Willd.) is a botanical supplement widely used as a nutraceutical. Food enriched with Se provides a feasible and economic approach for production of organic Se compounds. However, little is known about Se-enriched RP and the structure of Se-containing polysaccharides and proteins derived from Se-enriched RP. The organic form of Se accounted for 82.42% of total content. Purification by DEAE-52 and Sephadex G-100 column chromatography yielded three single fractions--RP-SeP-11, RP-SeP-22 and RP-SeP-33--with Se contents of 0.9562 × 10⁻³, 0.6113 × 10⁻³ and 0.3827 × 10⁻³ g kg⁻¹, respectively. RP-SeP-11 (3.5 kDa) was made of glucose, RP-SeP-22 (19.6 kDa) was composed of xylose and glucose, and RP-SeP-33 (97.9 kDa) was made up galactose, mannose and glucose. Two Se-containing proteins were obtained with Se content of 3.175 × 10⁻³ and 4.328 × 10⁻³ g kg⁻¹, respectively. One appeared as three subunits with molecular masses of 43.0, 29.0 and 17.8 kDa while the other appeared as two subunits with molecular masses of 43.0 and 26.3 kDa. The results provide a basis for promoting the utilisation of RP resources enriched with Se as a promising tool for the food industry and are significant for its contribution to Se biochemistry in plants. © 2013 Society of Chemical Industry.

Overexpression of periostin and distinct mesothelin forms predict malignant progression in a rat cholangiocarcinoma model

PubMed Central

Manzanares, Miguel Á.; Campbell, Deanna J.W.; Maldonado, Gabrielle T.

2017-01-01

Periostin and mesothelin have each been suggested to be predictors of poor survival for patients with intrahepatic cholangiocarcinoma, although the clinical prognostic value of both of these biomarkers remains uncertain. The aim of the current study was to investigate these biomarkers for their potential to act as tumor progression factors when assessed in orthotopic tumor and three‐dimensional culture models of rat cholangiocarcinoma progression. Using our orthotopic model, we demonstrated a strong positive correlation between tumor and serum periostin and mesothelin and increasing liver tumor mass and associated peritoneal metastases that also reflected differences in cholangiocarcinoma cell aggressiveness and malignant grade. Periostin immunostaining was most prominent in the desmoplastic stroma of larger sized more aggressive liver tumors and peritoneal metastases. In comparison, mesothelin was more highly expressed in the cholangiocarcinoma cells; the slower growing more highly differentiated liver tumors exhibited a luminal cancer cell surface immunostaining for this biomarker, and the rapidly growing less differentiated liver and metastatic tumor masses largely showed cytoplasmic mesothelin immunoreactivity. Two molecular weight forms of mesothelin were identified, one at ∼40 kDa and the other, a more heavily glycosylated form, at ∼50 kDa. Increased expression of the 40‐kDa mesothelin over that of the 50 kDa form predicted increased malignant progression in both the orthotopic liver tumors and in cholangiocarcinoma cells of different malignant potential in three‐dimensional culture. Moreover, coculturing of cancer‐associated myofibroblasts with cholangiocarcinoma cells promoted overexpression of the 40‐kDa mesothelin, which correlated with enhanced malignant progression in vitro. Conclusion: Periostin and mesothelin are useful predictors of tumor progression in our rat desmoplastic cholangiocarcinoma models. This supports their relevance to human intrahepatic cholangiocarcinoma. (Hepatology Communications 2018;2:155–172) PMID:29404524

Detection of Prosecretory Mitogen Lacritin in Nonprimate Tears Primarily as a C-Terminal-Like Fragment

PubMed Central

Laurie, Diane E.; Splan, Rebecca K.; Green, Kari; Still, Katherine M.; McKown, Robert L.; Laurie, Gordon W.

2012-01-01

Purpose. Lacritin is a human tear glycoprotein that promotes basal tear protein secretion in cultured rat lacrimal acinar cells and proliferation of subconfluent human corneal epithelial cells. When topically added to rabbit eyes, lacritin promotes basal tearing. Despite these activities on several species, lacritin's presence in nonprimate tears or other tissues has not been explored. Here we probed for lacritin in normal horse tears. Methods. Sequences were collected from the Ensembl genomic alignment of human LACRT gene with high-quality draft horse genome (EquCab2.0) and analyzed. Normal horse tears were collected and assayed by Western blotting, ELISA, and mass spectrometry. Newly generated rabbit antibodies, respectively, against N- and C-terminal regions of human lacritin were employed. Results. Identity was 75% and 45%, respectively, at nucleotide and protein levels. Structural features were conserved, including a C-terminal amphipathic α-helix. Anti-C-terminal antibodies strongly detected a ∼13 kDa band in horse tears that was validated by mass spectrometry. In human tears, the same antibody detected uncleaved lacritin (∼24 kDa) strongly and C-terminal fragments of ∼13 and ∼11 kDa weakly. Anti-N-terminal antibodies were slightly reactive with a ∼24 kDa horse antigen and showed no reaction with the anti-C-terminal–reactive ∼13 kDa species. Similar respective levels of horse C-terminal versus N-terminal immunoreactivity were apparent by ELISA. Conclusions. Lacritin is present in horse tears, largely as a C-terminal fragment homologous to the mitogenic and bactericidal region in human lacritin, suggesting potential benefit in corneal wound repair. PMID:22871838

High-precision Penning trap mass measurements of 9,10Be and the one-neutron halo nuclide 11Be

NASA Astrophysics Data System (ADS)

Ringle, R.; Brodeur, M.; Brunner, T.; Ettenauer, S.; Smith, M.; Lapierre, A.; Ryjkov, V. L.; Delheij, P.; Drake, G. W. F.; Lassen, J.; Lunney, D.; Dilling, J.

2009-05-01

Penning trap mass measurements of 9Be, 10Be (t1 / 2 = 1.51 My), and the one-neutron halo nuclide 11Be (t1 / 2 = 13.8 s) have been performed using TITAN at TRIUMF. The resulting 11Be mass excess (ME = 20 177.60 (58) keV) is in agreement with the current Atomic Mass Evaluation (AME03) [G. Audi, et al., Nucl. Phys. A 729 (2003) 337] value, but is over an order of magnitude more precise. The precision of the mass values of 9,10Be have been improved by about a factor of four and reveal a ≈ 2 σ deviation from the AME mass values. Results of new atomic physics calculations are presented for the isotope shift of 11Be relative to 9Be, and it is shown that the new mass values essentially remove atomic mass uncertainties as a contributing factor in determining the relative nuclear charge radius from the isotope shift. The new mass values of 10,11Be also allow for a more precise determination of the single-neutron binding energy of the halo neutron in 11Be.

Apparent negative mass in QCM sensors due to punctual rigid loading

NASA Astrophysics Data System (ADS)

Castro, P.; Resa, P.; Elvira, L.

2012-12-01

Quartz Crystal Microbalances (QCM) are highly sensitive piezoelectric sensors able to detect very small loads attached to them. These devices are widely employed in many applications including process control and industrial and environmental monitoring. Mass loading is usually related to frequency shift by the well-known Sauerbrey's equation, valid for thin rigid homogeneous films. However, a significant deviation from this equation can occur when the mass is not uniformly distributed over the surface. Whereas the effects of a thin film on a QCM have been thoroughly studied, there are relatively few results on punctual loads, even though particles are usually deposited randomly and non-uniformly on the resonator surface. In this work, we have studied the effect of punctual rigid loading on the resonant frequency shift of a QCM sensor, both experimentally and using finite element method (FEM). The FEM numerical analysis was done using COMSOL software, 3D modeling a linear elastic piezoelectric solid and introducing the properties of an AT-cut quartz crystal. It is shown that a punctual rigid mass deposition on the surface of a QCM sensor can lead to positive shifts of resonance frequency, contrary to Sauerbrey's equation.

Shift work, job strain and changes in the body mass index among women: a prospective study.

PubMed

Fujishiro, Kaori; Lividoti Hibert, Eileen; Schernhammer, Eva; Rich-Edwards, Janet W

2017-06-01

The effects of job strain and shift work on weight gain have not been studied jointly. Cross-sectional and longitudinal studies on shift work and weight gain have reported different results. This study examines potential effect modification by job strain on the link between shift work and weight gain, and concurrent and delayed effects of shift work on weight gain. Data came from 52 622 women who participated in the Nurses' Health Study II, a prospective cohort study. Using linear regression, we modelled change in body mass index (BMI) over 4 years as a function of change in job strain, cumulative exposure to rotating night shift previously and during the 4 years (ie, previous and concurrent exposures) and the interaction between job strain and concurrent shift work exposure. Age, race/ethnicity, pregnancy history, baseline BMI, job types and health behaviours at baseline were controlled for. Job strain and rotating shift work, concurrent and previous, all had independent associations with BMI change during the 4-year period. There was no evidence for effect modification by job strain. Concurrent and previous exposures to rotating night shift had different associations with BMI change: an inverted U-shape for concurrent exposure (ranging from 0.01 to 0.14 kg/m 2 increase), a dose-response for previous exposure (-0.02 to 0.09 kg/m 2 ). Job strain and rotating night shift work have independent contributions to weight gain. Reducing job strain and supporting night shift workers are both important intervention goals. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/.

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

Chen, L; Shen, C; Wang, J

Purpose: To reduce cone beam CT (CBCT) imaging dose, we previously proposed a progressive dose control (PDC) scheme to employ temporal correlation between CBCT images at different fractions for image quality enhancement. A temporal non-local means (TNLM) method was developed to enhance quality of a new low-dose CBCT using existing high-quality CBCT. To enhance a voxel value, the TNLM method searches for similar voxels in a window. Due to patient deformation among the two CBCTs, a large searching window was required, reducing image quality and computational efficiency. This abstract proposes a deformation-assisted TNLM (DA-TNLM) method to solve this problem. Methods:more » For a low-dose CBCT to be enhanced using a high-quality CBCT, we first performed deformable image registration between the low-dose CBCT and the high-quality CBCT to approximately establish voxel correspondence between the two. A searching window for a voxel was then set based on the deformation vector field. Specifically, the search window for each voxel was shifted by the deformation vector. A TNLM step was then applied using only voxels within this determined window to correct image intensity at the low-dose CBCT. Results: We have tested the proposed scheme on simulated CIRS phantom data and real patient data. The CITS phantom was scanned on Varian onboard imaging CBCT system with coach shifting and dose reducing for each time. The real patient data was acquired in four fractions with dose reduced from standard CBCT dose to 12.5% of standard dose. It was found that the DA-TNLM method can reduce total dose by over 75% on average in the first four fractions. Conclusion: We have developed a PDC scheme which can enhance the quality of image scanned at low dose using a DA-TNLM method. Tests in phantom and patient studies demonstrated promising results.« less

Color vision predicts processing modes of goal activation during action cascading.

PubMed

Jongkees, Bryant J; Steenbergen, Laura; Colzato, Lorenza S

2017-09-01

One of the most important functions of cognitive control is action cascading: the ability to cope with multiple response options when confronted with various task goals. A recent study implicates a key role for dopamine (DA) in this process, suggesting higher D1 efficiency shifts the action cascading strategy toward a more serial processing mode, whereas higher D2 efficiency promotes a shift in the opposite direction by inducing a more parallel processing mode (Stock, Arning, Epplen, & Beste, 2014). Given that DA is found in high concentration in the retina and modulation of retinal DA release displays characteristics of D2-receptors (Peters, Schweibold, Przuntek, & Müller, 2000), color vision discrimination might serve as an index of D2 efficiency. We used color discrimination, assessed with the Lanthony Desaturated Panel D-15 test, to predict individual differences (N = 85) in a stop-change paradigm that provides a well-established measure of action cascading. In this task it is possible to calculate an individual slope value for each participant that estimates the degree of overlap in task goal activation. When the stopping process of a previous task goal has not finished at the time the change process toward a new task goal is initiated (parallel processing), the slope value becomes steeper. In case of less overlap (more serial processing), the slope value becomes flatter. As expected, participants showing better color vision were more prone to activate goals in a parallel manner as indicated by a steeper slope. Our findings suggest that color vision might represent a predictor of D2 efficiency and the predisposed processing mode of goal activation during action cascading. Copyright © 2017 Elsevier Ltd. All rights reserved.

Ultra High Mass Range Mass Spectrometer System

DOEpatents

Reilly, Peter T. A. [Knoxville, TN

2005-12-06

Applicant's present invention comprises mass spectrometer systems that operate in a mass range from 1 to 10.sup.16 DA. The mass spectrometer system comprising an inlet system comprising an aerodynamic lens system, a reverse jet being a gas flux generated in an annulus moving in a reverse direction and a multipole ion guide; a digital ion trap; and a thermal vaporization/ionization detector system. Applicant's present invention further comprises a quadrupole mass spectrometer system comprising an inlet system having a quadrupole mass filter and a thermal vaporization/ionization detector system. Applicant's present invention further comprises an inlet system for use with a mass spectrometer system, a method for slowing energetic particles using an inlet system. Applicant's present invention also comprises a detector device and a method for detecting high mass charged particles.

Mass-stiffness substructuring of an elastic metasurface for full transmission beam steering

NASA Astrophysics Data System (ADS)

Lee, Hyuk; Lee, Jun Kyu; Seung, Hong Min; Kim, Yoon Young

2018-03-01

The metasurface concept has a significant potential due to its novel wavefront-shaping functionalities that can be critically useful for ultrasonic and solid wave-based applications. To achieve the desired functionalities, elastic metasurfaces should cover full 2π phase shift and also acquire full transmission within subwavelength scale. However, they have not been explored much with respect to the elastic regime, because the intrinsic proportionality of mass-stiffness within the continuum elastic media causes an inevitable trade-off between abrupt phase shift and sufficient transmission. Our goal is to engineer an elastic metasurface that can realize an inverse relation between (amplified) effective mass and (weakened) stiffness in order to satisfy full 2π phase shift as well as full transmission. To achieve this goal, we propose a continuum elastic metasurface unit cell that is decomposed into two substructures, namely a mass-tuning substructure with a local dipolar resonator and a stiffness-tuning substructure composed of non-resonant multiply-perforated slits. We demonstrate analytically, numerically, and experimentally that this unique substructured unit cell can satisfy the required phase shift with high transmission. The substructuring enables independent tuning of the elastic properties over a wide range of values. We use a mass-spring model of the proposed continuum unit cell to investigate the working mechanism of the proposed metasurface. With the designed metasurface consisting of substructured unit cells embedded in an aluminum plate, we demonstrate that our metasurface can successfully realize anomalous steering and focusing of in-plane longitudinal ultrasonic beams. The proposed substructuring concept is expected to provide a new principle for the design of general elastic metasurfaces that can be used to efficiently engineer arbitrary wave profiles.

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

Zhang, Yaping; Williams, Brent J.; Goldstein, Allen H.

Here, we present a rapid method for apportioning the sources of atmospheric organic aerosol composition measured by gas chromatography–mass spectrometry methods. Here, we specifically apply this new analysis method to data acquired on a thermal desorption aerosol gas chromatograph (TAG) system. Gas chromatograms are divided by retention time into evenly spaced bins, within which the mass spectra are summed. A previous chromatogram binning method was introduced for the purpose of chromatogram structure deconvolution (e.g., major compound classes) (Zhang et al., 2014). Here we extend the method development for the specific purpose of determining aerosol samples' sources. Chromatogram bins are arrangedmore » into an input data matrix for positive matrix factorization (PMF), where the sample number is the row dimension and the mass-spectra-resolved eluting time intervals (bins) are the column dimension. Then two-dimensional PMF can effectively do three-dimensional factorization on the three-dimensional TAG mass spectra data. The retention time shift of the chromatogram is corrected by applying the median values of the different peaks' shifts. Bin width affects chemical resolution but does not affect PMF retrieval of the sources' time variations for low-factor solutions. A bin width smaller than the maximum retention shift among all samples requires retention time shift correction. A six-factor PMF comparison among aerosol mass spectrometry (AMS), TAG binning, and conventional TAG compound integration methods shows that the TAG binning method performs similarly to the integration method. However, the new binning method incorporates the entirety of the data set and requires significantly less pre-processing of the data than conventional single compound identification and integration. In addition, while a fraction of the most oxygenated aerosol does not elute through an underivatized TAG analysis, the TAG binning method does have the ability to achieve molecular level resolution on other bulk aerosol components commonly observed by the AMS.« less

Pancreatic Islets and Insulinoma Cells Express a Novel Isoform of Group VIA Phospholipase A2 (iPLA2β) that Participates in Glucose-Stimulated Insulin Secretion and Is Not Produced by Alternate Splicing of the iPLA2β Transcript†

PubMed Central

Ramanadham, Sasanka; Song, Haowei; Hsu, Fong-Fu; Zhang, Sheng; Crankshaw, Mark; Grant, Gregory A.; Newgard, Christopher B.; Bao, Shunzhong; Ma, Zhongmin; Turk, John

2013-01-01

Many cells express a group VIA 84 kDa phospholipase A2 (iPLA2β) that is sensitive to inhibition by a bromoenol lactone (BEL) suicide substrate. Inhibition of iPLA2β in pancreatic islets and insulinoma cells suppresses, and overexpression of iPLA2β in INS-1 insulinoma cells amplifies, glucose-stimulated insulin secretion, suggesting that iPLA2β participates in secretion. Western blotting analyses reveal that glucose-responsive 832/13 INS-1 cells express essentially no 84 kDa iPLA2β-immunoreactive protein but predominantly express a previously unrecognized immunoreactive iPLA2β protein in the 70 kDa region that is not generated by a mechanism of alternate splicing of the iPLA2β transcript. To determine if the 70 kDa-immunoreactive protein is a short isoform of iPLA2β, protein from the 70 kDa region was digested with trypsin and analyzed by mass spectrometry. Such analyses reveal several peptides with masses and amino acid sequences that exactly match iPLA2β tryptic peptides. Peptide sequences identified in the 70 kDa tryptic digest include iPLA2β residues 7–53, suggesting that the N-terminus is preserved. We also report here that the 832/13 INS-1 cells express iPLA2β catalytic activity and that BEL inhibits secretagogue-stimulated insulin secretion from these cells but not the incorporation of arachidonic acid into membrane PC pools of these cells. These observations suggest that the catalytic iPLA2β activity expressed in 832/13 INS-1 cells is attributable to a short isoform of iPLA2β and that this isoform participates in insulin secretory but not in membrane phospholipid remodeling pathways. Further, the finding that pancreatic islets also express predominantly a 70 kDa iPLA2β-immunoreactive protein suggests that a signal transduction role of iPLA2β in the native β-cell might be attributable to a 70 kDa isoform of iPLA2β. PMID:14636061

Pancreatic islets and insulinoma cells express a novel isoform of group VIA phospholipase A2 (iPLA2 beta) that participates in glucose-stimulated insulin secretion and is not produced by alternate splicing of the iPLA2 beta transcript.

PubMed

Ramanadham, Sasanka; Song, Haowei; Hsu, Fong-Fu; Zhang, Sheng; Crankshaw, Mark; Grant, Gregory A; Newgard, Christopher B; Bao, Shunzhong; Ma, Zhongmin; Turk, John

2003-12-02

Many cells express a group VIA 84 kDa phospholipase A(2) (iPLA(2)beta) that is sensitive to inhibition by a bromoenol lactone (BEL) suicide substrate. Inhibition of iPLA(2)beta in pancreatic islets and insulinoma cells suppresses, and overexpression of iPLA(2)beta in INS-1 insulinoma cells amplifies, glucose-stimulated insulin secretion, suggesting that iPLA(2)beta participates in secretion. Western blotting analyses reveal that glucose-responsive 832/13 INS-1 cells express essentially no 84 kDa iPLA(2)beta-immunoreactive protein but predominantly express a previously unrecognized immunoreactive iPLA(2)beta protein in the 70 kDa region that is not generated by a mechanism of alternate splicing of the iPLA(2)beta transcript. To determine if the 70 kDa-immunoreactive protein is a short isoform of iPLA(2)beta, protein from the 70 kDa region was digested with trypsin and analyzed by mass spectrometry. Such analyses reveal several peptides with masses and amino acid sequences that exactly match iPLA(2)beta tryptic peptides. Peptide sequences identified in the 70 kDa tryptic digest include iPLA(2)beta residues 7-53, suggesting that the N-terminus is preserved. We also report here that the 832/13 INS-1 cells express iPLA(2)beta catalytic activity and that BEL inhibits secretagogue-stimulated insulin secretion from these cells but not the incorporation of arachidonic acid into membrane PC pools of these cells. These observations suggest that the catalytic iPLA(2)beta activity expressed in 832/13 INS-1 cells is attributable to a short isoform of iPLA(2)beta and that this isoform participates in insulin secretory but not in membrane phospholipid remodeling pathways. Further, the finding that pancreatic islets also express predominantly a 70 kDa iPLA(2)beta-immunoreactive protein suggests that a signal transduction role of iPLA(2)beta in the native beta-cell might be attributable to a 70 kDa isoform of iPLA(2)beta.

High fat diet promotes achievement of peak bone mass in young rats

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

Malvi, Parmanand; Piprode, Vikrant; Chaube, Balkrishna

Highlights: • High fat diet helps in achieving peak bone mass at younger age. • Shifting from high fat to normal diet normalizes obese parameters. • Bone parameters are sustained even after withdrawal of high fat diet. - Abstract: The relationship between obesity and bone is complex. Epidemiological studies demonstrate positive as well as negative correlation between obesity and bone health. In the present study, we investigated the impact of high fat diet-induced obesity on peak bone mass. After 9 months of feeding young rats with high fat diet, we observed obesity phenotype in rats with increased body weight, fatmore » mass, serum triglycerides and cholesterol. There were significant increases in serum total alkaline phosphatase, bone mineral density and bone mineral content. By micro-computed tomography (μ-CT), we observed a trend of better trabecular bones with respect to their microarchitecture and geometry. This indicated that high fat diet helps in achieving peak bone mass and microstructure at younger age. We subsequently shifted rats from high fat diet to normal diet for 6 months and evaluated bone/obesity parameters. It was observed that after shifting rats from high fat diet to normal diet, fat mass, serum triglycerides and cholesterol were significantly decreased. Interestingly, the gain in bone mineral density, bone mineral content and trabecular bone parameters by HFD was retained even after body weight and obesity were normalized. These results suggest that fat rich diet during growth could accelerate achievement of peak bone mass that is sustainable even after withdrawal of high fat diet.« less

MALDI imaging mass spectrometry analysis-A new approach for protein mapping in multiple sclerosis brain lesions.

PubMed

Maccarrone, Giuseppina; Nischwitz, Sandra; Deininger, Sören-Oliver; Hornung, Joachim; König, Fatima Barbara; Stadelmann, Christine; Turck, Christoph W; Weber, Frank

2017-03-15

Multiple sclerosis is a disease of the central nervous system characterized by recurrent inflammatory demyelinating lesions in the early disease stage. Lesion formation and mechanisms leading to lesion remyelination are not fully understood. Matrix Assisted Laser Desorption Ionisation Mass Spectrometry imaging (MALDI-IMS) is a technology which analyses proteins and peptides in tissue, preserves their spatial localization, and generates molecular maps within the tissue section. In a pilot study we employed MALDI imaging mass spectrometry to profile and identify peptides and proteins expressed in normal-appearing white matter, grey matter and multiple sclerosis brain lesions with different extents of remyelination. The unsupervised clustering analysis of the mass spectra generated images which reflected the tissue section morphology in luxol fast blue stain and in myelin basic protein immunohistochemistry. Lesions with low remyelination extent were defined by compounds with molecular weight smaller than 5300Da, while more completely remyelinated lesions showed compounds with molecular weights greater than 15,200Da. An in-depth analysis of the mass spectra enabled the detection of cortical lesions which were not seen by routine luxol fast blue histology. An ion mass, mainly distributed at the rim of multiple sclerosis lesions, was identified by liquid chromatography and tandem mass spectrometry as thymosin beta-4, a protein known to be involved in cell migration and in restorative processes. The ion mass of thymosin beta-4 was profiled by MALDI imaging mass spectrometry in brain slides of 12 multiple sclerosis patients and validated by immunohistochemical analysis. In summary, our results demonstrate the ability of the MALDI-IMS technology to map proteins within the brain parenchyma and multiple sclerosis lesions and to identify potential markers involved in multiple sclerosis pathogenesis and/or remyelination. Copyright © 2016 Elsevier B.V. All rights reserved.

Astrophysical Implications of a New Dynamical Mass for the Nearby White Dwarf 40 Eridani B

NASA Astrophysics Data System (ADS)

Bond, Howard E.; Bergeron, P.; Bédard, A.

2017-10-01

The bright, nearby DA-type white dwarf (WD) 40 Eridani B is orbited by the M dwarf 40 Eri C, allowing determination of the WD’s mass. Until recently, however, the mass depended on orbital elements determined four decades ago, and that mass was so low that it created several astrophysical puzzles. Using new astrometric measurements, the binary-star group at the U.S. Naval Observatory has revised the dynamical mass upward, to 0.573 ± 0.018 M ⊙. In this paper, we use model-atmosphere analysis to update other parameters of the WD, including effective temperature, surface gravity, radius, and luminosity. We then compare these results with WD interior models. Within the observational uncertainties, theoretical cooling tracks for CO-core WDs of its measured mass are consistent with the position of 40 Eri B in the H-R diagram; equivalently, the theoretical mass-radius relation (MRR) is consistent with the star’s location in the mass-radius plane. This consistency is, however, achieved only if we assume a “thin” outer hydrogen layer, with q H = M H/M WD ≃ 10-10. We discuss other evidence that a significant fraction of DA WDs have such thin H layers, in spite of the expectation from canonical stellar-evolution theory of “thick” H layers with q H ≃ 10-4. The cooling age of 40 Eri B is ˜122 Myr, and its total age is ˜1.8 Gyr. We present the MRRs for 40 Eri B and three other nearby WDs in visual binaries with precise mass determinations, and show that the agreement of current theory with observations is excellent in all cases.

Method for calibrating mass spectrometers

DOEpatents

Anderson, Gordon A [Benton City, WA; Brands, Michael D [Richland, WA; Bruce, James E [Schwenksville, PA; Pasa-Tolic, Ljiljana [Richland, WA; Smith, Richard D [Richland, WA

2002-12-24

A method whereby a mass spectra generated by a mass spectrometer is calibrated by shifting the parameters used by the spectrometer to assign masses to the spectra in a manner which reconciles the signal of ions within the spectra having equal mass but differing charge states, or by reconciling ions having known differences in mass to relative values consistent with those known differences. In this manner, the mass spectrometer is calibrated without the need for standards while allowing the generation of a highly accurate mass spectra by the instrument.

Purification and Characterization of Carbaryl Hydrolase from Blastobacter sp. Strain M501

PubMed Central

Hayatsu, Masahito; Nagata, Tadahiro

1993-01-01

A bacterium capable of hydrolyzing carbaryl (1-naphthyl-N-methylcarbamate) was isolated from a soil enrichment. This bacterium was characterized taxonomically as a Blastobacter sp. and designated strain M501. A carbaryl hydrolase present in this strain was purified to homogeneity by protamine sulfate treatment, ammonium sulfate precipitation, and hydrophobic, anion-exchange, gel filtration, and hydroxylapatite chromatographies. The native enzyme had a molecular mass of 166,000 Da and was composed of two subunits with molecular masses of 84,000 Da. The optimum pH and temperature of the enzyme activity were 9.0 and 45°C, respectively. The enzyme was not stable at temperatures above 40°C. The purified enzyme hydrolyzed seven N-methylcarbamate insecticides and also exhibited activity against 1-naphthyl acetate and 4-nitrophenyl acetate. Images PMID:16348989

Size-focusing synthesis of gold nanoclusters with p-mercaptobenzoic acid.

PubMed

Tvedte, Laura M; Ackerson, Christopher J

2014-09-18

Etching or size-focusing methods are now widespread for preparation of atomically monodisperse thiolate-protected gold nanoparticles. Size-focusing methods are not widespread, however, in the production of water-soluble gold nanoparticles. Reported here is a new method for size-focusing of large gold nanoparticles utilizing p-mercaptobenzoic acid. We observe preferential formation of three large gold nanoparticles with approximate masses of 23, 51, and 88 kDa. On the basis of the stability of these masses against further etching or growth, they appear to be especially stable sizes. These sizes are not prominent after etching challenges with organosoluble ligands, and the 51 and 88 kDa sizes appear to be novel stable thiolate-protected gold cluster sizes. The overall trend in particle size distribution over time is also unusual, with larger sizes dominating at longer time points.

Tools to discover anionic and nonionic polyfluorinated alkyl surfactants by liquid chromatography electrospray ionisation mass spectrometry.

PubMed

Trier, Xenia; Granby, Kit; Christensen, Jan H

2011-10-07

A tiered approach is proposed for the discovery of unknown anionic and nonionic polyfluorinated alkyl surfactants (PFASs) by reversed phase ultra high performance liquid chromatography (UHPLC)--negative electrospray ionisation--quadrupole time of flight mass spectrometry (UHPLC-ESI(-)-QTOF-MS). The chromatographic separation, ionisation and detection of PFASs mixtures, was achieved at high pH (pH=9.7) with NH(4)OH as additive. To distinguish PFASs from other chemicals we used the characteristic negative mass defects of PFASs, their specific losses of 20 Da (HF) and the presence of series of chromatographic peaks, belonging to homologues series with m/z of n×50 Da (CF(2)) or n×100 Da (CF(2)CF(2)). The elemental composition of the precursor ions were deducted from the accurate m/z values of the deprotonated molecules [M-H](-). In case of in-source fragmentation, the presence of dimers, e.g. [M(2)-H](-) and adduct ions such as [M-H+solvent](-) and [(M-H)(M-H+Na)(n)](-) were used to confirm the identity of the precursor ions. In relation to quantification of PFASs, we discuss how their surfactancy influence the ESI processes, challenge their handling in solution and choices of precursor-to-product ions for MSMS of e.g., structural PFAS isomers. The method has been used to discover PFASs in industrial blends and in extracts from food contact materials. Copyright © 2011 Elsevier B.V. All rights reserved.

Isolation and characterization of large spectrum and multiple bacteriocin-producing Enterococcus faecium strain from raw bovine milk.

PubMed

Gaaloul, N; ben Braiek, O; Hani, K; Volski, A; Chikindas, M L; Ghrairi, T

2015-02-01

To assess the antimicrobial properties of lactic acid bacteria from Tunisian raw bovine milk. A bacteriocin-producing Enterococcus faecium strain was isolated from raw cow milk with activity against Gram-positive and Gram-negative bacteria. Antimicrobial substances produced by this strain were sensitive to proteolytic enzymes and were thermostable and resistant to a broad range of pH (2-10). Mode of action of antimicrobial substances was determined as bactericidal. Maximum activity was reached at the end of the exponential growth phase when checked against Listeria ivanovii BUG 496 (2366.62 AU ml(-1)). However, maximum antimicrobial activity against Pseudomonas aeruginosa 28753 was recorded at the beginning of the exponential growth phase. Enterococcus faecium GGN7 was characterized as free from virulence factors and was susceptible to tested antibiotics. PCR analysis of the micro-organism's genome revealed the presence of genes coding for enterocins A and B. Mass spectrometry analysis of RP-HPLC active fractions showed molecular masses corresponding to enterocins A (4835.77 Da) and B (5471.56 Da), and a peptide with a molecular mass of 3215.5 Da active only against Gram-negative indicator strains. The latter was unique in the databases. Enterococcus faecium GGN7 produces three bacteriocins with different inhibitory spectra. Based on its antimicrobial properties and safety, Ent. faecium GGN7 is potentially useful for food biopreservation. The results suggest the bacteriocins from GGN7 strain could be useful for food biopreservation. © 2014 The Society for Applied Microbiology.

Multiconfiguration calculations of electronic isotope shift factors in Al i

NASA Astrophysics Data System (ADS)

Filippin, Livio; Beerwerth, Randolf; Ekman, Jörgen; Fritzsche, Stephan; Godefroid, Michel; Jönsson, Per

2016-12-01

The present work reports results from systematic multiconfiguration Dirac-Hartree-Fock calculations of electronic isotope shift factors for a set of transitions between low-lying levels of neutral aluminium. These electronic quantities together with observed isotope shifts between different pairs of isotopes provide the changes in mean-square charge radii of the atomic nuclei. Two computational approaches are adopted for the estimation of the mass- and field-shift factors. Within these approaches, different models for electron correlation are explored in a systematic way to determine a reliable computational strategy and to estimate theoretical error bars of the isotope shift factors.

Full Robotic Colorectal Resections for Cancer Combined With Other Major Surgical Procedures: Early Experience With the da Vinci Xi.

PubMed

Morelli, Luca; Di Franco, Gregorio; Guadagni, Simone; Palmeri, Matteo; Gianardi, Desirée; Bianchini, Matteo; Moglia, Andrea; Ferrari, Vincenzo; Caprili, Giovanni; D'Isidoro, Cristiano; Melfi, Franca; Di Candio, Giulio; Mosca, Franco

2017-08-01

The da Vinci Xi has been developed to overcome some of the limitations of the previous platform, thereby increasing the acceptance of its use in robotic multiorgan surgery. Between January 2015 and October 2015, 10 patients with synchronous tumors of the colorectum and others abdominal organs underwent robotic combined resections with the da Vinci Xi. Trocar positions respected the Universal Port Placement Guidelines provided by Intuitive Surgical for "left lower quadrant," with trocars centered on the umbilical area, or shifted 2 to 3 cm to the right or to the left, depending on the type of combined surgical procedure. All procedures were completed with the full robotic technique. Simultaneous procedures in same quadrant or left quadrant and pelvis, or left/right and upper, were performed with a single docking/single targeting approach; in cases of left/right quadrant or right quadrant/pelvis, we performed a dual-targeting operation. No external collisions or problems related to trocar positions were noted. No patient experienced postoperative surgical complications and the mean hospital stay was 6 days. The high success rate of full robotic colorectal resection combined with other surgical interventions for synchronous tumors, suggest the efficacy of the da Vinci Xi in this setting.

A long-term perspective on biomass burning in the Serra da Estrela, Portugal

NASA Astrophysics Data System (ADS)

Connor, Simon E.; Araújo, João; van der Knaap, Willem O.; van Leeuwen, Jacqueline F. N.

2012-11-01

Fire is currently perceived as a major threat to ecosystems and biodiversity in the mountains of the Mediterranean region. Portugal's highest mountain range, the Serra da Estrela, is one of the country's most important protected areas and also the most fire-prone. We present a ˜14,000-year fire history based on microscopic charred particles in an infilled glacial lake to better understand the antiquity of biomass burning and its effects on Mediterranean vegetation at the Atlantic margin. Results indicate the continuous occurrence of fire in the Serra da Estrela over the period of the record. Two periods of increased fire activity - around 12,000-11,000 calendar years before the present (cal. a BP) and 3500-2500 cal. a BP - were accompanied by major vegetation changes and followed by long periods of vegetation stabilisation. Cross-correlation analyses reveal that post-fire succession consistently began with herbaceous vegetation, followed by forest and shrubland stages. Past successional trends were often markedly different to those observed at present. Holocene climatic changes, including shifts in the North Atlantic Oscillation, played a pivotal role in the vegetation development and fire history of the Serra da Estrela. In the late Holocene, human use of fire became a major agent of vegetation change, accelerating the Holocene decline of forests.

Head-on collision between two dust acoustic solitary waves and study of rogue waves in multicomponent dusty plasma

NASA Astrophysics Data System (ADS)

Singh, Kuldeep; Kaur, Nimardeep; Saini, N. S.

2017-06-01

In this investigation, the study of head-on collision between two dust acoustic solitary waves (DASWs) and characteristics of rogue waves in a dusty plasma composed of dust fluid, kappa distributed ions, electrons, and positrons has been presented. Two Korteweg-de Vries equations are derived by employing the extended Poincaré-Lighthill-Kuo reductive perturbation method. The analytical phase shifts and trajectories after head-on collision of two DA solitary waves have been studied numerically. It is found that the presence of superthermal ions, electrons, as well as positrons; concentrations of electrons and positrons; and temperature of electrons and dust have an emphatic influence on the phase shifts after the head-on collision of two rarefactive DA solitary waves. The time evolution of two rarefactive DASWs has also been presented. Further, the generation of dust acoustic rogue waves (DARWs) has been studied in the framework of rational solution of nonlinear Schrödinger equation. The dependence of the rogue wave profile on the relevant physical parameters has been discussed in detail. It is emphasized that the real implementation of our present results may be of great importance in different regions of space and astrophysical environments, especially in the interstellar medium and Jupiter rings.

A thermostable manganese-containing superoxide dismutase from the thermophilic fungus Thermomyces lanuginosus.

PubMed

Li, Duo-Chuan; Gao, Jing; Li, Ya-Ling; Lu, Jing

2005-02-01

A thermostable superoxide dismutase (SOD) from a Thermomyces lanuginosus strain (P134) was purified to homogeneity by fractional ammonium sulfate precipitation, ion-exchange chromatography on DEAE-Sepharose, Phenyl-Sepharose hydrophobic interaction chromatography, and gel filtration on Sephacryl S-100. The molecular mass of a single band of the enzyme was estimated to be 22.4 kDa, using sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Using gel filtration on Sephacryl S-100, the molecular mass was estimated to be 89.1 kDa, indicating that this enzyme was composed of four identical subunits of 22.4 kDa each. The SOD was found to be inhibited by NaN3, but not by KCN or H2O2, suggesting that the SOD in T. lanuginosus was of the manganese superoxide dismutase type. The SOD exhibited maximal activity at pH 7.5. The optimum temperature for the activity was 55 degrees C. It was thermostable at 50 and 60 degrees C and retained 55% activity after 60 min at 70 degrees C. The half-life of the SOD at 80 degrees C was approximately 28 min and even retained 20% activity after 20 min at 90 degrees C.

An azoreductase, aerobic NADH-dependent flavoprotein discovered from Bacillus sp.: functional expression and enzymatic characterization.

PubMed

Ooi, Toshihiko; Shibata, Takeshi; Sato, Reiko; Ohno, Hiroaki; Kinoshita, Shinichi; Thuoc, Tran Linh; Taguchi, Seiichi

2007-05-01

The gene coding for an azoreductase, designated as an azrA, was cloned by polymerase chain reaction amplification from the genomic DNA of Bacillus sp. strain B29 isolated from soil. The azrA encoded a protein of 208 amino acids with calculated molecular mass of 22,766 Da. The enzyme was heterologously expressed in Escherichia coli with a strong band of 23 kDa on sodium dodecyl sulfate polyacrylamide gel electrophoresis. Purified recombinant AzrA was a homodimer with a native molecular mass of 48 kDa containing two molecules of flavin mononucleotide (FMN; oxidized). This activity was oxygen insensitive and was nicotinamide adenine dinucleotide (reduced form; NADH) dependent. Recombinant AzrA exhibited a broad pH stability between 6 and 10 with a temperature optimum of 60-80 degrees C. The enzyme cleaved the model azo compound of methyl red [MR, 4'-(dimethylamino)-azobenzene-2-carboxylic acid] into 2-aminobenzoic acid and N, N'-dimethyl-p-phenylenediamine by ping-pong mechanism. The enzyme was not only able to decolorize MR but also able to decolorize sulfonated azo dyes such as Orange I and Acid Red 88.

Purification and characterization of vanillin dehydrogenases from alkaliphile Micrococcus sp. TA1 and neutrophile Burkholderia cepacia TM1.

PubMed

Mitsui, Ryoji; Hirota, Mizuho; Tsuno, Takuo; Tanaka, Mitsuo

2010-02-01

Vanillin dehydrogenases (VDHs) were purified and characterized from two bacterial strains that have different pH dependencies for growth. The alkaliphile Micrococcus sp. TA1, isolated from an alkaline spa, can grow on several aromatic compounds such as ferulic acid, vanillin, vanillic acid, and protocatechuic acid under alkaline conditions. The neutrophile Burkholderia cepacia TM1, which was isolated previously, also grew on the above-mentioned compounds because they functioned as the sole carbon source under neutral conditions. Purified VDHs showed activities toward some aromatic aldehydes. These enzymes have the same subunit molecular mass of about 57 kDa as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, but differed in some of their observed properties. Native molecular masses also differed between the purified enzymes. These were 250 kDa for the enzyme from alkaliphilic strain TA1 and 110 kDa for that from neutrophilic strain TM1, as determined by gel filtration. The enzyme from strain TA1 required NADP(+) as a coenzyme for its activity, but that from strain TM1 required NAD(+). These results are important because this is the first report of an alkaliphilic bacterium consuming lignin monomers.

Bolevenine, a toxic protein from the Japanese toadstool Boletus venenatus.

PubMed

Matsuura, Masanori; Yamada, Mina; Saikawa, Yoko; Miyairi, Kazuo; Okuno, Toshikatsu; Konno, Katsuhiro; Uenishi, Jun'ichi; Hashimoto, Kimiko; Nakata, Masaya

2007-03-01

A toxic protein, called bolevenine, was isolated from the toxic mushroom Boletus venenatus based on its lethal effects on mice. On SDS-PAGE, in either the presence or absence of 2-mercaptoethanol, this protein showed a single band of approximately 12 kDa. In contrast, based on gel filtration and MALDI-TOFMS, its relative molecular mass was estimated to be approximately 30 kDa and approximately 33 kDa, respectively, indicating that the protein consists of three identical subunits. This toxin exhibited its lethal activity following injection at 10mg/kg into mice. The N-terminal amino acid sequence was determined up to 18, and found to be similar to the previously reported bolesatine, a toxic compound isolated from Boletus satanas.

Equivalence principle for quantum systems: dephasing and phase shift of free-falling particles

NASA Astrophysics Data System (ADS)

Anastopoulos, C.; Hu, B. L.

2018-02-01

We ask the question of how the (weak) equivalence principle established in classical gravitational physics should be reformulated and interpreted for massive quantum objects that may also have internal degrees of freedom (dof). This inquiry is necessary because even elementary concepts like a classical trajectory are not well defined in quantum physics—trajectories originating from quantum histories become viable entities only under stringent decoherence conditions. From this investigation we posit two logically and operationally distinct statements of the equivalence principle for quantum systems. Version A: the probability distribution of position for a free-falling particle is the same as the probability distribution of a free particle, modulo a mass-independent shift of its mean. Version B: any two particles with the same velocity wave-function behave identically in free fall, irrespective of their masses. Both statements apply to all quantum states, including those without a classical correspondence, and also for composite particles with quantum internal dof. We also investigate the consequences of the interaction between internal and external dof induced by free fall. For a class of initial states, we find dephasing occurs for the translational dof, namely, the suppression of the off-diagonal terms of the density matrix, in the position basis. We also find a gravitational phase shift in the reduced density matrix of the internal dof that does not depend on the particle’s mass. For classical states, the phase shift has a natural classical interpretation in terms of gravitational red-shift and special relativistic time-dilation.

Dendrite architecture organized by transcriptional control of the F-actin nucleator Spire.

PubMed

Ferreira, Tiago; Ou, Yimiao; Li, Sally; Giniger, Edward; van Meyel, Donald J

2014-02-01

The architectures of dendritic trees are crucial for the wiring and function of neuronal circuits because they determine coverage of receptive territories, as well as the nature and strength of sensory or synaptic inputs. Here, we describe a cell-intrinsic pathway sculpting dendritic arborization (da) neurons in Drosophila that requires Longitudinals Lacking (Lola), a BTB/POZ transcription factor, and its control of the F-actin cytoskeleton through Spire (Spir), an actin nucleation protein. Loss of Lola from da neurons reduced the overall length of dendritic arbors, increased the expression of Spir, and produced inappropriate F-actin-rich dendrites at positions too near the cell soma. Selective removal of Lola from only class IV da neurons decreased the evasive responses of larvae to nociception. The increased Spir expression contributed to the abnormal F-actin-rich dendrites and the decreased nocifensive responses because both were suppressed by reduced dose of Spir. Thus, an important role of Lola is to limit expression of Spir to appropriate levels within da neurons. We found Spir to be expressed in dendritic arbors and to be important for their development. Removal of Spir from class IV da neurons reduced F-actin levels and total branch number, shifted the position of greatest branch density away from the cell soma, and compromised nocifensive behavior. We conclude that the Lola-Spir pathway is crucial for the spatial arrangement of branches within dendritic trees and for neural circuit function because it provides balanced control of the F-actin cytoskeleton.

Use of matrix-assisted laser desorption ionization-time of flight mass spectrometry to identify MLST clade 4 Clostridium difficile isolates.

PubMed

Cheng, Jing-Wei; Liu, Chang; Kudinha, Timothy; Xiao, Meng; Yu, Shu-Ying; Yang, Chun-Xia; Wei, Ming; Liang, Guo-Wei; Shao, Dong-Hua; Kong, Fanrong; Tong, Zhao-Hui; Xu, Ying-Chun

2018-04-26

Clostridium difficile is the leading cause of health care-associated infections. Previous studies suggest that C. difficile MLST clade 4 strains with higher drug resistance rates constitute the major clone spreading in China. Thus development of a rapid and accurate typing method for these strains is needed to monitor the epidemiology of this clone and to guide clinical treatment. A total of 160 non-duplicate C. difficile isolates recovered from three large teaching hospitals in Beijing were studied. All the 41 clade 4 C. difficile isolates clustered together on the PCA dendrogram. Spectra peak statistics revealed that five markers (2691.43Da, 2704.91Da, 2711.93Da, 3247.27Da and 3290.76Da) can easily and reliably distinguish between clade 4 and non-clade 4 isolates, with area under the curve (AUC) values of 0.991, 0.997, 0.973, 1 and 1, respectively. In conclusion, MALDI-TOF MS is a very simple and accurate method for identifying C. difficile MLST clade 4 strains. Copyright © 2018 Elsevier Inc. All rights reserved.

Purification and characterization of a milk-clotting aspartic proteinase from globe artichoke (Cynara scolymus L.).

PubMed

Llorente, Berta E; Brutti, Cristina B; Caffini, Néstor O

2004-12-29

The study of proteinase expression in crude extracts from different organs of the globe artichoke (Cynara scolymus L.) disclosed that enzymes with proteolytic and milk-clotting activity are mainly located in mature flowers. Maximum proteolytic activity was recorded at pH 5.0, and inhibition studies showed that only pepstatin, specific for aspartic proteinases, presented a significant inhibitory effect. Such properties, in addition to easy enzyme inactivation by moderate heating, make this crude protease extract potentially useful for cheese production. Adsorption with activated carbon, together with anion exchange and affinity chromatography, led to the isolation of a heterodimeric milk-clotting proteinase consisting of 30- and 15-kDa subunits. MALDI-TOF MS of the 15-kDa chain determined a 15.358-Da mass, and the terminal amino sequence presented 96% homology with the smaller cardosin A subunit. The amino terminal sequence of the 30-kDa chain proved to be identical to the larger cardosin A subunit. Electrophoresis evidenced proteinase self-processing that was confirmed by immunoblots presenting 62-, 30-, and 15-kDa bands.

Loss of Mitochondrial Fission Depletes Axonal Mitochondria in Midbrain Dopamine Neurons

PubMed Central

Berthet, Amandine; Margolis, Elyssa B.; Zhang, Jue; Hsieh, Ivy; Zhang, Jiasheng; Hnasko, Thomas S.; Ahmad, Jawad; Edwards, Robert H.; Sesaki, Hiromi; Huang, Eric J.

2014-01-01

Disruptions in mitochondrial dynamics may contribute to the selective degeneration of dopamine (DA) neurons in Parkinson's disease (PD). However, little is known about the normal functions of mitochondrial dynamics in these neurons, especially in axons where degeneration begins, and this makes it difficult to understand the disease process. To study one aspect of mitochondrial dynamics—mitochondrial fission—in mouse DA neurons, we deleted the central fission protein dynamin-related protein 1 (Drp1). Drp1 loss rapidly eliminates the DA terminals in the caudate–putamen and causes cell bodies in the midbrain to degenerate and lose α-synuclein. Without Drp1, mitochondrial mass dramatically decreases, especially in axons, where the mitochondrial movement becomes uncoordinated. However, in the ventral tegmental area (VTA), a subset of midbrain DA neurons characterized by small hyperpolarization-activated cation currents (Ih) is spared, despite near complete loss of their axonal mitochondria. Drp1 is thus critical for targeting mitochondria to the nerve terminal, and a disruption in mitochondrial fission can contribute to the preferential death of nigrostriatal DA neurons. PMID:25339743

Growth hormone: its measurement and the need for assay harmonization.

PubMed

Wood, P

2001-09-01

Serum human growth hormone (hGH) assays show a wide range in bias and in cut-off values for provocative tests, which vary from 13.5 to 35-40 mU/L when they have been established. Studies using novel hGH assays show that methods that are absolutely specific for 22-kDa hGH may not identify bioactive hGH peaks and that 20:22-kDa hGH ratios are increased in acromegaly. Greater harmonization of serum hGH methods can be achieved by: changing from IS 80/505 to IS 98/574, which is calibrated in mass units of recombinant 22-kDa hGH; using monoclonal/polyclonal or polyclonal/polyclonal antibody combinations that measure both 20-kDa and 22-kDa hGH; the development of assays such as the immunofunctional hGH assay which has the convenience of an immunometric assay but gives results that correlate better with bioassays collaboration between manufacturers and laboratories to establish method-related cut-off limits for provocative tests of hGH status.

Loss of mitochondrial fission depletes axonal mitochondria in midbrain dopamine neurons.

PubMed

Berthet, Amandine; Margolis, Elyssa B; Zhang, Jue; Hsieh, Ivy; Zhang, Jiasheng; Hnasko, Thomas S; Ahmad, Jawad; Edwards, Robert H; Sesaki, Hiromi; Huang, Eric J; Nakamura, Ken

2014-10-22

Disruptions in mitochondrial dynamics may contribute to the selective degeneration of dopamine (DA) neurons in Parkinson's disease (PD). However, little is known about the normal functions of mitochondrial dynamics in these neurons, especially in axons where degeneration begins, and this makes it difficult to understand the disease process. To study one aspect of mitochondrial dynamics-mitochondrial fission-in mouse DA neurons, we deleted the central fission protein dynamin-related protein 1 (Drp1). Drp1 loss rapidly eliminates the DA terminals in the caudate-putamen and causes cell bodies in the midbrain to degenerate and lose α-synuclein. Without Drp1, mitochondrial mass dramatically decreases, especially in axons, where the mitochondrial movement becomes uncoordinated. However, in the ventral tegmental area (VTA), a subset of midbrain DA neurons characterized by small hyperpolarization-activated cation currents (Ih) is spared, despite near complete loss of their axonal mitochondria. Drp1 is thus critical for targeting mitochondria to the nerve terminal, and a disruption in mitochondrial fission can contribute to the preferential death of nigrostriatal DA neurons. Copyright © 2014 the authors 0270-6474/14/3414304-14$15.00/0.

Momentum peak shift and width of longitudinal momentum distribution of projectilelike fragments produced at E =290 MeV /nucleon

NASA Astrophysics Data System (ADS)

Momota, S.; Kanazawa, M.; Kitagawa, A.; Sato, S.

2018-04-01

Longitudinal momentum (PL) distributions of projectilelike fragments produced at E =290 MeV /nucleon are investigated. PL distributions of fragments produced by Ar and Kr beams with a wide variety of targets (C, Al, Nb, Tb, and Au) were measured using the fragment separator at HIMAC. PL distributions observed for fragments with a wide range of mass losses Δ A (1-30 for Ar beam and 1-64 for Kr beam), show a slightly, but definitely asymmetric nature. The peak shift and width were obtained from the observed PL distributions. No significant target dependence was found in either the peak shift or width. For the practical application, the variation in momentum peak shift with fragment mass (AF) was represented by a parabolic function. The width on the high-PL side (σHigh) is well reproduced by the Goldhaber formula, which is obtained from the contribution of the Fermi momentum. The behavior of the reduced width, σ0, obtained from σHigh via the Goldhaber formulation, is consistent with the mass-dependent Fermi momentum of a nucleon. The width on the low-PL side (σLow) is markedly larger than σHigh and exhibits a clear AF dependence.

HEALTH CONDITIONS LINKED TO AGE-RELATED MACULAR DEGENERATION ASSOCIATED WITH DARK ADAPTATION.

PubMed

Laíns, Inês; Miller, John B; Mukai, Ryo; Mach, Steven; Vavvas, Demetrios; Kim, Ivana K; Miller, Joan W; Husain, Deeba

2018-06-01

To determine the association between dark adaption (DA) and different health conditions linked with age-related macular degeneration (AMD). Cross-sectional study, including patients with AMD and a control group. Age-related macular degeneration was graded according to the Age-Related Eye Disease Study (AREDS) classification. We obtained data on medical history, medications, and lifestyle. Dark adaption was assessed with the extended protocol (20 minutes) of AdaptDx (MacuLogix). For analyses, the right eye or the eye with more advanced AMD was selected. Multivariate linear and logistic regressions were performed, accounting for age and AMD stage. Seventy-eight subjects (75.6% AMD; 24.4% controls) were included. Multivariate assessments revealed that body mass index (BMI; β = 0.30, P = 0.045), taking AREDS vitamins (β = 5.51, P < 0.001), and family history of AMD (β = 2.68, P = 0.039) were significantly associated with worse rod intercept times. Abnormal DA (rod intercept time ≥ 6.5 minutes) was significantly associated with family history of AMD (β = 1.84, P = 0.006), taking AREDS supplements (β = 1.67, P = 0.021) and alcohol intake (β = 0.07, P = 0.017). Besides age and AMD stage, a higher body mass index, higher alcohol intake, and a family history of AMD seem to impair DA. In this cohort, the use of AREDS vitamins was also statistically linked with impaired DA, most likely because of an increased severity of disease in subjects taking them.

Measuring masses of single bacterial whole cells with a quadrupole ion trap.

PubMed

Peng, Wen-Ping; Yang, Yi-Chang; Kang, Ming-Wei; Lee, Yuan T; Chang, Huan-Cheng

2004-09-29

A novel method has been developed to precisely measure the masses of single bacterial whole cells using a quadrupole ion trap as an electrodynamic balance. The bacterial cells were introduced into the ion trap by matrix-assisted laser desorption/ionization, confined in space by audio frequency ac fields, and detected by elastic light scattering. Mass measurement accuracy approaching 0.1% was achieved for Escherichia coli K-12 with a mass distribution of +/-3% from 60 repetitive measurements of the particles and their clusters. This is the first high-precision mass measurement reported for any intact microorganisms with masses greater than 1 x 1010 Da. The method opens new avenues for high-precision mass measurement of single microbial particles and offers an alternative approach for rapid identification of microorganisms by mass spectrometry.

PG1258+593 and its common proper motion magnetic white dwarf counterpart

NASA Astrophysics Data System (ADS)

Girven, J.; Gänsicke, B. T.; Külebi, B.; Steeghs, D.; Jordan, S.; Marsh, T. R.; Koester, D.

2010-05-01

We confirm SDSSJ130033.48+590407.0 as a common proper motion companion to the well-studied hydrogen-atmosphere (DA) white dwarf PG1258+593 (GD322). The system lies at a distance of 68 +/- 3pc, where the angular separation of 16.1 +/- 0.1arcsec corresponds to a minimum binary separation of 1091 +/- 7au. SDSSJ1300+5904 is a cool (Teff = 6300 +/- 300K) magnetic white dwarf (B ~= 6mG). PG1258+593 is a DA white dwarf with Teff = 14790 +/- 77K and logg = 7.87 +/- 0.02. Using the white dwarf mass-radius relation implies the masses of SDSSJ1300+5904 and PG1258+593 are 0.54 +/- 0.06 and 0.54 +/- 0.01Msolar, respectively, and therefore a cooling age difference of 1.67 +/- 0.05Gyr. Adopting main-sequence lifetimes from stellar models, we derive an upper limit of 2.2Msolar for the mass of the progenitor of PG1258+593. A plausible range of initial masses is 1.4-1.8 Msolar for PG1258+593 and 2-3 Msolar for SDSSJ1300+5904. Our analysis shows that white dwarf common proper motion binaries can potentially constrain the white dwarf initial mass-final mass relation and the formation mechanism for magnetic white dwarfs. The magnetic field of SDSSJ1300+5904 is consistent with an Ap progenitor star. A common envelope origin of the system cannot be excluded, but requires a triple system as progenitor.

Expression and Cellular Distribution of Ubiquitin in Response to Injury in the Developing Spinal Cord of Monodelphis domestica

PubMed Central

Noor, Natassya M.; Møllgård, Kjeld; Wheaton, Benjamin J.; Steer, David L.; Truettner, Jessie S.; Dziegielewska, Katarzyna M.; Dietrich, W. Dalton; Smith, A. Ian; Saunders, Norman R.

2013-01-01

Ubiquitin, an 8.5 kDa protein associated with the proteasome degradation pathway has been recently identified as differentially expressed in segment of cord caudal to site of injury in developing spinal cord. Here we describe ubiquitin expression and cellular distribution in spinal cord up to postnatal day P35 in control opossums (Monodelphis domestica) and in response to complete spinal transection (T10) at P7, when axonal growth through site of injury occurs, and P28 when this is no longer possible. Cords were collected 1 or 7 days after injury, with age-matched controls and segments rostral to lesion were studied. Following spinal injury ubiquitin levels (western blotting) appeared reduced compared to controls especially one day after injury at P28. In contrast, after injury mRNA expression (qRT-PCR) was slightly increased at P7 but decreased at P28. Changes in isoelectric point of separated ubiquitin indicated possible post-translational modifications. Cellular distribution demonstrated a developmental shift between earliest (P8) and latest (P35) ages examined, from a predominantly cytoplasmic immunoreactivity to a nuclear expression; staining level and shift to nuclear staining was more pronounced following injury, except 7 days after transection at P28. After injury at P7 immunostaining increased in neurons and additionally in oligodendrocytes at P28. Mass spectrometry showed two ubiquitin bands; the heavier was identified as a fusion product, likely to be an ubiquitin precursor. Apparent changes in ubiquitin expression and cellular distribution in development and response to spinal injury suggest an intricate regulatory system that modulates these responses which, when better understood, may lead to potential therapeutic targets. PMID:23626776

Age-related changes in glial cells of dopamine midbrain subregions in rhesus monkeys.

PubMed

Kanaan, Nicholas M; Kordower, Jeffrey H; Collier, Timothy J

2010-06-01

Aging remains the strongest risk factor for developing Parkinson's disease (PD), and there is selective vulnerability in midbrain dopamine (DA) neuron degeneration in PD. By tracking normal aging-related changes with an emphasis on regional specificity, factors involved in selective vulnerability and resistance to degeneration can be studied. Towards this end, we sought to determine whether age-related changes in microglia and astrocytes in rhesus monkeys are region-specific, suggestive of involvement in regional differences in vulnerability to degeneration that may be relevant to PD pathogenesis. Gliosis in midbrain DA subregions was measured by estimating glia number using unbiased stereology, assessing fluorescence intensity for proteins upregulated during activation, and rating morphology. With normal aging, microglia exhibited increased staining intensity and a shift to more activated morphologies preferentially in the vulnerable substantia nigra-ventral tier (vtSN). Astrocytes did not exhibit age-related changes consistent with an involvement in regional vulnerability in any measure. Our results suggest advancing age is associated with chronic mild inflammation in the vtSN, which may render these DA neurons more vulnerable to degeneration. Copyright 2008 Elsevier Inc. All rights reserved.

Solution-Processed Donor-Acceptor Polymer Nanowire Network Semiconductors For High-Performance Field-Effect Transistors

PubMed Central

Lei, Yanlian; Deng, Ping; Li, Jun; Lin, Ming; Zhu, Furong; Ng, Tsz-Wai; Lee, Chun-Sing; Ong, Beng S.

2016-01-01

Organic field-effect transistors (OFETs) represent a low-cost transistor technology for creating next-generation large-area, flexible and ultra-low-cost electronics. Conjugated electron donor-acceptor (D-A) polymers have surfaced as ideal channel semiconductor candidates for OFETs. However, high-molecular weight (MW) D-A polymer semiconductors, which offer high field-effect mobility, generally suffer from processing complications due to limited solubility. Conversely, the readily soluble, low-MW D-A polymers give low mobility. We report herein a facile solution process which transformed a lower-MW, low-mobility diketopyrrolopyrrole-dithienylthieno[3,2-b]thiophene (I) into a high crystalline order and high-mobility semiconductor for OFETs applications. The process involved solution fabrication of a channel semiconductor film from a lower-MW (I) and polystyrene blends. With the help of cooperative shifting motion of polystyrene chain segments, (I) readily self-assembled and crystallized out in the polystyrene matrix as an interpenetrating, nanowire semiconductor network, providing significantly enhanced mobility (over 8 cm2V−1s−1), on/off ratio (107), and other desirable field-effect properties that meet impactful OFET application requirements. PMID:27091315

Identification by GeLC-MS/MS of trypsin inhibitor in sarcoplasmic proteins of three tropical fish and characterization of their inhibitory properties.

PubMed

Siriangkanakun, Siriphon; Li-Chan, Eunice C Y; Yongsawadigul, Jirawat

2014-07-01

Sarcoplasmic proteins from 3 fish species were fractionated by 50% to 70% ammonium sulfate precipitation. Lyophilized fractionated sarcoplasmic proteins of threadfin bream (TB-SP), bigeye snapper (BS-SP), and yellow croaker (YC-SP) showed 80% to 92% trypsin inhibitory activity. Trypsin inhibitory activity staining gel electrophoresis revealed bands at 32, 33, 37, 45, 48, and 50 kDa for the 3 species, and a band at 95 kDa was observed for TB-SP and YC-SP. Alpha-1-antitrypsin with molecular mass of 45 to 50 kDa was identified in YC-SP by gel-based liquid chromatography-tandem mass spectrometry (GeLC-MS/MS). Other major protein bands appeared on trypsin activity staining included phosphorylase, glyceraldehyde-3-phosphate dehydrogenase, and creatine kinase with molecular mass of 95 and 35 to 40 kDa, respectively. But, these 3 proteins did not show true trypsin inhibitory activity. Trypsin inhibitory activity of fractionated sarcoplasmic proteins showed good stability, with >80% activity retained at 60 °C and up to 0.6 M NaCl. TB-SP showed the highest inhibitory activity against autolysis of washed threadfin bream mince at 65 °C. Addition of 0.5% or 1% TB-SP improved textural properties of threadfin bream surimi gels preincubated at 37 or 65 °C followed by heating at 90 °C. Therefore, TB-SP could be a promising protein ingredient for enhancing surimi gel texture. Threadfin bream, bigeye snapper, and yellow croaker are the main species used as raw material for tropical surimi production. Sarcoplasmic proteins from 3 species contain trypsin inhibitor(s) that can minimize proteolytic activity and improve gel texture of proteinase-laden fish muscle. Therefore, sarcoplasmic proteins that are byproducts from surimi processing of these species could be recovered, fractionated, and utilized as a functional protein ingredient. © 2014 Institute of Food Technologists®

A Robust Two-Dimensional Separation of Intact Proteins for Bottom-Up Tandem Mass Spectrometry of the Human CSF Proteome

PubMed Central

Bora, Adriana; Anderson, Carol; Bachani, Muznabanu; Nath, Avindra; Cotter, Robert J.

2012-01-01

The cerebrospinal fluid (CSF) is produced in the brain by cells in the choroid plexus at a rate of 500mL/day. It is the only body fluid in direct contact with the brain. Thus, any changes in the CSF composition will reflect pathological processes and make CSF a potential source of biomarkers for different disease states. Proteomics offers a comprehensive view of the proteins found in CSF. In this study, we use a recently developed non-gel based method of sample preparation of CSF followed by liquid chromatography high accuracy mass spectrometry (LC-MS) for MS and MS/MS analyses, allowing unambiguous identification of peptides/proteins. Gel-eluted liquid fraction entrapment electrophoresis (Gelfree) is used to separate a CSF complex protein mixture in 12 user-selectable liquid-phase molecular weight fractions. Using this high throughput workflow we have been able to separate CSF intact proteins over a broad mass range 3.5 kDa-100 kDa with high resolution between 15 kDa and 100 kDa in 2 hours and 40 min. We have completely eliminated albumin and were able to interrogate the low abundance CSF proteins in a highly reproducible manner from different CSF samples in the same time. Using LC-MS as a downstream analysis, we identified 368 proteins using MidiTrap G-10 desalting columns and 166 proteins (including 57 unique proteins) using Zeba spin columns with 5% false discovery rate (FDR). Prostaglandin D2 synthase, Chromogranin A, Apolipoprotein E, Chromogranin B, Secretogranin III, Cystatin C, VGF nerve growth factor, Cadherin 2 are a few of the proteins that were characterized. The Gelfree-LC-MS is a robust method for the analysis of the human proteome that we will use to develop biomarkers for several neurodegenerative diseases and to quantitate these markers using multiple reaction monitoring. PMID:22537003

A microchip electrophoresis-mass spectrometric platform with double cell lysis nano-electrodes for automated single cell analysis.

PubMed

Li, Xiangtang; Zhao, Shulin; Hu, Hankun; Liu, Yi-Ming

2016-06-17

Capillary electrophoresis-based single cell analysis has become an essential approach in researches at the cellular level. However, automation of single cell analysis has been a challenge due to the difficulty to control the number of cells injected and the irreproducibility associated with cell aggregation. Herein we report the development of a new microfluidic platform deploying the double nano-electrode cell lysis technique for automated analysis of single cells with mass spectrometric detection. The proposed microfluidic chip features integration of a cell-sized high voltage zone for quick single cell lysis, a microfluidic channel for electrophoretic separation, and a nanoelectrospray emitter for ionization in MS detection. Built upon this platform, a microchip electrophoresis-mass spectrometric method (MCE-MS) has been developed for automated single cell analysis. In the method, cell introduction, cell lysis, and MCE-MS separation are computer controlled and integrated as a cycle into consecutive assays. Analysis of large numbers of individual PC-12 neuronal cells (both intact and exposed to 25mM KCl) was carried out to determine intracellular levels of dopamine (DA) and glutamic acid (Glu). It was found that DA content in PC-12 cells was higher than Glu content, and both varied from cell to cell. The ratio of intracellular DA to Glu was 4.20±0.8 (n=150). Interestingly, the ratio drastically decreased to 0.38±0.20 (n=150) after the cells are exposed to 25mM KCl for 8min, suggesting the cells released DA promptly and heavily while they released Glu at a much slower pace in response to KCl-induced depolarization. These results indicate that the proposed MCE-MS analytical platform may have a great potential in researches at the cellular level. Copyright © 2016 Elsevier B.V. All rights reserved.

Surface Induced Dissociation Coupled with High Resolution Mass Spectrometry Unveils Heterogeneity of a 211 kDa Multicopper Oxidase Protein Complex

NASA Astrophysics Data System (ADS)

Zhou, Mowei; Yan, Jing; Romano, Christine A.; Tebo, Bradley M.; Wysocki, Vicki H.; Paša-Tolić, Ljiljana

2018-01-01

Manganese oxidation is an important biogeochemical process that is largely regulated by bacteria through enzymatic reactions. However, the detailed mechanism is poorly understood due to challenges in isolating and characterizing these unknown enzymes. A manganese oxidase, Mnx, from Bacillus sp. PL-12 has been successfully overexpressed in active form as a protein complex with a molecular mass of 211 kDa. We have recently used surface induced dissociation (SID) and ion mobility-mass spectrometry (IM-MS) to release and detect folded subcomplexes for determining subunit connectivity and quaternary structure. The data from the native mass spectrometry experiments led to a plausible structural model of this multicopper oxidase, which has been difficult to study by conventional structural biology methods. It was also revealed that each Mnx subunit binds a variable number of copper ions. Becasue of the heterogeneity of the protein and limited mass resolution, ambiguities in assigning some of the observed peaks remained as a barrier to fully understanding the role of metals and potential unknown ligands in Mnx. In this study, we performed SID in a modified Fourier transform-ion cyclotron resonance (FTICR) mass spectrometer. The high mass accuracy and resolution offered by FTICR unveiled unexpected artificial modifications on the protein that had been previously thought to be iron bound species based on lower resolution spectra. Additionally, isotopically resolved spectra of the released subcomplexes revealed the metal binding stoichiometry at different structural levels. This method holds great potential for in-depth characterization of metalloproteins and protein-ligand complexes. [Figure not available: see fulltext.

Surface-assisted laser desorption ionization mass spectrometry techniques for application in forensics.

PubMed

Guinan, Taryn; Kirkbride, Paul; Pigou, Paul E; Ronci, Maurizio; Kobus, Hilton; Voelcker, Nicolas H

2015-01-01

Matrix-assisted laser desorption ionization (MALDI) mass spectrometry (MS) is an excellent analytical technique for the rapid and sensitive analysis of macromolecules (>700 Da), such as peptides, proteins, nucleic acids, and synthetic polymers. However, the detection of smaller organic molecules with masses below 700 Da using MALDI-MS is challenging due to the appearance of matrix adducts and matrix fragment peaks in the same spectral range. Recently, nanostructured substrates have been developed that facilitate matrix-free laser desorption ionization (LDI), contributing to an emerging analytical paradigm referred to as surface-assisted laser desorption ionization (SALDI) MS. Since SALDI enables the detection of small organic molecules, it is rapidly growing in popularity, including in the field of forensics. At the same time, SALDI also holds significant potential as a high throughput analytical tool in roadside, work place and athlete drug testing. In this review, we discuss recent advances in SALDI techniques such as desorption ionization on porous silicon (DIOS), nano-initiator mass spectrometry (NIMS) and nano assisted laser desorption ionization (NALDI™) and compare their strengths and weaknesses with particular focus on forensic applications. These include the detection of illicit drug molecules and their metabolites in biological matrices and small molecule detection from forensic samples including banknotes and fingerprints. Finally, the review highlights recent advances in mass spectrometry imaging (MSI) using SALDI techniques. © 2014 Wiley Periodicals, Inc.

PRESSURE SHIFT AND GRAVITATIONAL REDSHIFT OF BALMER LINES IN WHITE DWARFS: REDISCUSSION

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

Halenka, Jacek; Olchawa, Wieslaw; Madej, Jerzy

2015-08-01

The Stark-induced shift and asymmetry, the so-called pressure shift (PS) of H{sub α} and H{sub β} Balmer lines in spectra of DA white dwarfs (WDs), have been examined in detail as masking effects in measurements of the gravitational redshift in WDs. The results are compared with our earlier ones from a quarter of a century ago. In these earlier papers, the standard, symmetrical Stark line profiles, as a dominant constituent of the Balmer line profiles but shifted as a whole by the PS effect, were applied to all spectrally active layers of the WD atmosphere. At present, in each ofmore » the WD layers, the Stark line profiles (especially of H{sub β}) are inherently asymmetrical and shifted due to the effects of strong inhomogeneity of the perturbing fields in plasma. To calculate the Stark line profiles in successive layers of the WD atmosphere we used the modified Full Computer Simulation Method, able to take adequately into account the complexity of local elementary quantum processes in plasma. In the case of the H{sub α} line, the present value of Stark-induced shift of the synthetic H{sub α} line profile is about half the previous one and it is negligible in comparison with the gravitational redshift. In the case of the H{sub β} line, the present value of Stark-induced shift of the synthetic H{sub β} line profile is about twice the previous one. The source of this extra shift is the asymmetry of H{sub β} peaks.« less

Application of phase correction to improve the interpretation of crude oil spectra obtained using 7 T Fourier transform ion cyclotron resonance mass spectrometry.

PubMed

Cho, Yunju; Qi, Yulin; O'Connor, Peter B; Barrow, Mark P; Kim, Sunghwan

2014-01-01

In this study, a phase-correction technique was applied to the study of crude oil spectra obtained using a 7 T Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS). 7 T FT-ICR MS had not been widely used for oil analysis due to the lower resolving power compared with high field FT-ICR MS. For low field instruments, usage of data that has not been phase-corrected results in an inability to resolve critical mass splits of C3 and SH4 (3.4 mDa), and (13)C and CH (4.5 mDa). This results in incorrect assignments of molecular formulae, and discontinuous double bond equivalents (DBE) and carbon number distributions of S1, S2, and hydrocarbon classes are obtained. Application of phase correction to the same data, however, improves the reliability of assignments and produces continuous DBE and carbon number distributions. Therefore, this study clearly demonstrates that phase correction improves data analysis and the reliability of assignments of molecular formulae in crude oil anlayses.

Physiochemical properties and kinetics of glucoamylase produced from deoxy-d-glucose resistant mutant of Aspergillus niger for soluble starch hydrolysis.

PubMed

Riaz, Muhammad; Rashid, Muhammad Hamid; Sawyer, Lindsay; Akhtar, Saeed; Javed, Muhammad Rizwan; Nadeem, Habibullah; Wear, Martin

2012-01-01

Glucoamylases (GAs) from a wild and a deoxy-d-glucose-resistant mutant of a locally isolated Aspergillus niger were purified to apparent homogeneity. The subunit molecular mass estimated by SDS-PAGE was 93 kDa for both strains, while the molecular masses determined by MALDI-TOF for wild and mutant GAs were 72.876 and 72.063 kDa, respectively. The monomeric nature of the enzymes was confirmed through activity staining. Significant improvement was observed in the kinetic properties of the mutant GA relative to the wild type enzyme. Kinetic constants of starch hydrolysis for A. niger parent and mutant GAs calculated on the basis of molecular masses determined through MALDI-TOF were as follows: k cat = 343 and 727 s -1 , K m = 0.25 and 0.16 mg mL -1 , k cat / K m (specificity constant) = 1374 and 4510 mg mL -1 s -1 , respectively. Thermodynamic parameters for soluble starch hydrolysis also suggested that mutant GA was more efficient compared to the parent enzyme.

Platelet cytosolic 44-kDa protein is a substrate of cholera toxin-induced ADP-ribosylation and is not recognized by antisera against the. alpha. subunit of the stimulatory guanine nucleotide-binding regulatory protein

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

Molina Y Vedia, L.M.; Reep, B.R.; Lapetina, E.G.

1988-08-01

ADP-ribosylation induced by cholera toxin and pertussis toxin was studied in particulate and cytosolic fractions of human platelets. Platelets were disrupted by a cycle of freezing and thawing in the presence of a hyposmotic buffer containing protease inhibitors. In both fractions, the A subunit of cholera toxin ADP-ribosylates two proteins with molecular masses of 42 and 44 kDa, whereas pertussis toxin ADP-ribosylates a 41-kDa polypeptide. Two antisera against the {alpha} subunit of the stimulatory guanine nucleotide-binding regulatory protein recognize only the 42-kDa polypeptide. Cholera toxin-induced ADP-ribosylation of the 42- and 44-kDa proteins is reduced by pretreatment of platelets with iloprost,more » a prostacyclin analog. The 44-kDa protein, which is substrate of cholera toxin, could be extracted completely from the membrane and recovered in the cytosolic fraction when the cells were disrupted by Dounce homogenization and the pellet was extensively washed. A 44-kDa protein can also be labeled with 8-azidoguanosine 5{prime}-({alpha}-{sup 32}P)triphosphate in the cytosol and membranes. These finding indicate that cholera and pertussis toxins produced covalent modifications of proteins present in particulate and cytosolic platelet fractions. Moreover, the 44-kDa protein might be an {alpha} subunit of a guanine nucleotide-binding regulatory protein that is not recognized by available antisera.« less

Charge Exchange Reaction in Dopant-Assisted Atmospheric Pressure Chemical Ionization and Atmospheric Pressure Photoionization.

PubMed

Vaikkinen, Anu; Kauppila, Tiina J; Kostiainen, Risto

2016-08-01

The efficiencies of charge exchange reaction in dopant-assisted atmospheric pressure chemical ionization (DA-APCI) and dopant-assisted atmospheric pressure photoionization (DA-APPI) mass spectrometry (MS) were compared by flow injection analysis. Fourteen individual compounds and a commercial mixture of 16 polycyclic aromatic hydrocarbons were chosen as model analytes to cover a wide range of polarities, gas-phase ionization energies, and proton affinities. Chlorobenzene was used as the dopant, and methanol/water (80/20) as the solvent. In both techniques, analytes formed the same ions (radical cations, protonated molecules, and/or fragments). However, in DA-APCI, the relative efficiency of charge exchange versus proton transfer was lower than in DA-APPI. This is suggested to be because in DA-APCI both dopant and solvent clusters can be ionized, and the formed reagent ions can react with the analytes via competing charge exchange and proton transfer reactions. In DA-APPI, on the other hand, the main reagents are dopant-derived radical cations, which favor ionization of analytes via charge exchange. The efficiency of charge exchange in both DA-APPI and DA-APCI was shown to depend heavily on the solvent flow rate, with best efficiency seen at lowest flow rates studied (0.05 and 0.1 mL/min). Both DA-APCI and DA-APPI showed the radical cation of chlorobenzene at 0.05-0.1 mL/min flow rate, but at increasing flow rate, the abundance of chlorobenzene M(+.) decreased and reagent ion populations deriving from different gas-phase chemistry were recorded. The formation of these reagent ions explains the decreasing ionization efficiency and the differences in charge exchange between the techniques. Graphical Abstract ᅟ.

Partially Reduced Graphene Oxide Modified Tetrahedral Amorphous Carbon Thin-Film Electrodes as a Platform for Nanomolar Detection of Dopamine

DOE PAGES

Wester, Niklas; Sainio, Sami; Palomäki, Tommi; ...

2017-03-16

Here, we present for the first time tetrahedral amorphous carbon (ta-C)—a partially reduced graphene oxide (PRGO) hybrid electrode nanomaterial platform for electrochemical sensing of dopamine (DA). Graphene oxide was synthesized with the modified Hummer’s method. Before modification of ta-C by drop casting, partial reduction of the GO was carried out to improve electrochemical properties and adhesion to the ta-C thin film. A facile nitric acid treatment that slightly reoxidized the surface and modified the surface chemistry was subsequently performed to further improve the electrochemical properties of the electrodes. The largest relative increase was seen in carboxyl groups. The HNO 3more » treatment increased the sensitivity toward DA and AA and resulted in a cathodic shift in the oxidation of AA. The fabricated hybrid electrodes were characterized with scanning electron microscopy (SEM), Raman spectroscopy, Fourier transform infrared spectroscopy (FT-IR), X-ray photoelectron spectroscopy (XPS), X-ray absorption spectroscopy (XAS), and electrochemical impedance spectroscopy (EIS). Moreover, compared to the plain ta-C electrode the hybrid electrode was shown to exhibit superior sensitivity and selectivity toward DA in the presence of ascorbic acid (AA), enabling simultaneous sensing of AA and DA close to the physiological concentrations by cyclic voltammetry (CV) and by differential pulse voltammetry (DPV). Two linear ranges of 0–1 μM and 1–100 μM and a detection limit (S/N = 3.3) of 2.6 nM for DA were determined by means of cyclic voltammetry. Thus, the current work provides a fully CMOS-compatible carbon based hybrid nanomaterial that shows potential for in vivo measurements of DA.« less

Partially Reduced Graphene Oxide Modified Tetrahedral Amorphous Carbon Thin-Film Electrodes as a Platform for Nanomolar Detection of Dopamine

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

Wester, Niklas; Sainio, Sami; Palomäki, Tommi

Here, we present for the first time tetrahedral amorphous carbon (ta-C)—a partially reduced graphene oxide (PRGO) hybrid electrode nanomaterial platform for electrochemical sensing of dopamine (DA). Graphene oxide was synthesized with the modified Hummer’s method. Before modification of ta-C by drop casting, partial reduction of the GO was carried out to improve electrochemical properties and adhesion to the ta-C thin film. A facile nitric acid treatment that slightly reoxidized the surface and modified the surface chemistry was subsequently performed to further improve the electrochemical properties of the electrodes. The largest relative increase was seen in carboxyl groups. The HNO 3more » treatment increased the sensitivity toward DA and AA and resulted in a cathodic shift in the oxidation of AA. The fabricated hybrid electrodes were characterized with scanning electron microscopy (SEM), Raman spectroscopy, Fourier transform infrared spectroscopy (FT-IR), X-ray photoelectron spectroscopy (XPS), X-ray absorption spectroscopy (XAS), and electrochemical impedance spectroscopy (EIS). Moreover, compared to the plain ta-C electrode the hybrid electrode was shown to exhibit superior sensitivity and selectivity toward DA in the presence of ascorbic acid (AA), enabling simultaneous sensing of AA and DA close to the physiological concentrations by cyclic voltammetry (CV) and by differential pulse voltammetry (DPV). Two linear ranges of 0–1 μM and 1–100 μM and a detection limit (S/N = 3.3) of 2.6 nM for DA were determined by means of cyclic voltammetry. Thus, the current work provides a fully CMOS-compatible carbon based hybrid nanomaterial that shows potential for in vivo measurements of DA.« less

Design, biometric simulation and optimization of a nano-enabled scaffold device for enhanced delivery of dopamine to the brain.

PubMed

Pillay, Samantha; Pillay, Viness; Choonara, Yahya E; Naidoo, Dinesh; Khan, Riaz A; du Toit, Lisa C; Ndesendo, Valence M K; Modi, Girish; Danckwerts, Michael P; Iyuke, Sunny E

2009-12-01

This study focused on the design, biometric simulation and optimization of an intracranial nano-enabled scaffold device (NESD) for the site-specific delivery of dopamine (DA) as a strategy to minimize the peripheral side-effects of conventional forms of Parkinson's disease therapy. The NESD was modulated through biometric simulation and computational prototyping to produce a binary crosslinked alginate scaffold embedding stable DA-loaded cellulose acetate phthalate (CAP) nanoparticles optimized in accordance with Box-Behnken statistical designs. The physicomechanical properties of the NESD were characterized and in vitro and in vivo release studies performed. Prototyping predicted a 3D NESD model with enhanced internal micro-architecture. SEM and TEM revealed spherical, uniform and non-aggregated DA-loaded nanoparticles with the presence of CAP (FTIR bands at 1070, 1242 and 2926 cm(-1)). An optimum nanoparticle size of 197 nm (PdI=0.03), a zeta potential of -34.00 mV and a DEE of 63% was obtained. The secondary crosslinker BaCl(2) imparted crystallinity resulting in significant thermal shifts between native CAP (T(g)=160-170 degrees C; T(m)=192 degrees C) and CAP nanoparticles (T(g)=260 degrees C; T(m)=268 degrees C). DA release displayed an initial lag phase of 24 h and peaked after 3 days, maintaining favorable CSF (10 microg/mL) versus systemic concentrations (1-2 microg/mL) over 30 days and above the inherent baseline concentration of DA (1 microg/mL) following implantation in the parenchyma of the frontal lobe of the Sprague-Dawley rat model. The strategy of coupling polymeric scaffold science and nanotechnology enhanced the site-specific delivery of DA from the NESD.

Tryptic digestion of human GPIIIa. Isolation and biochemical characterization of the 23 kDa N-terminal glycopeptide carrying the antigenic determinant for a monoclonal antibody (P37) which inhibits platelet aggregation.

PubMed Central

Calvete, J J; Rivas, G; Maruri, M; Alvarez, M V; McGregor, J L; Hew, C L; Gonzalez-Rodriguez, J

1988-01-01

Early digestion of pure human platelet glycoprotein IIIa (GPIIIa) leads to a single cleavage of the molecule at 23 kDa far from one of the terminal amino acids. Automated Edman degradation demonstrates that GPIIIa and the smaller (23 kDa) tryptic fragment share the same N-terminal amino acid sequence. A further cleavage occurs in the larger fragment (80 kDa), reducing its apparent molecular mass by 10 kDa. The 23 kDa fragment remains attached to the larger ones in unreduced samples. Stepwise reduction of early digested GPIIIa with dithioerythritol selectively reduces the single disulphide bond joining the smaller (23 kDa) to the larger (80/70 kDa) fragments. Two fractions were obtained by size-exclusion chromatography of early digested GPIIIa after partial or full reduction and alkylation. The larger-size fraction contains the 80/70 kDa fragments, while the 23 kDa fragment is isolated in the smaller. The amino acid compositions of these fractions do not differ very significantly from the composition of GPIIIa; however the 23 kDa fragment contains only 10.2% by weight of sugars and is richer in neuraminic acid. Disulphide bonds are distributed four in the 23 kDa glycopeptide and 20-21 in the 80/70 kDa glycopeptide. The epitope for P37, a monoclonal antibody which inhibits platelet aggregation [Melero & González-Rodríguez (1984) Eur. J. Biochem. 141, 421-427] is situated within the first 17 kDa of the N-terminal region of GPIIIa, which gives a special functional interest to this extracellular region of GPIIIa. On the other hand, the epitopes for GPIIIa-specific monoclonal antibodies, P6, P35, P40 and P97, which do not interfere with platelet aggregation, are located within the larger tryptic fragment (80/70 kDa). Thus, the antigenic areas available in the extracellular surface of GPIIIa for these five monoclonal antibodies are now more precisely delineated. Images Fig. 1. Fig. 2. Fig. 3. Fig. 4. PMID:2455507

Rho0 production and possible modification in Au+Au and p+p collisions at square root [sNN] = 200 GeV.

PubMed

Adams, J; Adler, C; Aggarwal, M M; Ahammed, Z; Amonett, J; Anderson, B D; Arkhipkin, D; Averichev, G S; Badyal, S K; Balewski, J; Barannikova, O; Barnby, L S; Baudot, J; Bekele, S; Belaga, V V; Bellwied, R; Berger, J; Bezverkhny, B I; Bhardwaj, S; Bhati, A K; Bichsel, H; Billmeier, A; Bland, L C; Blyth, C O; Bonner, B E; Botje, M; Boucham, A; Brandin, A; Bravar, A; Cadman, R V; Cai, X Z; Caines, H; Calderón de la Barca Sánchez, M; Carroll, J; Castillo, J; Cebra, D; Chaloupka, P; Chattopadhyay, S; Chen, H F; Chen, Y; Chernenko, S P; Cherney, M; Chikanian, A; Christie, W; Coffin, J P; Cormier, T M; Cramer, J G; Crawford, H J; Das, D; Das, S; Derevschikov, A A; Didenko, L; Dietel, T; Dong, W J; Dong, X; Draper, J E; Du, F; Dubey, A K; Dunin, V B; Dunlop, J C; Dutta Majumdar, M R; Eckardt, V; Efimov, L G; Emelianov, V; Engelage, J; Eppley, G; Erazmus, B; Estienne, M; Fachini, P; Faine, V; Faivre, J; Fatemi, R; Filimonov, K; Filip, P; Finch, E; Fisyak, Y; Flierl, D; Foley, K J; Fu, J; Gagliardi, C A; Gagunashvili, N; Gans, J; Ganti, M S; Gaudichet, L; Geurts, F; Ghazikhanian, V; Ghosh, P; Gonzalez, J E; Grachov, O; Grebenyuk, O; Gronstal, S; Grosnick, D; Guertin, S M; Gupta, A; Gutierrez, T D; Hallman, T J; Hamed, A; Hardtke, D; Harris, J W; Heinz, M; Henry, T W; Heppelmann, S; Hippolyte, B; Hirsch, A; Hjort, E; Hoffmann, G W; Horsley, M; Huang, H Z; Huang, S L; Hughes, E; Humanic, T J; Igo, G; Ishihara, A; Jacobs, P; Jacobs, W W; Janik, M; Jiang, H; Johnson, I; Jones, P G; Judd, E G; Kabana, S; Kaplan, M; Keane, D; Khodyrev, V Yu; Kiryluk, J; Kisiel, A; Klay, J; Klein, S R; Klyachko, A; Koetke, D D; Kollegger, T; Kopytine, M; Kotchenda, L; Kovalenko, A D; Kramer, M; Kravtsov, P; Kravtsov, V I; Krueger, K; Kuhn, C; Kulikov, A I; Kumar, A; Kunde, G J; Kunz, C L; Kutuev, R Kh; Kuznetsov, A A; Lamont, M A C; Landgraf, J M; Lange, S; Lasiuk, B; Laue, F; Lauret, J; Lebedev, A; Lednický, R; LeVine, M J; Li, C; Li, Q; Lindenbaum, S J; Lisa, M A; Liu, F; Liu, L; Liu, Z; Liu, Q J; Ljubicic, T; Llope, W J; Long, H; Longacre, R S; Lopez-Noriega, M; Love, W A; Ludlam, T; Lynn, D; Ma, J; Ma, Y G; Magestro, D; Mahajan, S; Mangotra, L K; Mahapatra, D P; Majka, R; Manweiler, R; Margetis, S; Markert, C; Martin, L; Marx, J; Matis, H S; Matulenko, Yu A; McClain, C J; McShane, T S; Meissner, F; Melnick, Yu; Meschanin, A; Miller, M L; Milosevich, Z; Minaev, N G; Mironov, C; Mischke, A; Mishra, D; Mitchell, J; Mohanty, B; Molnar, L; Moore, C F; Mora-Corral, M J; Morozov, D A; Morozov, V; De Moura, M M; Munhoz, M G; Nandi, B K; Nayak, S K; Nayak, T K; Nelson, J M; Netrakanti, P K; Nikitin, V A; Nogach, L V; Norman, B; Nurushev, S B; Odyniec, G; Ogawa, A; Okorokov, V; Oldenburg, M; Olson, D; Paic, G; Pal, S K; Panebratsev, Y; Panitkin, S Y; Pavlinov, A I; Pawlak, T; Peitzmann, T; Perevoztchikov, V; Perkins, C; Peryt, W; Petrov, V A; Phatak, S C; Picha, R; Planinic, M; Pluta, J; Porile, N; Porter, J; Poskanzer, A M; Potekhin, M; Potrebenikova, E; Potukuchi, B V K S; Prindle, D; Pruneau, C; Putschke, J; Rai, G; Rakness, G; Raniwala, R; Raniwala, S; Ravel, O; Ray, R L; Razin, S V; Reichhold, D; Reid, J G; Renault, G; Retiere, F; Ridiger, A; Ritter, H G; Roberts, J B; Rogachevski, O V; Romero, J L; Rose, A; Roy, C; Ruan, L J; Sahoo, R; Sakrejda, I; Salur, S; Sandweiss, J; Savin, I; Schambach, J; Scharenberg, R P; Schmitz, N; Schroeder, L S; Schweda, K; Seger, J; Seyboth, P; Shahaliev, E; Shao, M; Shao, W; Sharma, M; Shestermanov, K E; Shimanskii, S S; Singaraju, R N; Simon, F; Skoro, G; Smirnov, N; Snellings, R; Sood, G; Sorensen, P; Sowinski, J; Speltz, J; Spinka, H M; Srivastava, B; Stanislaus, T D S; Stock, R; Stolpovsky, A; Strikhanov, M; Stringfellow, B; Struck, C; Suaide, A A P; Sugarbaker, E; Suire, C; Sumbera, M; Surrow, B; Symons, T J M; Szanto de Toledo, A; Szarwas, P; Tai, A; Takahashi, J; Tang, A H; Thein, D; Thomas, J H; Timoshenko, S; Tokarev, M; Tonjes, M B; Trainor, T A; Trentalange, S; Tribble, R E; Tsai, O; Ullrich, T; Underwood, D G; Van Buren, G; VanderMolen, A M; Varma, R; Vasilevski, I; Vasiliev, A N; Vernet, R; Vigdor, S E; Viyogi, Y P; Voloshin, S A; Vznuzdaev, M; Waggoner, W; Wang, F; Wang, G; Wang, G; Wang, X L; Wang, Y; Wang, Z M; Ward, H; Watson, J W; Webb, J C; Wells, R; Westfall, G D; Whitten, C; Wieman, H; Willson, R; Wissink, S W; Witt, R; Wood, J; Wu, J; Xu, N; Xu, Z; Xu, Z Z; Yamamoto, E; Yepes, P; Yurevich, V I; Yuting, B; Zanevski, Y V; Zhang, H; Zhang, W M; Zhang, Z P; Zhaomin, Z P; Zizong, Z P; Zołnierczuk, P A; Zoulkarneev, R; Zoulkarneeva, J; Zubarev, A N

2004-03-05

We report results on rho(770)(0)-->pi(+)pi(-) production at midrapidity in p+p and peripheral Au+Au collisions at sqrt[s(NN)]=200 GeV. This is the first direct measurement of rho(770)(0)-->pi(+)pi(-) in heavy-ion collisions. The measured rho(0) peak in the invariant mass distribution is shifted by approximately 40 MeV/c(2) in minimum bias p+p interactions and approximately 70 MeV/c(2) in peripheral Au+Au collisions. The rho(0) mass shift is dependent on transverse momentum and multiplicity. The modification of the rho(0) meson mass, width, and shape due to phase space and dynamical effects are discussed.

Upregulation of the β-form of 14-3-3 protein in telencephalon of goldfish (Carassius auratus): its possible role in spatial learning.

PubMed

Subramanian, Dharaneedharan; Ramalingam, Rajkumar; Karuppasamy, Radhakrishnan; Subramanian, Thanga Leela; Chellam, Balasundaram; Rajan, Koilmani Emmanuvel

2012-10-03

In the present study, we observed variations in the expression pattern of proteins isolated from the telencephalon of goldfish (Carassius auratus). The expression of a 28 kDa protein was elevated in the individuals trained in a spatial task when compared with the untrained individuals. The ∼28 kDa protein was analyzed using liquid chromatography and mass spectrometry; further, the data were analyzed using the MASCOT search engine. The analysis showed that the ∼28 kDa protein is a β form of 14-3-3 protein with 35.1% identity. In addition, the semiquantitative PCR confirmed the variation in the expression of 14-3-3 between the trained and the untrained groups. Subsequently, we examined the effect of upregulation of 14-3-3 (β) in the neurotransmitters; that is, serotonin (5-hydroxytryptamine, 5-HT) and dopamine (DA). Notably, the level of 5-HT and DA was found to be significantly elevated in the telencephalon of individuals trained in the spatial task than in the untrained individuals. Our results suggest that the spatial learning increases the expression of 14-3-3 (β), which in turn leads to an increase in the level of 5-HT and DA. The upregulated 5-HT and DA may facilitate synapse formation during spatial learning in a novel environment.

High Molecular Weight Isoforms of Growth Hormone In Cells of the Immune System

PubMed Central

Weigent, Douglas A.

2013-01-01

A substantial body of research exists to support the idea that cells of the immune system produce growth hormone (GH). However, the structure and mechanism of action of lymphocyte-derived GH continues to remain largely unknown. Here we present the results of Western analysis of whole cell extracts showing that different molecular weight isoforms of GH of approximately 100 kDa, 65 kDa, and 48 kDa can be detected in primary mouse cells of the immune system and in the mouse EL4 cell line. The identity of the 65 kDa and 48 kDa isoforms of GH were confirmed by mass spectrometry. The various isoforms were detected in both enriched T and B spleen cell populations. The large molecular weight isoform appears to reside primarily in the cytoplasm whereas the lower molecular weight 65 kDa and 48 kDa isoforms were detected primarily in the nucleus. These results also suggest that GH isoforms are induced by oxidative stress. In EL4 cells overexpressing GH, the expression of luciferase controlled by a promoter containing the antioxidant response element is increased almost three-fold above control. The data suggest that the induction of isoforms of the GH molecule in cells of the immune system may be an important mechanism of adaptation and/or protection of lymphoid cells under conditions of oxidative stress. PMID:21741628

Extracting the mass dependence and quantum numbers of short-range correlated pairs from A ( e , e ' p ) and A ( e , e ' p p ) scattering

DOE PAGES

Colle, C.; Hen, O.; Cosyn, W.; ...

2015-08-06

Tmore » he addition of Molybdenum was used to modify the nanostructure and enhance coercivity of rare-earth-free Zr 2Co 11-based nanocrystalline permanent magnets. he effect of Mo addition on magnetic domain structures of melt spun nanocrystalline Zr 16Co 84-xMo x( x = 0 , 0.5, 1, 1.5, and 2.0) ribbons has been investigated. It was found that magnetic properties and local domain structures are strongly influenced by Mo doping. he coercivity of the samples increases with the increase in Mo content ( x ≤ 1.5 ). he maximum energy product ( B H ) max increases with increasing x from 0.5 MGOe for x = 0 to a maximum value of 4.2 MGOe for x = 1.5 . he smallest domain size with a relatively short magnetic correlation length of 128 nm and largest root-mean-square phase shift Φ rms value of 0.66° are observed for the x = 1.5 . he optimal Mo addition promotes magnetic domain structure refinement and thus leads to a significant increase in coercivity and energy product in this sample. MDH is primarily a dimer in solution, although an oligomeric species with a molecular mass of ~450–560 kDa forms at higher protein concentrations. A new measurement of the rare, doubly radiative decay eta->pi 0 γγ was conducted with the Crystal Ball and APS multiphoton spectrometers together with the photon tagging facility at the Mainz Microtron MAMI.« less

Extracting the mass dependence and quantum numbers of short-range correlated pairs from A ( e , e ' p ) and A ( e , e ' p p ) scattering

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

Colle, C.; Hen, O.; Cosyn, W.

Tmore » he addition of Molybdenum was used to modify the nanostructure and enhance coercivity of rare-earth-free Zr 2Co 11-based nanocrystalline permanent magnets. he effect of Mo addition on magnetic domain structures of melt spun nanocrystalline Zr 16Co 84-xMo x( x = 0 , 0.5, 1, 1.5, and 2.0) ribbons has been investigated. It was found that magnetic properties and local domain structures are strongly influenced by Mo doping. he coercivity of the samples increases with the increase in Mo content ( x ≤ 1.5 ). he maximum energy product ( B H ) max increases with increasing x from 0.5 MGOe for x = 0 to a maximum value of 4.2 MGOe for x = 1.5 . he smallest domain size with a relatively short magnetic correlation length of 128 nm and largest root-mean-square phase shift Φ rms value of 0.66° are observed for the x = 1.5 . he optimal Mo addition promotes magnetic domain structure refinement and thus leads to a significant increase in coercivity and energy product in this sample. MDH is primarily a dimer in solution, although an oligomeric species with a molecular mass of ~450–560 kDa forms at higher protein concentrations. A new measurement of the rare, doubly radiative decay eta->pi 0 γγ was conducted with the Crystal Ball and APS multiphoton spectrometers together with the photon tagging facility at the Mainz Microtron MAMI.« less

Biochemical and mass spectrometric characterization of the human CB2 cannabinoid receptor expressed in Pichia pastoris--importance of correct processing of the N-terminus.

PubMed

Zhang, Rundong; Kim, Tae-Kang; Qiao, Zhaun-Hong; Cai, Jian; Pierce, William M; Song, Zhao-Hui

2007-10-01

This study was conducted to optimize the expression of human CB2 cannabinoid receptors in methylotrophic yeast Pichia pastoris (P. pastoris). Two major species of expressed CB2 proteins were seen on Western blot, i.e., a 42 kDa band which matches the calculated molecular weight for tagged CB2, and a 52/55 kDa doublet. Treatment of membranes with N-glycosidase F or inclusion of tunicamycin in the culture medium during induction resulted in the disappearance of the 55 kDa, but not the 52 kDa band, suggesting that the 3 kDa extra in the 55 kDa band is due to N-glycosylation, but the 10 kDa extra in the 52 kDa band is not due to N-glycosylation. Anti-FLAG M1 antibody had a much higher preference for the 42 kDa band over the 52/55 kDa doublet, and a 10 kDa fragment recognized by anti-FLAG M2 antibody was generated by CNBr digestion of the 52/55 doublet. These data strongly support the hypothesis that the 10 kDa increase in molecular weight was due to unprocessed alpha-factor sequence. This conclusion was further validated by finding several peptide sequences for alpha-factor fragments at the N-terminal of the CB2 receptor using pepsin/chymotrypsin digestion and LC/MS/MS approaches. Importantly, unprocessed alpha-factor was found to be associated with poor ligand binding. In addition, controlling the level of CB2 protein expression was found to be critical for minimizing the presence of unprocessed alpha-factor sequence. The information gained from this study should aid the proper expression of not only CB2 receptor but also other members of the GPCR family in P. pastoris.

Density and spin modes in imbalanced normal Fermi gases from collisionless to hydrodynamic regime

NASA Astrophysics Data System (ADS)

Narushima, Masato; Watabe, Shohei; Nikuni, Tetsuro

2018-03-01

We study the mass- and population-imbalance effect on density (in-phase) and spin (out-of-phase) collective modes in a two-component normal Fermi gas. By calculating the eigenmodes of the linearized Boltzmann equation as well as the density/spin dynamic structure factor, we show that mass- and population-imbalance effects offer a variety of collective mode crossover behaviors from collisionless to hydrodynamic regimes. The mass-imbalance effect shifts the crossover regime to the higher-temperature, and a significant peak of the spin dynamic structure factor emerges only in the collisionless regime. This is in contrast to the case of mass- and population-balanced normal Fermi gases, where the spin dynamic response is always absent. Although the population-imbalance effect does not shift the crossover regime, the spin dynamic structure factor survives both in the collisionless and hydrodynamic regimes.

Shift work schedule and night work load: Effects on body mass index - a four-year longitudinal study.

PubMed

Buchvold, Hogne Vikanes; Pallesen, Ståle; Waage, Siri; Bjorvatn, Bjørn

2018-05-01

Objectives The aim of this study was to investigate changes in body mass index (BMI) between different work schedules and different average number of yearly night shifts over a four-year follow-up period. Methods A prospective study of Norwegian nurses (N=2965) with different work schedules was conducted: day only, two-shift rotation (day and evening shifts), three-shift rotation (day, evening and night shifts), night only, those who changed towards night shifts, and those who changed away from schedules containing night shifts. Paired student's t-tests were used to evaluate within subgroup changes in BMI. Multiple linear regression analysis was used to evaluate between groups effects on BMI when adjusting for BMI at baseline, sex, age, marital status, children living at home, and years since graduation. The same regression model was used to evaluate the effect of average number of yearly night shifts on BMI change. Results We found that night workers [mean difference (MD) 1.30 (95% CI 0.70-1.90)], two shift workers [MD 0.48 (95% CI 0.20-0.75)], three shift workers [MD 0.46 (95% CI 0.30-0.62)], and those who changed work schedule away from [MD 0.57 (95% CI 0.17-0.84)] or towards night work [MD 0.63 (95% CI 0.20-1.05)] all had significant BMI gain (P<0.01) during the follow-up period. However, day workers had a non-significant BMI gain. Using adjusted multiple linear regressions, we found that night workers had significantly larger BMI gain compared to day workers [B=0.89 (95% CI 0.06-1.72), P<0.05]. We did not find any significant association between average number of yearly night shifts and BMI change using our multiple linear regression model. Conclusions After adjusting for possible confounders, we found that BMI increased significantly more among night workers compared to day workers.

Mass transit ridership and self-reported hearing health in an urban population.

PubMed

Gershon, Robyn R M; Sherman, Martin F; Magda, Lori A; Riley, Halley E; McAlexander, Tara P; Neitzel, Richard

2013-04-01

Information on prevalence and risk factors associated with self-reported hearing health among mass transit riders is extremely limited, even though evidence suggests mass transit may be a source of excessive exposure to noise. Data on mass transit ridership were collected from 756 study participants using a self-administered questionnaire. Hearing health was measured using two symptom items (tinnitus and temporary audiometric threshold shift), two subjective measures (self-rated hearing and hearing ability), and two medical-related questions (hearing testing and physician-diagnosed hearing loss). In logistic regression analyses that controlled for possible confounders, including demographic variables, occupational noise exposure, nonoccupational noise exposure (including MP3 player use) and use of hearing protection, frequent and lengthy mass transit (all forms) ridership (1,100 min or more per week vs. 350 min or less per week) was the strongest predictor of temporary threshold shift symptoms. Noise abatement strategies, such as engineering controls, and the promotion of hearing protection use should be encouraged to reduce the risk of adverse impacts on the hearing health of mass transit users.

Activity Theory and the Transformation of Pedagogic Practice

ERIC Educational Resources Information Center

Yamazumi, Katsuhiro

2006-01-01

Today, work and other societal practices are experiencing accelerating paradigm shifts from mass-production-based systems toward new systems based on networking between organizations, collaboration, and partnerships. This shift requires new paradigms in the fields of education, learning, and development. As human activity quickly changes to…

Radiation inactivation method provides evidence that membrane-bound mitochondrial creatine kinase is an oligomer

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

Quemeneur, E.; Eichenberger, D.; Goldschmidt, D.

1988-06-30

Lyophilized suspensions of rabbit heart mitochondria have been irradiated with varying doses of gamma rays. Mitochondrial creatine kinase activity was inactivated exponentially with a radiation inactivation size of 352 or 377 kDa depending upon the initial medium. These values are in good agreement with the molecular mass previously deduced from by permeation experiments: 357 kDa. This is the first direct evidence showing that the native form of mitochondrial creatine kinase is associated to the inner membrane as an oligomer, very likely an octamer.

Intact and Top-Down Characterization of Biomolecules and Direct Analysis Using Infrared Matrix-Assisted Laser Desorption Electrospray Ionization Coupled to FT-ICR Mass Spectrometry

PubMed Central

Sampson, Jason S.; Murray, Kermit K.; Muddiman, David C.

2013-01-01

We report the implementation of an infrared laser onto our previously reported matrix-assisted laser desorption electrospray ionization (MALDESI) source with ESI post-ionization yielding multiply charged peptides and proteins. Infrared (IR)-MALDESI is demonstrated for atmospheric pressure desorption and ionization of biological molecules ranging in molecular weight from 1.2 to 17 kDa. High resolving power, high mass accuracy single-acquisition Fourier transform ion cyclotron resonance (FT-ICR) mass spectra were generated from liquid-and solid-state peptide and protein samples by desorption with an infrared laser (2.94 µm) followed by ESI post-ionization. Intact and top-down analysis of equine myoglobin (17 kDa) desorbed from the solid state with ESI post-ionization demonstrates the sequencing capabilities using IR-MALDESI coupled to FT-ICR mass spectrometry. Carbohydrates and lipids were detected through direct analysis of milk and egg yolk using both UV- and IR-MALDESI with minimal sample preparation. Three of the four classes of biological macromolecules (proteins, carbohydrates, and lipids) have been ionized and detected using MALDESI with minimal sample preparation. Sequencing of O-linked glycans, cleaved from mucin using reductive β-elimination chemistry, is also demonstrated. PMID:19185512

Does testis weight decline towards the Subarctic? A case study on the common frog, Rana temporaria

NASA Astrophysics Data System (ADS)

Hettyey, Attila; Laurila, Anssi; Herczeg, Gábor; Jönsson, K. Ingemar; Kovács, Tibor; Merilä, Juha

2005-04-01

Interpopulation comparisons of variation in resource availability and in allocation patterns along altitudinal and latitudinal gradients allow insights into the mechanisms shaping the life history of animals. Patterns of between-population differences in female life history traits have been studied intensively across a wide range of taxa, but similar investigations in males have remained scarce. To study if testis weight—a measure of reproductive investment—varies on a geographical scale in anurans, we focussed on the variation in relative testis weight (RelTW) and asymmetry in 22 populations of the common frog Rana temporaria along a 1,600-km latitudinal transect across the Scandinavian peninsula. We found that RelTW decreased towards the north. Body mass and body length both had independent positive effects on testes mass. We found evidence for directional asymmetry (DA) in testis weight with the right testis being larger than the left. The level of DA in testis weight was not related to latitude, but both body mass and testes mass had independent positive effects on asymmetry. We discuss the northwards decrease in RelTW in terms of a decreased reproductive investment as a possible consequence of harsher environmental conditions, and perhaps also, weaker sexual selection in the north than in the south.

MALDI mass spectrometry of biomolecules and synthetic polymers using alkali hexacyanoferrate (II) complexes and glycerol as matrix

NASA Astrophysics Data System (ADS)

Zollner, Peter; Stubiger, Gerald; Schmid, Erich; Pittenauer, Ernst; Allmaier, Gunter

1997-12-01

K4[Fe(CN)6]/glycerol and Na4[Fe(CN)6]/glycerol have been investigated as liquid matrix systems for UV-MALDI MS applying a N2 laser. Analyte molecules were detected as sodium or potassium adduct ions and, in the case of proteins, as well as protonated molecular ions. Mass accuracies were comparable to those found with standard solid matrix systems with -0.06 to +0.05% deviation in the reflectron mode and with -0.24 to +0.13% in the linear mode. Useful results could be obtained within a mass range of 15 000 Da for single-charged proteins and 8000 Da for potassium cationized polyethylene glycols. Detection limits were found for hydrophilic compounds in the low picomol range and for lipophilic compounds as triacylglycerols or peracetylated and partially benzylated carbohydrates in the low femtomol range. As shown by scanning electron microscopic investigations, the generation of a thin homogenous matrix layer was essential for a successful mass spectrometric experiment. A very careful cleaning of the target surface with glacial acid prior to matrix deposition improved the formation of such a matrix film that maximum sensitivity as well as good reproducibility of the experiments could be achieved.

Purification and characterization of a soluble glycoprotein from garlic (Allium sativum) and its in vitro bioactivity.

PubMed

Wang, Yan; Zou, Tingting; Xiang, Minghui; Jin, Chenzhong; Zhang, Xuejiao; Chen, Yong; Jiang, Qiuqing; Hu, Yihong

2016-10-02

A soluble glycoprotein was purified to homogeneity from ripe garlic (Allium sativum) bulbs using ammonium sulfate precipitation, Sephadex G-100 gel filtration, and diethylaminoethyl-52 cellulose anion-exchange chromatography. A native mass of 55.7 kDa estimated on gel permeation chromatography and a molecular weight of 13.2 kDa observed on sodium dodecyl sulfate-polyacrylamide gel electrophoresis supported that the glycoprotein is a homotetramer. β-Elimination reaction result suggested that the glycoprotein is an N-linked type. Fourier-transform infrared spectroscopy proved that it contains sugar. Gas chromatography-mass spectrometer analysis showed that its sugar component was galactose. The glycoprotein has 1,1-diphenyl-2-picrylhydrazil free radical scavenging activity and the peroxidation inhibition ability to polyunsaturated fatty acid. These results indicated that the glycoprotein has potential for food additives, functional foods, and even biotechnological and medical applications.

Mass spectrometry for the biophysical characterization of therapeutic monoclonal antibodies.

PubMed

Zhang, Hao; Cui, Weidong; Gross, Michael L

2014-01-21

Monoclonal antibodies (mAbs) are powerful therapeutics, and their characterization has drawn considerable attention and urgency. Unlike small-molecule drugs (150-600 Da) that have rigid structures, mAbs (∼150 kDa) are engineered proteins that undergo complicated folding and can exist in a number of low-energy structures, posing a challenge for traditional methods in structural biology. Mass spectrometry (MS)-based biophysical characterization approaches can provide structural information, bringing high sensitivity, fast turnaround, and small sample consumption. This review outlines various MS-based strategies for protein biophysical characterization and then reviews how these strategies provide structural information of mAbs at the protein level (intact or top-down approaches), peptide, and residue level (bottom-up approaches), affording information on higher order structure, aggregation, and the nature of antibody complexes. Copyright © 2013 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

Purification and characterization of a new type lactose-binding Ulex europaeus lectin by affinity chromatography.

PubMed

Konami, Y; Yamamoto, K; Osawa, T

1991-02-01

A new type lactose-binding lectin was purified from extracts of Ulex europaeus seeds by affinity chromatography on a column of galactose-Sepharose 4B, followed by gel filtration on Sephacryl S-300. This lectin, designated as Ulex europaeus lectin III (UEA-III), was found to be inhibited by lactose. The dimeric lectin is a glycoprotein with a molecular mass of 70,000 Da; it consists of two apparently identical subunits of a molecular mass of 34,000 Da. Compositional analysis showed that this lectin contains 30% carbohydrate and a large amount of aspartic acid, serine and valine, but no sulfur-containing amino acids. The N-terminal amino-acid sequences of L-fucose-binding Ulex europaeus lectin I (UEA-I) and di-N-acetylchitobiose-binding Ulex europaeus lectin II (UEA-II), both of which we have already purified and characterized, and that of UEA-III were determined and compared.

The endolysins of bacteriophages CMP1 and CN77 are specific for the lysis of Clavibacter michiganensis strains.

PubMed

Wittmann, Johannes; Eichenlaub, Rudolf; Dreiseikelmann, Brigitte

2010-08-01

Putative endolysin genes of bacteriophages CMP1 and CN77, which infect Clavibacter michiganensis subsp. michiganensis and C. michiganensis subsp. nebraskensis, respectively, were cloned and expressed in Escherichia coli. The His-tagged endolysin of CMP1 consists of 306 amino acids and has a calculated molecular mass of 34.8 kDa, while the His-tagged endolysin of CN77 has 290 amino acids with a molecular mass of 31.9 kDa. The proteins were purified and their bacteriolytic activity was demonstrated. The bacteriolytic activity of both enzymes showed a host range which was limited to the respective C. michiganensis subspecies and did not affect other bacteria, even those closely related to Clavibacter. Due to the high specificity of the CMP1 and CN77 endolysins they may be useful tools for biocontrol of plant-pathogenic C. michiganensis without affecting other bacteria in the soil.

Giardia muris trophozoite antigenic targets for mouse intestinal IgA antibody.

PubMed

Heyworth, M F; Vergara, J A

1994-02-01

The aim of this work was to characterize Giardia muris trophozoite proteins that are targets for intestinal anti-trophozoite IgA in G. muris-infected mice. Intestinal secretions were obtained from immunocompetent BALB/c mice that had been infected with G. muris cysts 4-5 weeks previously and from control uninfected BALB/c mice. Flow cytometry of G. muris trophozoites that had been incubated with intestinal secretions and with fluorescein isothiocyanate-conjugated anti-mouse IgA showed that anti-trophozoite IgA was present in intestinal secretions obtained from infected BALB/c mice. By immunoblotting on G. muris trophozoite proteins separated by one-dimensional gel electrophoresis, this IgA recognized at least one trophozoite protein of molecular mass of approximately 80 kDa. The 80-kDa G. muris protein(s) has a molecular mass similar to that described for cysteine-rich surface proteins of the human parasite Giardia lamblia.

Quantitative analysis of modified proteins and their positional isomers by tandem mass spectrometry: human histone H4.

PubMed

Pesavento, James J; Mizzen, Craig A; Kelleher, Neil L

2006-07-01

Here we show that fragment ion abundances from dissociation of ions created from mixtures of multiply modified histone H4 (11 kDa) or of N-terminal synthetic peptides (2 kDa) correspond to their respective intact ion abundances measured by Fourier transform mass spectrometry. Isomeric mixtures of modified forms of the same protein are resolved and quantitated with a precision of

Role of afferent input and mechanical load for size regulation of rat soleus muscle

NASA Astrophysics Data System (ADS)

Kawano, Fuminori; Matsuka, Yoshikazu; Oke, Yoshihiko; Higo, Yoko; Terada, Masahiro; Umemoto, Shiori; Kawabe, Naoko; Wang, Xiao Dong; Shinoda, Yo; Lan, Yong Bo; Fukuda, Hiroyuki; Ohmi, Shinobu; Ohira, Yoshinobu

2005-08-01

Effects of deafferentation on the phosphorylation of ribosomal protein S6 (S6), 27 kDa heat shock protein (HSP27) and extracellular signal-regulated kinase (ERK) 1/2 were studied in rat soleus muscle. Adult male Wistar rats were randomly separated into the pre- and post- experimental control, functionally overloaded (FO), sham-operated, deafferentated (DA), FO+DA, and hindlimb-unloaded (U) groups. The distal tendons of left plantaris and gastrocnemius muscles were transected in the FO rats. The left dorsal roots of the spinal cord at the L4-5 segmental levels were transected in the DA rats. The rats in U were tail-suspended. The sampling of the soleus muscle was performed 2 weeks after the treatments shown above. The cytoplasmic fraction of the soleus muscle homogenate was used for the quantitative analyses of the phosphorylation levels of S6, HSP27, and ERK 1/2. The phosphorylation levels of these proteins were up-regulated by FO. On the contrary, the phosphorylation of all of these proteins was down-regulated by U and DA. Further, the FO-related increase of the protein phosphorylation was inhibited by additional treatment with DA. These results indicated that the afferent feedback plays crucial roles in the intramuscular regulation of the soleus muscle mass.

Efficacy of paracetamol on patent ductus arteriosus closure may be dose dependent: evidence from human and murine studies.

PubMed

El-Khuffash, Afif; Jain, Amish; Corcoran, David; Shah, Prakesh S; Hooper, Christopher W; Brown, Naoko; Poole, Stanley D; Shelton, Elaine L; Milne, Ginger L; Reese, Jeff; McNamara, Patrick J

2014-09-01

We evaluated the clinical effectiveness of variable courses of paracetamol on patent ductus arteriosus (PDA) closure and examined its effect on the in vitro term and preterm murine ductus arteriosus (DA). Neonates received one of the following three paracetamol regimens: short course of oral paracetamol (SCOP), long course of oral paracetamol (LCOP), and intravenous paracetamol (IVP) for 2-6 d. Pressure myography was used to examine changes in vasomotor tone of the preterm and term mouse DA in response to paracetamol or indomethacin. Their effect on prostaglandin synthesis by DA explants was measured by mass spectroscopy. Twenty-one preterm infants were included. No changes in PDA hemodynamics were seen in SCOP infants (n = 5). The PDA became less significant and eventually closed in six LCOP infants (n = 7). PDA closure was achieved in eight IVP infants (n = 9). On pressure myograph, paracetamol induced a concentration-dependent constriction of the term mouse DA, up to 30% of baseline (P < 0.01), but required >1 µmol/l. Indomethacin induced greater DA constriction and suppression of prostaglandin synthesis (P < 0.05). The clinical efficacy of paracetamol on PDA closure may depend on the duration of treatment and the mode of administration. Paracetamol is less potent than indomethacin for constriction of the mouse DA in vitro.

Thermal effects on nonlinear vibration of a carbon nanotube-based mass sensor using finite element analysis

NASA Astrophysics Data System (ADS)

Kang, Dong-Keun; Kim, Chang-Wan; Yang, Hyun-Ik

2017-01-01

In the present study we carried out a dynamic analysis of a CNT-based mass sensor by using a finite element method (FEM)-based nonlinear analysis model of the CNT resonator to elucidate the combined effects of thermal effects and nonlinear oscillation behavior upon the overall mass detection sensitivity. Mass sensors using carbon nanotube (CNT) resonators provide very high sensing performance. Because CNT-based resonators can have high aspect ratios, they can easily exhibit nonlinear oscillation behavior due to large displacements. Also, CNT-based devices may experience high temperatures during their manufacture and operation. These geometrical nonlinearities and temperature changes affect the sensing performance of CNT-based mass sensors. However, it is very hard to find previous literature addressing the detection sensitivity of CNT-based mass sensors including considerations of both these nonlinear behaviors and thermal effects. We modeled the nonlinear equation of motion by using the von Karman nonlinear strain-displacement relation, taking into account the additional axial force associated with the thermal effect. The FEM was employed to solve the nonlinear equation of motion because it can effortlessly handle the more complex geometries and boundary conditions. A doubly clamped CNT resonator actuated by distributed electrostatic force was the configuration subjected to the numerical experiments. Thermal effects upon the fundamental resonance behavior and the shift of resonance frequency due to attached mass, i.e., the mass detection sensitivity, were examined in environments of both high and low (or room) temperature. The fundamental resonance frequency increased with decreasing temperature in the high temperature environment, and increased with increasing temperature in the low temperature environment. The magnitude of the shift in resonance frequency caused by an attached mass represents the sensing performance of a mass sensor, i.e., its mass detection sensitivity, and it can be seen that this shift is affected by the temperature change and the amount of electrostatic force. The thermal effects on the mass detection sensitivity are intensified in the linear oscillation regime and increase with increasing CNT length; this intensification can either improve or worsen the detection sensitivity.

Vector and scalar charmonium resonances with lattice QCD

DOE PAGES

Lang, C. B.; Leskovec, Luka; Mohler, Daniel; ...

2015-09-15

We perform an exploratory lattice QCD simulation of DD¯ scattering, aimed at determining the masses as well as the decay widths of charmonium resonances above open charm threshold. Neglecting coupling to other channels, the resulting phase shift for DD¯ scattering in p-wave yields the well-known vector resonance ψ(3770). For m π = 156 MeV, the extracted resonance mass and the decay width agree with experiment within large statistical uncertainty. The scalar charmonium resonances present a puzzle, since only the ground state Χc0(1P) is well understood, while there is no commonly accepted candidate for its first excitation. We simulate DD¯ scatteringmore » in s-wave in order to shed light on this puzzle. The resulting phase shift supports the existence of a yet-unobserved narrow resonance with a mass slightly below 4 GeV. A scenario with this narrow resonance and a pole at Χc0(1P) agrees with the energy-dependence of our phase shift. In addition, further lattice QCD simulations and experimental efforts are needed to resolve the puzzle of the excited scalar charmonia.« less

The respiratory arsenate reductase from Bacillus selenitireducens strain MLS10

USGS Publications Warehouse

Afkar, E.; Lisak, J.; Saltikov, C.; Basu, P.; Oremland, R.S.; Stolz, J.F.

2003-01-01

The respiratory arsenate reductase from the Gram-positive, haloalkaliphile, Bacillus selenitireducens strain MLS10 was purified and characterized. It is a membrane bound heterodimer (150 kDa) composed of two subunits ArrA (110 kDa) and ArrB (34 kDa), with an apparent Km for arsenate of 34 ??M and Vmax of 2.5 ??mol min-1 mg-1. Optimal activity occurred at pH 9.5 and 150 g l-1 of NaCl. Metal analysis (inductively coupled plasma mass spectrometry) of the holoenzyme and sequence analysis of the catalytic subunit (ArrA; the gene for which was cloned and sequenced) indicate it is a member of the DMSO reductase family of molybdoproteins. ?? 2003 Federation of European Microbiological Societies. Published by Elsevier B.V. All rights reserved.

Theories of Variable Mass Particles and Low Energy Nuclear Phenomena

NASA Astrophysics Data System (ADS)

Davidson, Mark

2014-02-01

Variable particle masses have sometimes been invoked to explain observed anomalies in low energy nuclear reactions (LENR). Such behavior has never been observed directly, and is not considered possible in theoretical nuclear physics. Nevertheless, there are covariant off-mass-shell theories of relativistic particle dynamics, based on works by Fock, Stueckelberg, Feynman, Greenberger, Horwitz, and others. We review some of these and we also consider virtual particles that arise in conventional Feynman diagrams in relativistic field theories. Effective Lagrangian models incorporating variable mass particle theories might be useful in describing anomalous nuclear reactions by combining mass shifts together with resonant tunneling and other effects. A detailed model for resonant fusion in a deuterium molecule with off-shell deuterons and electrons is presented as an example. Experimental means of observing such off-shell behavior directly, if it exists, is proposed and described. Brief explanations for elemental transmutation and formation of micro-craters are also given, and an alternative mechanism for the mass shift in the Widom-Larsen theory is presented. If variable mass theories were to find experimental support from LENR, then they would undoubtedly have important implications for the foundations of quantum mechanics, and practical applications may arise.

Identification of a Hemagglutinin from Gallibacterium anatis.

PubMed

Montes-García, J F; Vaca, S; Vazquez-Cruz, C; Soriano-Vargas, E; Aguilar-Romero, F; Blackall, P J; Negrete-Abascal, E

2016-04-01

Gallibacterium anatis has the ability to hemagglutinate rabbit erythrocytes; however, no bacterial component has yet been associated with this function. In the present work, a protein of approximately 65 kDa with hemagglutinating activity for glutaraldehyde-fixed chicken erythrocytes was purified by ion interchange chromatography from G. anatis F149(T) secreted proteins. The protein was recognized by a rabbit polyclonal serum against a hemagglutinin from Avibacterium paragallinarum. The 65 kDa purified protein presented identity with a G. anatis filamentous hemagglutinin by mass spectrometric analysis. As well, the bacterial surface of G. anatis was labeled by immune gold assays using a polyclonal serum against the 65-kDa protein. A similar protein was recognized in four other G. anatis strains by immunoblots using the same antiserum. The protein binds sheep or pig biotinylated fibrinogen, suggesting an interaction with basement membrane eukaryotic cells components, and the protein is present in G. anatis biofilms. Overall, the results suggest that the 65 kDa hemagglutinin is a common antigen and a potential virulence factor in G. anatis.

Isolation and purification of two bacteriocins 3D produced by Enterococcus faecium with inhibitory activity against Listeria monocytogenes.

PubMed

Bayoub, Kaoutar; Mardad, Ilham; Ammar, Emna; Serrano, Aurelio; Soukri, Abdelaziz

2011-02-01

Strain 3D, isolated from fermented traditional Moroccan dairy product, and identified as Enterococcus faecium, was studied for its capability to produce two bacteriocins acting against Listeria monocytogenes. Bacteriocins 3 Da and 3Db were heat stable inactivated by proteinase K, pepsin, and trypsin but not when treated with catalase. The evidenced bacteriocins were stable in a wide pH range from 2 to 11 and bactericidal activity was kept during storage at 4°C. However, the combination of temperature and pH exhibited a stability of the bacteriocins. RP-HPLC purification of the anti-microbial compounds shows two active fractions eluted at 16 and 30.5 min, respectively. Mass spectrometry analysis showed that E. faecium 3D produce two bacteriocins Enterocin 3 Da (3893.080 Da) and Enterocin 3Db (4203.350 Da). This strain is food-grade organism and its bacteriocins were heat-stable peptides at basic, neutral, and acid pH: such bacteriocins may be of interest as food preservatives.

Purification of nitrate reductase from Nicotiana plumbaginifolia by affinity chromatography using 5'AMP-sepharose and monoclonal antibodies.

PubMed

Moureaux, T; Leydecker, M T; Meyer, C

1989-02-15

Nitrate reductase was purified from leaves of Nicotiana plumbaginifolia using either 5'AMP-Sepharose chromatography or two steps of immunoaffinity chromatography involving monoclonal antibodies directed against nitrate reductase from maize and against ribulose-1,5-bisphosphate carboxylase from N. plumbaginifolia. Nitrate reductase obtained by the first method was purified 1000-fold to a specific activity of 9 units/mg protein. The second method produced an homogenous enzyme, purified 21,000-fold to a specific activity of 80 units/mg protein. SDS/PAGE of nitrate reductase always resulted in two bands of 107 and 99.5 kDa. The 107-kDa band was the nitrate reductase subunit of N. plumbaginifolia; the smaller one of 99.5 kDa is thought, as commonly reported, to result from proteolysis of the larger protein. The molecular mass of 107 kDa is close to the values calculated from the coding sequences of the two nitrate reductase genes recently cloned from tobacco (Nicotiana tabacum cv Xanthi).

Analysis of Biological Particles by Mass Spectrometry

DTIC Science & Technology

1984-10-23

approximately 20 nm thick layers of gold /palladium. f. 4( RESULTS Particle beam characterization. The objectives of this part of the study were to determine the...5 and produce a spectral shift in the light aborption chara- cteristics of the dye. Measuring this shift by conventional spectroscopy has 4 been the

Association of Rotating Night Shift Work with BMI and Abdominal Obesity among Nurses and Midwives.

PubMed

Peplonska, Beata; Bukowska, Agnieszka; Sobala, Wojciech

2015-01-01

Mounting epidemiological evidence suggests that night shift work may contribute to the etiology of increased body weight. The present study aimed to examine association between rotating night shift work and body mass index (BMI), and abdominal adiposity respectively among nurses and midwives. A cross-sectional study was conducted among 724 female nurses and midwives, aged 40-60 years (354 rotating night shift and 370 daytime workers) in Łódź, Poland, between 2008 and 2011. Information about occupational history and potential confounders was collected during personal interviews. Anthropometric measurements of body weight, height, waist (WC) and hip (HC) circumference were made, and body mass index (BMI), waist to hip ratio (WHR) and waist to height ratio (WHtR) were calculated. GLM regression models and multinomial logit regression models were fitted to explore the association between night shift work and anthropometric parameters, with adjustment for age, body silhouette at age 20, current smoking status, packyears, marital status, and menopausal hormone therapy use. Cumulative night shift work showed significant associations with BMI, WC, HC and WHtR, with BMI increasing by 0.477 kg/m2 per 1000 night duties and by 0.432 kg/m2 per 10000 night shift hours, WC increasing respectively by 1.089 cm and 0.99 cm, and HC by 0.72 cm and WHtR by 0.007 cm for both metrics. Both current and cumulative night work was associated with obesity (BMI≥30kg/m2), with OR=3.9 (95%CI:1.5-9.9), in women reporting eight or more night shifts per month. The results of the study support the previously reported relations between night shift work and development of obesity.

Association of Rotating Night Shift Work with BMI and Abdominal Obesity among Nurses and Midwives

PubMed Central

Peplonska, Beata; Bukowska, Agnieszka; Sobala, Wojciech

2015-01-01

Background Mounting epidemiological evidence suggests that night shift work may contribute to the etiology of increased body weight. The present study aimed to examine association between rotating night shift work and body mass index (BMI), and abdominal adiposity respectively among nurses and midwives. Methods A cross-sectional study was conducted among 724 female nurses and midwives, aged 40-60 years (354 rotating night shift and 370 daytime workers) in Łódź, Poland, between 2008 and 2011. Information about occupational history and potential confounders was collected during personal interviews. Anthropometric measurements of body weight, height, waist (WC) and hip (HC) circumference were made, and body mass index (BMI), waist to hip ratio (WHR) and waist to height ratio (WHtR) were calculated. GLM regression models and multinomial logit regression models were fitted to explore the association between night shift work and anthropometric parameters, with adjustment for age, body silhouette at age 20, current smoking status, packyears, marital status, and menopausal hormone therapy use. Results Cumulative night shift work showed significant associations with BMI, WC, HC and WHtR, with BMI increasing by 0.477 kg/m2 per 1000 night duties and by 0.432 kg/m2 per 10000 night shift hours, WC increasing respectively by 1.089 cm and 0.99 cm, and HC by 0.72 cm and WHtR by 0.007 cm for both metrics. Both current and cumulative night work was associated with obesity (BMI≥30kg/m2), with OR=3.9 (95%CI:1.5-9.9), in women reporting eight or more night shifts per month. Conclusion The results of the study support the previously reported relations between night shift work and development of obesity. PMID:26196859

A 15-year prospective study of shift work and disability pension

PubMed Central

Tüchsen, F; Christensen, K B; Lund, T; Feveile, H

2008-01-01

Objective: To estimate the hazard ratio for disability pension associated with shift work. Methods: Cohorts of shift and day workers were identified in three waves of the Danish Work Environment Cohort Study and followed up for incidence of disability pension in a national register of social transfer payment. A total of 3980 female and 4025 male employees were included in the cohorts. Information about shift work status, age, smoking habits, body mass index and ergonomic work environment were updated according to responses in subsequent waves of the survey when possible. Respondents reporting shift work were classified as shift workers in the following waves as well. Respondents were followed in the register from the time of first interview and were censored at the time of their 60th birthday, emigration, death or end of follow-up (18 June 2006). The authors used the Cox proportional hazards model to estimate hazard ratios for incidence of disability pension and 95% confidence intervals. Results: The authors observed 253 new disability pensions among women and 173 among men during 56 903 and 57 886 person-years at risk respectively, Among women, shift work predicted disability after adjustment for age, general health and socioeconomic status HR 1.39 (95% CI 1.07 to 1.82). After further adjustment for body mass index, smoking habits, socioeconomic status and ergonomic exposures the association remained statistically significant HR 1.34 (95% CI 1.02 to 1.75). Shift work was not associated with disability among men. Conclusion: Shift work might be moderately associated with disability pension among women; however, more powerful studies are needed to establish the possible association. PMID:18198201

Long work hours and adiposity among police officers in a US northeast city.

PubMed

Gu, Ja K; Charles, Luenda E; Burchfiel, Cecil M; Fekedulegn, Desta; Sarkisian, Khachatur; Andrew, Michael E; Ma, Claudia; Violanti, John M

2012-11-01

To investigate the associations between long work hours and adiposity measures in police officers. Participants included 408 officers from the Buffalo Cardio-Metabolic Occupational Police Stress study who were examined between 2004 and 2009. Total work hours were abstracted from payroll records and questionnaires. Analysis of variance and covariance models were used. Among male officers who worked the midnight shift, mean values of waist circumference and body mass index increased with longer work hours after adjustment for age, physical activity, energy intake, sleep duration, smoking status, police rank, activities after work (eg, child/family care, sports), and household income. Adiposity measures were not associated with work hours among women on any shift. Working longer hours was significantly associated with larger waist circumferences and higher body mass index among male police officers working the midnight shift.

Monoclonal antibody against a serotype antigen of Porphyromonas (Bacteroides) endodontalis and characteristics of the antigen.

PubMed Central

Hanazawa, S; Sagiya, T; Amano, S; Nishikawa, H; Kitano, S

1990-01-01

Recent studies have demonstrated the presence of three serotypes (O1K1, O1K2, and O1K-) of Porphyromonas (Bacteroides) endodontalis. In the present study, a hybridoma cell line producing monoclonal antibody (BEE11) specific for serotype O1K1 of P. endodontalis was established. The specificity of the antibody was evaluated by enzyme-linked immunosorbent assay and immunoslot blot analysis. BEE11 antibody reacted with strains ATCC 35406, HG 400, and HG 421 of the bacterium. However, it did not react with HG 422 or HG 948. Also, the antibody did not react with any of the black-pigmented Bacteroides strains tested. Although the antibody reacted with total cell envelope and capsule materials, it did not do so with lipopolysaccharide. The antibody reacted with antigen material having a molecular mass of 110 kilodaltons (kDa), as judged from fractionation by Superose 12 prep gel chromatography. When the peak fraction from the Superose 12 column was subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blot (immunoblot) analysis, the reactivity was detected as a single band at an apparent molecular mass of about 52 kDa. The antigen material purified partially by high-performance liquid chromatography was sensitive to trypsin, V8 protease, and heating to 80 degrees C but not to neuraminidase. Therefore, the present study shows that BEE11 antibody recognizes a serotype antigen of P. endodontalis which may be a dimer consisting of monomers having molecular masses of approximately 52 kDa and sensitivity to proteases and heat. Images PMID:2370106

The core of tau-paired helical filaments studied by scanning transmission electron microscopy and limited proteolysis.

PubMed

von Bergen, Martin; Barghorn, Stefan; Müller, Shirley A; Pickhardt, Marcus; Biernat, Jacek; Mandelkow, Eva-Maria; Davies, Peter; Aebi, Ueli; Mandelkow, Eckhard

2006-05-23

In Alzheimer's disease and frontotemporal dementias the microtubule-associated protein tau forms intracellular paired helical filaments (PHFs). The filaments formed in vivo consist mainly of full-length molecules of the six different isoforms present in adult brain. The substructure of the PHF core is still elusive. Here we applied scanning transmission electron microscopy (STEM) and limited proteolysis to probe the mass distribution of PHFs and their surface exposure. Tau filaments assembled from the three repeat domain have a mass per length (MPL) of approximately 60 kDa/nm and filaments from full-length tau (htau40DeltaK280 mutant) have approximately 160 kDa/nm, compared with approximately 130 kDa/nm for PHFs from Alzheimer's brain. Polyanionic cofactors such as heparin accelerate assembly but are not incorporated into PHFs. Limited proteolysis combined with N-terminal sequencing and mass spectrometry of fragments reveals a protease-sensitive N-terminal half and semiresistant PHF core starting in the first repeat and reaching to the C-terminus of tau. Continued proteolysis leads to a fragment starting at the end of the first repeat and ending in the fourth repeat. PHFs from tau isoforms with four repeats revealed an additional cleavage site within the middle of the second repeat. Probing the PHFs with antibodies detecting epitopes either over longer stretches in the C-terminal half of tau or in the fourth repeat revealed that they grow in a polar manner. These data describe the physical parameters of the PHFs and enabled us to build a model of the molecular arrangement within the filamentous structures.

Purification and characterization of rat liver minoxidil sulphotransferase.

PubMed Central

Hirshey, S J; Falany, C N

1990-01-01

Minoxidil (Mx), a pyrimidine N-oxide, is used therapeutically as an antihypertensive agent and to induce hair growth in patients with male pattern baldness. Mx NO-sulphate has been implicated as the agent active in producing these effects. This paper describes the purification of a unique sulphotransferase (ST) from rat liver cytosol that is capable of catalysing the sulphation of Mx. By using DEAE-Sepharose CL-6B chromatography, hydroxyapatite chromatography and ATP-agarose affinity chromatography, Mx-ST activity was purified 240-fold compared with the activity in cytosol. The purified enzyme was also capable of sulphating p-nitrophenol (PNP) at low concentrations (less than 10 microM). Mx-ST was purified to homogeneity, as evaluated by SDS/PAGE and reverse-phase h.p.l.c. The active form of the enzyme had a molecular mass of 66,000-68,000 Da as estimated by gel exclusion chromatography and a subunit molecular mass of 35,000 Da. The apparent Km values for Mx, 3'-phosphoadenosine 5'-phosphosulphate and PNP were 625 microM, 5.0 microM and 0.5 microM respectively. However, PNP displayed potent substrate inhibition at concentrations above 1.2 microM. Antibodies raised in rabbits to the pure enzyme detected a single band in rat liver cytosol with a subunit molecular mass of 35,000 Da, as determined by immunoblotting. The anti-(rat Mx-ST) antibodies also reacted with the phenol-sulphating form of human liver phenol sulphotransferase, suggesting some structural similarity between these proteins. Images Fig. 5. Fig. 6. Fig. 7. PMID:2241904

Extremely stable soluble high molecular mass multi-protein complex with DNase activity in human placental tissue.

PubMed

Burkova, Evgeniya E; Dmitrenok, Pavel S; Sedykh, Sergey E; Buneva, Valentina N; Soboleva, Svetlana E; Nevinsky, Georgy A

2014-01-01

Human placenta is an organ which protects, feeds, and regulates the grooving of the embryo. Therefore, identification and characterization of placental components including proteins and their multi-protein complexes is an important step to understanding the placenta function. We have obtained and analyzed for the first time an extremely stable multi-protein complex (SPC, ∼ 1000 kDa) from the soluble fraction of three human placentas. By gel filtration on Sepharose-4B, the SPC was well separated from other proteins of the placenta extract. Light scattering measurements and gel filtration showed that the SPC is stable in the presence of NaCl, MgCl2, acetonitrile, guanidinium chloride, and Triton in high concentrations, but dissociates efficiently in the presence of 8 M urea, 50 mM EDTA, and 0.5 M NaCl. Such a stable complex is unlikely to be a casual associate of different proteins. According to SDS-PAGE and MALDI mass spectrometry data, this complex contains many major glycosylated proteins with low and moderate molecular masses (MMs) 4-14 kDa and several moderately abundant (79.3, 68.5, 52.8, and 27.2 kDa) as well as minor proteins with higher MMs. The SPC treatment with dithiothreitol led to a disappearance of some protein bands and revealed proteins with lower MMs. The SPCs from three placentas efficiently hydrolyzed plasmid supercoiled DNA with comparable rates and possess at least two DNA-binding sites with different affinities for a 12-mer oligonucleotide. Progress in study of placental protein complexes can promote understanding of their biological functions.

Shift work-related problems in 16-h night shift nurses (1): Development of an automated data processing system for questionnaires, heart rate, physical activity and posture.

PubMed

Fukuda, H; Takahashi, M; Miki, K; Haratani, T; Kurabayashi, L; Hisanaga, N; Arito, H; Takahashi, H; Egoshi, M; Sakurai, M

1999-04-01

To assess the shift work-related problems associated with a 16-h night shift in a two-shift system, we took the following important factors into consideration; the interaction between circadian rhythms and the longer night shift, the type of morningness and eveningness experienced, the subjective sleep feeling, the subjects' daily behavior, the effectiveness of taking a nap during the long night shift, and finally the effectiveness of using several different kinds of measuring devices. Included among the measuring devices used were a standard questionnaire, repetitive self-assessment of subjective symptoms and daily behavior at short intervals, and a continuous recording of such objective indices as physical activity and heart rate. A potential problem lies in the fact that field studies that use such measures tend to produce a mass of data, and are thus faced with the accompanying technical problem of analyzing such a large amount of data (time, effort and cost). To solve the data analysis problem, we developed an automated data processing system. Through the use of an image scanner with a paper feeder, standard paper, an optical character recognition function and common application software, we were able to analyze a mass of data continuously and automatically within a short time. Our system should prove useful for field studies that produce a large amount of data collected with several different kinds of measuring devices.

Algorithms for calculating mass-velocity and Darwin relativistic corrections with n-electron explicitly correlated Gaussians with shifted centers

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

Stanke, Monika, E-mail: monika@fizyka.umk.pl; Palikot, Ewa, E-mail: epalikot@doktorant.umk.pl; Adamowicz, Ludwik, E-mail: ludwik@email.arizona.edu

2016-05-07

Algorithms for calculating the leading mass-velocity (MV) and Darwin (D) relativistic corrections are derived for electronic wave functions expanded in terms of n-electron explicitly correlated Gaussian functions with shifted centers and without pre-exponential angular factors. The algorithms are implemented and tested in calculations of MV and D corrections for several points on the ground-state potential energy curves of the H{sub 2} and LiH molecules. The algorithms are general and can be applied in calculations of systems with an arbitrary number of electrons.

Plasma volume methodology: Evans blue, hemoglobin-hematocrit, and mass density transformations

NASA Technical Reports Server (NTRS)

Greenleaf, J. E.; Hinghofer-Szalkay, H.

1985-01-01

Methods for measuring absolute levels and changes in plasma volume are presented along with derivations of pertinent equations. Reduction in variability of the Evans blue dye dilution technique using chromatographic column purification suggests that the day-to-day variability in the plasma volume in humans is less than + or - 20 m1. Mass density determination using the mechanical-oscillator technique provides a method for measuring vascular fluid shifts continuously for assessing the density of the filtrate, and for quantifying movements of protein across microvascular walls. Equations for the calculation of volume and density of shifted fluid are presented.

Isoform composition and stoichiometry of the approx. 90-kDa heat shock protein associated with glucocorticoid receptors

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

Mendel, D.B.; Orti, E.

1988-05-15

The authors observed that the approx. 90-kDa non-steroid-binding component of nonactivated glucocorticoid receptors purified from WEHI-7 mouse thymoma cells (which has been identified as the approx. 90-kDa heat shock protein) consistently migrates as a doublet during polyacrylamide gel electrophoresis under denaturing and reducing conditions. It has recently been reported that murine Meth A cells contain a tumor-specific transplantation antigen (TSTA) which is related or identical to the approx. 90-kDa heat shock protein. The observation that TSTA and the approx. 90-kDa heat shock protein isolated from these cells exists as two isoforms of similar molecular mass and charge has suggested thatmore » the doublet observed is also due to the existence of two isoforms. They have therefore conducted this study to determine whether TSTA and the approx. 90-kDa component of glucocorticoid receptors are indeed related, to establish whether the receptor preferentially binds one isoform of the approx. 90-kDa heat shock protein, and to investigate the stoichiometry of the nonactivated receptor complex. They used the BuGr1 and AC88 monoclonal antibodies to purify, respectively, receptor-associated and free approx. 90-kDa heat shock protein from WEHI-7 cells grown for 48 h with (/sup 35/S)methionine to metabolically label proteins to steady state. The long-term metabolic labeling approach has also enabled them to directly determine that the purified non-activated glucocorticoid receptor contains a single steroid-binding protein and two approx. 90-kDa non-steroid-binding subunits. The consistency with which a approx. 1:2 stoichiometric ratio of steroid binding to approx. 90-kDa protein is observed supports the view that the approx. 90-kDa heat shock protein is a true component of nonactivated glucocorticoid-receptor complexes.« less

The notions of mass in gravitational and particle physics

NASA Astrophysics Data System (ADS)

Castellani, Gianluca

It is presently thought that the mass of all of the elementary particles is determined by the Higgs field. This scalar field couples directly into the trace of the energy momentum tensor of the elementary particles. The attraction between two or more masses arises from the exchange of gravitational quantum particles of spin 2, called gravitons. The gravitational field couples directly into the energy momentum tensor. Then there is a close connection between the Higgs field, that originates the mass, and the gravitational field that dictates how the masses interact. Our purpose in this thesis is to discuss this close connection in terms of fundamental definitions of inertial and gravitational masses. On a practical level we explore two properties of mass from the viewpoint of coupling into the Higgs field: (i) The coupling of the both the Higgs and gravity to the energy-pressure tensor allows for the decay of the Higgs particle into two gravitons. We use the self energy part of the Higgs propagator to calculate the electromagnetic, weak, fermionic and gravitational decay rate of the Higgs particle. We show that the former process appears to dominate the other decay modes. Since the gravitons are detectable with virtually zero probability, the number of Higgs particles with observable decay products will be much less than previously expected. (ii) Some new experimental results seem to indicate that the mass of the heavy elementary particles like the Z,W+,W- and especially the top quark, depends on the particle environment in which these particles are produced. The presence of a Higgs field due to neighboring particles could be responsible for induced mass shifts. Further measurements of mass shift effects might give an indirect proof of the Higgs particle. Such can be in principle done by re-analyzing some of the production data e +e- → ZZ (or W+W-) already collected at the LEP experiment. About the physical property of the top quark, it is too early to arrive at any conclusion. In the foreseeable future, there will be more extended top quark production statistics from the Tevatron accelerator so that the mass shift hypothesis can be experimentally probed.

Backbone and stereospecific (13)C methyl Ile (δ1), Leu and Val side-chain chemical shift assignments of Crc.

PubMed

Sharma, Rakhi; Sahu, Bhubanananda; Ray, Malay K; Deshmukh, Mandar V

2015-04-01

Carbon catabolite repression (CCR) allows bacteria to selectively assimilate a preferred compound among a mixture of several potential carbon sources, thus boosting growth and economizing the cost of adaptability to variable nutrients in the environment. The RNA-binding catabolite repression control (Crc) protein acts as a global post-transcriptional regulator of CCR in Pseudomonas species. Crc triggers repression by inhibiting the expression of genes involved in transport and catabolism of non-preferred substrates, thus indirectly favoring assimilation of preferred one. We report here a nearly complete backbone and stereospecific (13)C methyl side-chain chemical shift assignments of Ile (δ1), Leu and Val of Crc (~ 31 kDa) from Pseudomonas syringae Lz4W.

Normal-phase liquid chromatography retention behavior of polycyclic aromatic hydrocarbon and their methyl-substituted derivatives on an aminopropyl stationary phase.

PubMed

Wilson, Walter B; Hayes, Hugh V; Sander, Lane C; Campiglia, Andres D; Wise, Stephen A

2017-09-01

Retention indices for 124 polycyclic aromatic hydrocarbons (PAHs) and 62 methyl-substituted (Me-) PAHs were determined using normal-phase liquid chromatography (NPLC) on a aminopropyl (NH 2 ) stationary phase. PAH retention behavior on the NH 2 phase is correlated to the total number of aromatic carbons in the PAH structure. Within an isomer group, non-planar isomers generally elute earlier than planar isomers. MePAHs generally elute slightly later but in the same region as the parent PAHs. Correlations between PAH retention behavior on the NH 2 phase and PAH thickness (T) values were investigated to determine the influence of non-planarity for isomeric PAHs with four to seven aromatic rings. Correlation coefficients ranged from r = 0.19 (five-ring peri-condensed molecular mass (MM) 252 Da) to r = -0.99 (five-ring cata-condensed MM 278 Da). In the case of the smaller PAHs (MM ≤ 252 Da), most of the PAHs had a planar structure and provided a low correlation. In the case of larger PAHs (MM ≥ 278 Da), nonplanarity had a significant influence on the retention behavior and good correlation between retention and T was obtained for the MM 278 Da, MM 302 Da, MM 328 Da, and MM 378 Da isomer sets. Graphical abstract NPLC separation of the three-, four-, five-, and six-ring PAH isomers with different number of aromatic carbon atoms and degrees of non-planarity (Thickness, T). The inserted figure plots the number of aromatic carbon atoms vs. the log I value for the 124 parent PAHs.

Maize 27 kDa gamma-zein is a potential allergen for early weaned pigs.

PubMed

Krishnan, Hari B; Kerley, Monty S; Allee, Gary L; Jang, Sungchan; Kim, Won-Seok; Fu, Chunjiang J

2010-06-23

Soybean and maize are extensively used in animal feed, primarily in poultry, swine, and cattle diets. Soybean meal can affect pig performance in the first few weeks following weaning and elicit specific antibodies in weaned piglets. Though maize is a major component of pig feed, it is not known if any of the maize proteins can elicit immunological response in young pigs. In this study, we have identified a prominent 27 kDa protein from maize as an immunodominant protein in young pigs. This protein, like some known allergens, exhibited resistance to pepsin digestion in vitro. Several lines of evidence identify the immunodominant 27 kDa protein as a gamma-zein, a maize seed storage protein. First, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis of different solubility classes of maize seed proteins revealed the presence of an abundant 27 kDa protein in the prolamin (zein) fraction. Antibodies raised against the purified maize 27 kDa gamma-zein also reacted against the same protein recognized by the young pig serum. Additionally, matrix-assisted laser desorption ionization time-of-flight mass spectrometry analysis of the peptides generated by trypsin digestion of the immunodominant 27 kDa protein showed significant homology to the maize 27 kDa gamma-zein. Since eliminating the allergenic protein will have a great impact on the nutritive value of the maize meal and expand its use in the livestock industry, it will be highly desirable to develop maize cultivars completely lacking the 27 kDa allergenic protein.

Proteolytic processing of connective tissue growth factor in normal ocular tissues and during corneal wound healing.

PubMed

Robinson, Paulette M; Smith, Tyler S; Patel, Dilan; Dave, Meera; Lewin, Alfred S; Pi, Liya; Scott, Edward W; Tuli, Sonal S; Schultz, Gregory S

2012-12-13

Connective tissue growth factor (CTGF) is a fibrogenic cytokine that is up-regulated by TGF-β and mediates most key fibrotic actions of TGF-β, including stimulation of synthesis of extracellular matrix and differentiation of fibroblasts into myofibroblasts. This study addresses the role of proteolytic processing of CTGF in human corneal fibroblasts (HCF) stimulated with TGF-β, normal ocular tissues and wounded corneas. Proteolytic processing of CTGF in HCF cultures, normal animal eyes, and excimer laser wounded rat corneas were examined by Western blot. The identity of a 21-kDa band was determined by tandem mass spectrometry, and possible alternative splice variants of CTGF were assessed by 5' Rapid Amplification of cDNA Ends (RACE). HCF stimulated by TGF-β contained full length 38-kDa CTGF and fragments of 25, 21, 18, and 13 kDa, while conditioned medium contained full length 38- and a 21-kDa fragment of CTGF that contained the middle "hinge" region of CTGF. Fragmentation of recombinant CTGF incubated in HCF extracts was blocked by the aspartate protease inhibitor, pepstatin. Normal mouse, rat, and rabbit whole eyes and rabbit ocular tissues contained abundant amounts of C-terminal 25- and 21-kDa fragments and trace amounts of 38-kDa CTGF, although no alternative transcripts were detected. All forms of CTGF (38, 25, and 21 kDa) were detected during healing of excimer ablated rat corneas, peaking on day 11. Proteolytic processing of 38-kDa CTGF occurs during corneal wound healing, which may have important implications in regulation of corneal scar formation.

Analysis of intact glucuronides and sulfates of serotonin, dopamine, and their phase I metabolites in rat brain microdialysates by liquid chromatography-tandem mass spectrometry.

PubMed

Uutela, Päivi; Reinilä, Ruut; Harju, Kirsi; Piepponen, Petteri; Ketola, Raimo A; Kostiainen, Risto

2009-10-15

A method for the analysis of intact glucuronides and sulfates of common neurotransmitters serotonin (5-HT) and dopamine (DA) as well as of 5-hydroxy-3-indoleacetic acid (5-HIAA), 3,4-dihydroxyphenylacetic acid (DOPAC), and homovanillic acid (HVA) in rat brain microdialysates by liquid chromatography-tandem mass spectrometry (LC-MS/MS) was developed. Enzyme-assisted synthesis using rat liver microsomes as a biocatalyst was employed for the production of 5-HT-, 5-HIAA-, DOPAC-, and HVA-glucuronides for reference compounds. The sulfate conjugates were synthesized either chemically or enzymatically using a rat liver S9 fraction. The LC-MS/MS method was validated by determining the limits of detection and quantitation, linearity, and repeatability for the quantitative analysis of 5-HT and DA and their glucuronides, as well as of 5-HIAA, DOPAC, and HVA and their sulfate-conjugates. In this study, 5-HT-glucuronide was for the first time detected in rat brain. The concentration of 5-HT-glucuronide (1.0-1.7 nM) was up to 2.5 times higher than that of free 5-HT (0.4-2.1 nM) in rat brain microdialysates, whereas the concentration of DA-glucuronide (1.0-1.4 nM) was at the same level or lower than the free DA (1.2-2.4 nM). The acidic metabolites of neurotransmitters, 5-HIAA, HVA, and DOPAC, were found in free and sulfated form, whereas their glucuronidation was not observed.

Comparative effect of lurasidone and blonanserin on cortical glutamate, dopamine, and acetylcholine efflux: role of relative serotonin (5-HT)2A and DA D2 antagonism and 5-HT1A partial agonism.

PubMed

Huang, Mei; Panos, John J; Kwon, Sunoh; Oyamada, Yoshihiro; Rajagopal, Lakshmi; Meltzer, Herbert Y

2014-03-01

Atypical antipsychotic drugs (AAPDs) have been suggested to be more effective in improving cognitive impairment in schizophrenia than typical APDs, a conclusion supported by differences in receptor affinities and neurotransmitter efflux in the cortex and the hippocampus. More potent serotonin (5-HT)2A than dopamine (DA) D2 receptors antagonism, and direct or indirect 5-HT1A agonism, characterize almost all AAPDs. Blonanserin, an AAPD, has slightly greater affinity for D2 than 5-HT2A receptors. Using microdialysis and ultra performance liquid chromatography-mass spectrometry/mass spectrometry, we compared the abilities of the typical APD, haloperidol, three AAPDs, blonanserin, lurasidone, and olanzapine, and a selective 5-HT1A partial agonist, tandospirone, and all, except haloperidol, were found to ameliorate the cognitive deficits produced by the N-methyl-d-aspartate antagonist, phencyclidine, altering the efflux of neurotransmitters and metabolites in the rat cortex and nucleus accumbens. Blonanserin, lurasidone, olanzapine, and tandospirone, but not haloperidol, increased the efflux of cortical DA and its metabolites, homovanillic acid and 3,4-dihydroxyphenylacetic acid. Olanzapine and lurasidone increased the efflux of acetylcholine; lurasidone increased glutamate as well. None of the compounds significantly altered the efflux of 5-HT or its metabolite, 5-hydroxyindole acetic acid, or GABA, serine, and glycine. The ability to increase cortical DA efflux was the only shared effect of the compounds which ameliorates the deficit in cognition in rodents following phencyclidine. © 2013 International Society for Neurochemistry.

Isolation and characterization of flagellar filament from zoospores of Dermatophilus congolensis.

PubMed

Hiraizumi, Mieko; Tagawa, Yuichi

2014-09-17

Highly motile zoospores from Dermatophilus congolensis bovine isolates from clinical dermatophilosis in Japan were obtained by culturing at 27°C in an ambient atmosphere on heart infusion agar supplemented with 5% defibrinated sheep blood for 72h or in heart infusion broth for 48h with gentle shaking. After vigorous mechanical agitation of the zoospore suspension, the flagellar filaments detached from motile zoospores and were isolated in the clear gelatinous part of the final pellet by differential centrifugation. Typical morphology of a flagellar filament, with a width of approximately 15nm, was observed in the isolated flagellar filament by electron microscopy. A single major protein (flagellin) band with an apparent molecular mass of 35kDa was detected in the flagellar filament of D. congolensis strain AM-1 and that of 33kDa was detected in strain IT-2 by SDS-PAGE. In immunoblot analysis of whole-cell proteins from seven isolates of D. congolensis, antiserum to strain AM-1 zoospores reacted with the 35-kDa antigen band of strain AM-1, but not with any antigen band of other strains in a similar molecular mass range. In contrast, antiserum to strain IT-2 zoospores reacted with antigen bands at 33kDa from six strains, except strain AM-1. Similar strain-specific reactions of these anti-zoospore sera with isolated flagellar filaments from strains AM-1 and IT-2 were confirmed by immunoblot, indicating the presence of antigenic variations of flagellins of D. congolensis zoospores. Copyright © 2014 Elsevier B.V. All rights reserved.

Molecular cloning and heterologous expression of the isopullulanase gene from Aspergillus niger A.T.C.C. 9642.

PubMed Central

Aoki, H; Yopi; Sakano, Y

1997-01-01

Isopullulanase (IPU) from Aspergillus niger A.T.C.C. (American Type Culture Collection) 9642 hydrolyses pullulan to isopanose. IPU is important for the production of isopanose and is used in the structural analysis of oligosaccharides with alpha-1,4 and alpha-1,6 glucosidic linkages. We have isolated the ipuA gene encoding IPU from the filamentous fungi A. niger A.T.C.C. 9642. The ipuA gene encodes an open reading frame of 1695 bp (564 amino acids). IPU contained a signal sequence of 19 amino acids, and the molecular mass of the mature form was calculated to be 59 kDa. IPU has no amino-acid-sequence similarity with the other pullulan-hydrolysing enzymes, which are pullulanase, neopullulanase and glucoamylase. However, IPU showed a high amino-acid-sequence similarity with dextranases from Penicillium minioluteum (61%) and Arthrobacter sp. (56%). When the ipuA gene was expressed in Aspergillus oryzae, the expressed protein (recombinant IPU) had IPU activity and was immunologically reactive with antibodies raised against native IPU. The substrate specificity, thermostability and pH profile of recombinant IPU were identical with those of the native enzyme, but recombinant IPU (90 kDa) was larger than the native enzyme (69-71 kDa). After deglycosylation with peptide-N-glycosidase F, the deglycosylated recombinant IPU had the same molecular mass as deglycosylated native enzyme (59 kDa). This result suggests that the carbohydrate chain of recombinant IPU differed from that of the native enzyme. PMID:9169610

Resolution and some properties of enzymes involved in enantioselective transformation of 1,3-dichloro-2-propanol to (R)-3-chloro-1,2-propanediol by Corynebacterium sp. strain N-1074.

PubMed Central

Nakamura, T; Nagasawa, T; Yu, F; Watanabe, I; Yamada, H

1992-01-01

During the course of the transformation of 1,3-dichloro-2-propanol (DCP) into (R)-3-chloro-1,2-propanediol [(R)-MCP] with the cell extract of Corynebacterium sp. strain N-1074, epichlorohydrin (ECH) was transiently formed. The cell extract was fractionated into two DCP-dechlorinating activities (fractions Ia and Ib) and two ECH-hydrolyzing activities (fractions IIa and IIb) by TSKgel DEAE-5PW column chromatography. Fractions Ia and Ib catalyzed the interconversion of DCP to ECH, and fractions IIa and IIb catalyzed the transformation of ECH into MCP. Fractions Ia and IIa showed only low enantioselectivity for each reaction, whereas fractions Ib and IIb exhibited considerable enantioselectivity, yielding R-rich ECH and MCP, respectively. Enzymes Ia and Ib were isolated from fractions Ia and Ib, respectively. Enzyme Ia had a molecular mass of about 108 kDa and consisted of four subunits identical in molecular mass (about 28 kDa). Enzyme Ib was a protein of 115 kDa, composed of two different polypeptides (about 35 and 32 kDa). The specific activity of enzyme Ib for DCP was about 30-fold higher than that of enzyme Ia. Both enzymes catalyzed the transformation of several halohydrins into the corresponding epoxides with liberation of halides and its reverse reaction. Their substrate specificities and immunological properties differed from each other. Enzyme Ia seemed to be halohydrin hydrogen-halide-lyase which was already purified from Escherichia coli carrying a gene from Corynebacterium sp. strain N-1074. Images PMID:1447132

Wastewater characterisation by combining size fractionation, chemical composition and biodegradability.

PubMed

Ravndal, Kristin T; Opsahl, Eystein; Bagi, Andrea; Kommedal, Roald

2017-12-18

The potential for resource recovery from wastewater can be evaluated based on a detailed characterisation of wastewater. In this paper, results from fractionation and characterisation of two distinct wastewaters are reported. Using tangential flow filtration, the wastewater was fractionated into 10 size fractions ranging from 1 kDa to 1 mm, wherein the chemical composition and biodegradability were determined. Carbohydrates were dominant in particulate size fractions larger than 100 μm, indicating a potential of cellulose recovery from these fractions. While the particulate size fractions between 0.65 and 100 μm show a potential as a source for biofuel production due to an abundance of saturated C16 and C18 lipids. Both wastewaters were dominated by particulate (>0.65 μm), and oligo- and monomeric (<1 kDa) COD. Polymeric (1-1000 kDa) and colloidal (1000 kDa-0.65 μm) fractions had a low COD content, expected due to degradation in the sewer system upstream of the wastewater treatment plant. Biodegradation rates of particulate fractions increase with decreasing size. However, this was not seen in polymeric fractions where degradation rate was governed by chemical composition. Analytical validation of molecular weight and particle size distribution showed below filter cut-off retention of particles and polymers close to nominal cut-off, shifting the actual size distribution. Copyright © 2017. Published by Elsevier Ltd.

Glycosylated Porphyra-334 and Palythine-Threonine from the Terrestrial Cyanobacterium Nostoc commune

PubMed Central

Nazifi, Ehsan; Wada, Naoki; Yamaba, Minami; Asano, Tomoya; Nishiuchi, Takumi; Matsugo, Seiichi; Sakamoto, Toshio

2013-01-01

Mycosporine-like amino acids (MAAs) are water-soluble UV-absorbing pigments, and structurally different MAAs have been identified in eukaryotic algae and cyanobacteria. In this study novel glycosylated MAAs were found in the terrestrial cyanobacterium Nostoc commune (N. commune). An MAA with an absorption maximum at 334 nm was identified as a hexose-bound porphyra-334 derivative with a molecular mass of 508 Da. Another MAA with an absorption maximum at 322 nm was identified as a two hexose-bound palythine-threonine derivative with a molecular mass of 612 Da. These purified MAAs have radical scavenging activities in vitro, which suggests multifunctional roles as sunscreens and antioxidants. The 612-Da MAA accounted for approximately 60% of the total MAAs and contributed approximately 20% of the total radical scavenging activities in a water extract, indicating that it is the major water-soluble UV-protectant and radical scavenger component. The hexose-bound porphyra-334 derivative and the glycosylated palythine-threonine derivatives were found in a specific genotype of N. commune, suggesting that glycosylated MAA patterns could be a chemotaxonomic marker for the characterization of the morphologically indistinguishable N. commune. The glycosylation of porphyra-334 and palythine-threonine in N. commune suggests a unique adaptation for terrestrial environments that are drastically fluctuating in comparison to stable aquatic environments. PMID:24065157

One Size Does Not Fit All: The Effect of Chain Length and Charge Density of Poly(ethylene imine) Based Copolymers on Delivery of pDNA, mRNA, and RepRNA Polyplexes.

PubMed

Blakney, Anna K; Yilmaz, Gokhan; McKay, Paul F; Becer, C Remzi; Shattock, Robin J

2018-05-03

Nucleic acid delivery systems are commonly translated between different modalities, such as DNA and RNA of varying length and structure, despite physical differences in these molecules that yield disparate delivery efficiency with the same system. Here, we synthesized a library of poly(2-ethyl-2-oxazoline)/poly(ethylene imine) copolymers with varying molar mass and charge densities in order to probe how pDNA, mRNA, and RepRNA polyplex characteristics affect transfection efficiency. The library was utilized in a full factorial design of experiment (DoE) screening, with outputs of luciferase expression, particle size, surface charge, and particle concentration. The optimal copolymer molar mass and charge density was found as 83 kDa/100%, 72 kDa/100%, and 45 kDa/80% for pDNA, RepRNA, and mRNA, respectively. While 10 of the synthesized copolymers enhanced the transfection efficiency of pDNA and mRNA, only 2 copolymers enhanced RepRNA transfection efficiency, indicating a narrow and more stringent design space for RepRNA. These findings suggest that there is not a "one size fits all" polymer for different nucleic acid species.

[Blood plasma protein adsorption capacity of perfluorocarbon emulsion stabilized by proxanol 268 (in vitro and in vivo studies)].

PubMed

Sklifas, A N; Zhalimov, V K; Temnov, A A; Kukushkin, N I

2012-01-01

The adsorption abilities of the perfluorocarbon emulsion stabilized by Proxanol 268 were investigated in vitro and in vivo. In vitro, the saturation point for the blood plasma proteins was nearly reached after five minutes of incubation of the emulsion with human/rabbit blood plasma and was stable for all incubation periods studied. The decrease in volume ratio (emulsion/plasma) was accompanied by the increase in the adsorptive capacity of the emulsion with maximal values at 1/10 (3.2 and 1.5 mg of proteins per 1 ml of the emulsion, for human and rabbit blood plasma, respectively) that was unchanged at lower ratios. In vivo, in rabbits, intravenously injected with the emulsion, the proteins with molecular masses of 12, 25, 32, 44, 55, 70, and 200 kDa were adsorbed by the emulsion (as in vitro) if it was used 6 hours or less before testing. More delayed testing (6 h) revealed elimination of proteins with molecular masses of 25 and 44 kDa and an additional pool of adsorpted new ones of 27, 50, and 150 kDa. Specific adsorptive capacity of the emulsion enhanced gradually after emulsion injection and reached its maximum (3.5-5 mg of proteins per 1 ml of the emulsion) after 24 hours.

FLB1, a human protein of epididymal origin that is involved in the sperm-oocyte recognition process.

PubMed

Boué, F; Duquenne, C; Lassalle, B; Lefèvre, A; Finaz, C

1995-02-01

CA6 antibody was selected out of a monoclonal antibody library raised against human sperm proteins primarily for its ability to recognize an epididymal antigen and to modify sperm adhesion to zona-free hamster oocytes. In the present study, CA6 was shown to decrease sperm binding to zona-free hamster and human oocytes by 40-92% and 38-48%, respectively. The corresponding protein, which was referred to as FLB1, was found to be secreted by the epididymis and to bind specifically to a human, macaque, and rodent subacrosomal sperm region. Western blotting revealed a molecular mass of 94 kDa in human epididymal extracts and of 100 kDa in human, macaque, mouse, rat, and hamster sperm, suggesting further modifications after its binding to sperm. An equivalent protein was not observed in human liver, ovary, testis, plasma, or epidermis. Two-dimensional electrophoresis showed that FLB1 is formed of two subunits with the same 47-kDa molecular mass and slightly different pI (5.8, 5.9). Microsequencing of the protein revealed a partial homology with human cytokeratins 1 and 10. These results suggest that FLB1 is an epididymis-specific cytokeratin-like protein that is involved in the sperm-oocyte recognition process.

Application of surface-enhanced laser desorption/ionization time-of-flight mass spectrometry technology for the diagnosis of colorectal adenoma.

PubMed

Zhou, Zhong-Yin; Tao, DI-DI; Cao, Ji-Wang; Luo, He-Sheng

2013-06-01

The aim of the present study was to identify a specific biological marker for the diagnosis of colorectal adenomas through the analysis of variations in serum protein profiling in colorectal adenoma patients. The study was conducted at the Renmin Hospital of Wuhan University (Wuhan, China) between September 2011 and May 2012. Surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF-MS) was performed to compare the serum protein profiles of 50 patients with colorectal adenoma and 50 healthy individuals. The obtained protein profiles were analyzed using Biomarker Wizard software. Twenty protein peaks were identified to exhibit differences in average intensity between colorectal adenomas compared with normal controls, including peaks 8,565.84, 8,694.51 and 5,910.50 Da, in which the intensity between the patients and control individuals was significantly different. Two peaks, 8,565.84 and 8,694.51 Da, were observed to be highly expressed in the colorectal adenomas, however, expression was low in the control samples. By contrast, 5,910.50 Da expression was low in the colorectal adenomas and high in the controls. The results of the current study indicate that the three protein peaks may represent specific biomarkers for colorectal adenomas.

Electrospray Ionization with High-Resolution Mass Spectrometry as a Tool for Lignomics: Lignin Mass Spectrum Deconvolution

NASA Astrophysics Data System (ADS)

Andrianova, Anastasia A.; DiProspero, Thomas; Geib, Clayton; Smoliakova, Irina P.; Kozliak, Evguenii I.; Kubátová, Alena

2018-05-01

The capability to characterize lignin, lignocellulose, and their degradation products is essential for the development of new renewable feedstocks. Electrospray ionization high-resolution time-of-flight mass spectrometry (ESI-HR TOF-MS) method was developed expanding the lignomics toolkit while targeting the simultaneous detection of low and high molecular weight (MW) lignin species. The effect of a broad range of electrolytes and various ionization conditions on ion formation and ionization effectiveness was studied using a suite of mono-, di-, and triarene lignin model compounds as well as kraft alkali lignin. Contrary to the previous studies, the positive ionization mode was found to be more effective for methoxy-substituted arenes and polyphenols, i.e., species of a broadly varied MW structurally similar to the native lignin. For the first time, we report an effective formation of multiply charged species of lignin with the subsequent mass spectrum deconvolution in the presence of 100 mmol L-1 formic acid in the positive ESI mode. The developed method enabled the detection of lignin species with an MW between 150 and 9000 Da or higher, depending on the mass analyzer. The obtained M n and M w values of 1500 and 2500 Da, respectively, were in good agreement with those determined by gel permeation chromatography. Furthermore, the deconvoluted ESI mass spectrum was similar to that obtained with matrix-assisted laser desorption/ionization (MALDI)-HR TOF-MS, yet featuring a higher signal-to-noise ratio. The formation of multiply charged species was confirmed with ion mobility ESI-HR Q-TOF-MS. [Figure not available: see fulltext.

Mass Spectrometric Identification of Phospholipids in Human Tears and Tear Lipocalin

PubMed Central

Dean, Austin W.; Glasgow, Ben J.

2012-01-01

Purpose. The purpose of this article was to identify by mass spectrometry phosphocholine lipids in stimulated human tears and determine the molecules bound to tear lipocalin or other proteins. Methods. Tear proteins were separated isocratically from pooled stimulated human tears by gel filtration fast performance liquid chromatography. Separation of tear lipocalin was confirmed by SDS tricine gradient PAGE. Protein fractions were extracted with chloroform/methanol and analyzed with electrospray ionization MS/MS triple quadrupole mass spectrometry in precursor ion scan mode for select leaving groups. For quantification, integrated ion counts were derived from standard curves of authentic compounds of phosphatidylcholine (PC) and phosphatidylserine. Results. Linear approximation was possible from integration of the mass spectrometrically obtained ion peaks at 760 Da for the PC standard. Tears contained 194 ng/mL of the major intact PC (34:2), m/z 758.6. Ten other monoisotopic phosphocholines were found in tears. A peak at 703.3 Da was assigned as a sphingomyelin. Four lysophosphatidylcholines (m/z 490–540) accounted for about 80% of the total integrated ion count. The [M+H]+ compound, m/z 496.3, accounted for 60% of the signal intensity. Only the tear lipocalin–bearing fractions showed phosphocholines (104 ng/mL). Although the intact phospholipids bound to tear lipocalin corresponded precisely in mass and relative signal intensity to that found in tears, we did not identify phosphocholines between m/z 490 and 540 in any of the gel-filtration fractions. Conclusions. Phospholipids, predominantly lysophospholipids, are present in tears. The higher mass intact PCs in tears are native ligands of tear lipocalin. PMID:22395887

Mass spectrometric identification of phospholipids in human tears and tear lipocalin.

PubMed

Dean, Austin W; Glasgow, Ben J

2012-04-02

The purpose of this article was to identify by mass spectrometry phosphocholine lipids in stimulated human tears and determine the molecules bound to tear lipocalin or other proteins. Tear proteins were separated isocratically from pooled stimulated human tears by gel filtration fast performance liquid chromatography. Separation of tear lipocalin was confirmed by SDS tricine gradient PAGE. Protein fractions were extracted with chloroform/methanol and analyzed with electrospray ionization MS/MS triple quadrupole mass spectrometry in precursor ion scan mode for select leaving groups. For quantification, integrated ion counts were derived from standard curves of authentic compounds of phosphatidylcholine (PC) and phosphatidylserine. Linear approximation was possible from integration of the mass spectrometrically obtained ion peaks at 760 Da for the PC standard. Tears contained 194 ng/mL of the major intact PC (34:2), m/z 758.6. Ten other monoisotopic phosphocholines were found in tears. A peak at 703.3 Da was assigned as a sphingomyelin. Four lysophosphatidylcholines (m/z 490-540) accounted for about 80% of the total integrated ion count. The [M+H](+) compound, m/z 496.3, accounted for 60% of the signal intensity. Only the tear lipocalin-bearing fractions showed phosphocholines (104 ng/mL). Although the intact phospholipids bound to tear lipocalin corresponded precisely in mass and relative signal intensity to that found in tears, we did not identify phosphocholines between m/z 490 and 540 in any of the gel-filtration fractions. Phospholipids, predominantly lysophospholipids, are present in tears. The higher mass intact PCs in tears are native ligands of tear lipocalin.

Electrospray Ionization with High-Resolution Mass Spectrometry as a Tool for Lignomics: Lignin Mass Spectrum Deconvolution

NASA Astrophysics Data System (ADS)

Andrianova, Anastasia A.; DiProspero, Thomas; Geib, Clayton; Smoliakova, Irina P.; Kozliak, Evguenii I.; Kubátová, Alena

2018-03-01

The capability to characterize lignin, lignocellulose, and their degradation products is essential for the development of new renewable feedstocks. Electrospray ionization high-resolution time-of-flight mass spectrometry (ESI-HR TOF-MS) method was developed expanding the lignomics toolkit while targeting the simultaneous detection of low and high molecular weight (MW) lignin species. The effect of a broad range of electrolytes and various ionization conditions on ion formation and ionization effectiveness was studied using a suite of mono-, di-, and triarene lignin model compounds as well as kraft alkali lignin. Contrary to the previous studies, the positive ionization mode was found to be more effective for methoxy-substituted arenes and polyphenols, i.e., species of a broadly varied MW structurally similar to the native lignin. For the first time, we report an effective formation of multiply charged species of lignin with the subsequent mass spectrum deconvolution in the presence of 100 mmol L-1 formic acid in the positive ESI mode. The developed method enabled the detection of lignin species with an MW between 150 and 9000 Da or higher, depending on the mass analyzer. The obtained M n and M w values of 1500 and 2500 Da, respectively, were in good agreement with those determined by gel permeation chromatography. Furthermore, the deconvoluted ESI mass spectrum was similar to that obtained with matrix-assisted laser desorption/ionization (MALDI)-HR TOF-MS, yet featuring a higher signal-to-noise ratio. The formation of multiply charged species was confirmed with ion mobility ESI-HR Q-TOF-MS. [Figure not available: see fulltext.

Relationships between Molecular Composition and Optical Properties of Dissolved Organic Matter

NASA Astrophysics Data System (ADS)

Cooper, W. T.; Tfaily, M.; Osborne, D.; Paul, A.; Podgorski, D. C.; Corbett, J.; Chanton, J.

2009-12-01

Our focus is on the relationships between the optical properties of dissolved organic matter (DOM) and its molecular composition. For example, we demonstrated that changes in the absorption and fluorescence characteristics of DOM in outwelling from Brazilian mangrove forests correlated with decreases in highly unsaturated organic compounds as DOM was transported from mangrove porewaters to the continental shelf. In that work we combined ultrahigh resolution Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) that provided detailed compositional information with absorption and Excitation/Emission Matrix (EEM) spectroscopy This presentation will highlight new results utilizing the combination of optical spectroscopy and FT-ICR mass spectrometry to illuminate the processes which control DOM cycling. Our focus will be on the contributions of the heteroatom components of DOM (i.e. organic sulfur and organic nitrogen) to its optical properties and how changes in optical properties correlate with important environmental processes like humification and bioavailability. Figure 1 below includes a narrow 0.20 Dalton window from a mass spectrum which demonstrates the ability of ultrahigh resolution mass spectrometry to resolve and identify nitrogen heteroatom compounds in DOM. Our study sites include the Glacial Lake Agassiz Peatlands (GLAP) in northern Minnesota and wetlands in the Caloosahatchee River basin in South Florida. Figure 1. Isolated 0.20 Da window of an ESI-FT-ICR mass spectrum of DOM from a GLAP bog. Labels identify N1 (d,e,f) and N3 classes of nitrogen heteroatoms. The 0.0031 Da mass spacing is used to confirm the N3 class.

Normal-phase liquid chromatography retention behavior of polycyclic aromatic sulfur heterocycles and alkyl-substituted polycyclic aromatic sulfur heterocycle isomers on an aminopropyl stationary phase.

PubMed

Wilson, Walter B; Hayes, Hugh V; Sander, Lane C; Campiglia, Andres D; Wise, Stephen A

2018-02-01

Retention indices for 67 polycyclic aromatic sulfur heterocycles (PASHs) and 80 alkyl-substituted PASHs were determined using normal-phase liquid chromatography (NPLC) on an aminopropyl (NH 2 ) stationary phase. The retention behavior of PASH on the NH 2 phase is correlated with the number of aromatic carbon atoms and two structural characteristics have a significant influence on their retention: non-planarity (thickness, T) and the position of the sulfur atom in the bay-region of the structure. Correlations between solute retention on the NH 2 phase and T of PASHs were investigated for three cata-condensed (cata-) PASH isomer groups: (a) 13 four-ring molecular mass (MM) 234 Da cata-PASHs, (b) 20 five-ring MM 284 Da cata-PASHs, and (c) 12 six-ring MM 334 Da cata-PASHs. Correlation coefficients ranged from r = -0.49 (MM 234 Da) to r = -0.65 (MM 334 Da), which were significantly lower than structurally similar PAH isomer groups (r = -0.70 to r = -0.99). The NPLC retention behavior of the PASHs are compared to similar results for PAHs.

Molecular cloning, expression and characterization of a serine proteinase inhibitor gene from Entamoeba histolytica.

PubMed

Riahi, Yael; Siman-Tov, Rama; Ankri, Serge

2004-02-01

Serine proteinase inhibitors (serpins) are irreversible suicide inhibitors of proteinases that regulate a wide range of biological processes, including pathogen evasion of the host defence system. We report the cloning and characterization of a gene encoding a serpin from the protozoan parasite Entamoeba histolytica (Ehserp) that may function in this manner. The protein encoded by Ehserp contains 371 amino acids with a predicted mass of 42.6 kDa. Antibodies to a 42 kDa recombinant Ehserp react specifically with two bands of 42 and 49 kDa in trophozoite extracts. Ehserp has a cytoplasmic localization and is secreted by trophozoites incubated in the presence of mammalian cells, but not by resting trophozoites. A panel of mammalian serine proteinases was screened, but none of them was inhibited by the recombinant Ehserp. In contrast, the 49 kDa Ehserp present in the secretion product (SP) of activated macrophages interacted with human neutrophil cathepsin G to form a complex resistant to sodium dodecyl sulphate. We discuss the nature of the 42 and 49 kDa Ehserp and the possible roles that Ehserp may play in the survival of the parasite inside the host.

FoxO1 in dopaminergic neurons regulates energy homeostasis and targets tyrosine hydroxylase

PubMed Central

Doan, Khanh V.; Kinyua, Ann W.; Yang, Dong Joo; Ko, Chang Mann; Moh, Sang Hyun; Shong, Ko Eun; Kim, Hail; Park, Sang-Kyu; Kim, Dong-Hoon; Kim, Inki; Paik, Ji-Hye; DePinho, Ronald A.; Yoon, Seul Gi; Kim, Il Yong; Seong, Je Kyung; Choi, Yun-Hee; Kim, Ki Woo

2016-01-01

Dopaminergic (DA) neurons are involved in the integration of neuronal and hormonal signals to regulate food consumption and energy balance. Forkhead transcriptional factor O1 (FoxO1) in the hypothalamus plays a crucial role in mediation of leptin and insulin function. However, the homoeostatic role of FoxO1 in DA system has not been investigated. Here we report that FoxO1 is highly expressed in DA neurons and mice lacking FoxO1 specifically in the DA neurons (FoxO1 KODAT) show markedly increased energy expenditure and interscapular brown adipose tissue (iBAT) thermogenesis accompanied by reduced fat mass and improved glucose/insulin homoeostasis. Moreover, FoxO1 KODAT mice exhibit an increased sucrose preference in concomitance with higher dopamine and norepinephrine levels. Finally, we found that FoxO1 directly targets and negatively regulates tyrosine hydroxylase (TH) expression, the rate-limiting enzyme of the catecholamine synthesis, delineating a mechanism for the KO phenotypes. Collectively, these results suggest that FoxO1 in DA neurons is an important transcriptional factor that directs the coordinated control of energy balance, thermogenesis and glucose homoeostasis. PMID:27681312

Current use of high-resolution mass spectrometry in drug screening relevant to clinical and forensic toxicology and doping control.

PubMed

Ojanperä, Ilkka; Kolmonen, Marjo; Pelander, Anna

2012-05-01

Clinical and forensic toxicology and doping control deal with hundreds or thousands of drugs that may cause poisoning or are abused, are illicit, or are prohibited in sports. Rapid and reliable screening for all these compounds of different chemical and pharmaceutical nature, preferably in a single analytical method, is a substantial effort for analytical toxicologists. Combined chromatography-mass spectrometry techniques with standardised reference libraries have been most commonly used for the purpose. In the last ten years, the focus has shifted from gas chromatography-mass spectrometry to liquid chromatography-mass spectrometry, because of progress in instrument technology and partly because of the polarity and low volatility of many new relevant substances. High-resolution mass spectrometry (HRMS), which enables accurate mass measurement at high resolving power, has recently evolved to the stage that is rapidly causing a shift from unit-resolution, quadrupole-dominated instrumentation. The main HRMS techniques today are time-of-flight mass spectrometry and Orbitrap Fourier-transform mass spectrometry. Both techniques enable a range of different drug-screening strategies that essentially rely on measuring a compound's or a fragment's mass with sufficiently high accuracy that its elemental composition can be determined directly. Accurate mass and isotopic pattern acts as a filter for confirming the identity of a compound or even identification of an unknown. High mass resolution is essential for improving confidence in accurate mass results in the analysis of complex biological samples. This review discusses recent applications of HRMS in analytical toxicology.

Top-down MALDI-in-source decay-FTICR mass spectrometry of isotopically resolved proteins.

PubMed

Nicolardi, Simone; Switzar, Linda; Deelder, André M; Palmblad, Magnus; van der Burgt, Yuri E M

2015-03-17

An accurate mass measurement of a known protein provides information on potential amino acid deletions and post-translational modifications. Although this field is dominated by strategies based on electrospray ionization, mass spectrometry (MS) methods using matrix-assisted laser desorption/ionization (MALDI) have the advantage of yielding predominantly singly charged precursor ions, thus avoiding peak overlap from different charge states of multiple species. Such MALDI-MS methods require mass measurement at ultrahigh resolution, which is provided by Fourier transform ion cyclotron resonance (FTICR) mass analyzers. Recently, using a MALDI-FTICR-MS platform equipped with a 15 T magnet, we reported on the mass analysis of intact human serum peptides and small proteins with isotopic resolution up to ∼15 kDa and identified new proteoforms from an accurate measurement of mass distances. In the current study, we have used this FTICR system after an upgrade with a novel dynamically harmonized ICR cell, i.e., ParaCell, for mapping isotopically resolved intact proteins up to about 17 kDa and performed top-down MALDI in-source decay (ISD) analysis. Standard proteins myoglobin (m/z-value 16,950) and ribonuclease B (m/z-value 14,900) were measured with resolving powers of 62,000 and 61,000, respectively. Furthermore, it will be shown that (singly charged) MALDI-ISD fragment ions can be measured at isotopic resolution up to m/z-value 12,000 (e.g., resolving power 39,000 at m/z-value 12,000) providing more reliable identifications. Moreover, examples are presented of pseudo-MS(3) experiments on ISD fragment ions from RNase B by collisional-induced dissociation (CID).

Astrophysical Implications of a New Dynamical Mass for the Nearby White Dwarf 40 Eridani B

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

Bond, Howard E.; Bergeron, P.; Bédard, A., E-mail: heb11@psu.edu

The bright, nearby DA-type white dwarf (WD) 40 Eridani B is orbited by the M dwarf 40 Eri C, allowing determination of the WD’s mass. Until recently, however, the mass depended on orbital elements determined four decades ago, and that mass was so low that it created several astrophysical puzzles. Using new astrometric measurements, the binary-star group at the U.S. Naval Observatory has revised the dynamical mass upward, to 0.573 ± 0.018 M {sub ☉}. In this paper, we use model-atmosphere analysis to update other parameters of the WD, including effective temperature, surface gravity, radius, and luminosity. We then comparemore » these results with WD interior models. Within the observational uncertainties, theoretical cooling tracks for CO-core WDs of its measured mass are consistent with the position of 40 Eri B in the H-R diagram; equivalently, the theoretical mass–radius relation (MRR) is consistent with the star’s location in the mass–radius plane. This consistency is, however, achieved only if we assume a “thin” outer hydrogen layer, with q {sub H} = M {sub H}/ M {sub WD} ≃ 10{sup −10}. We discuss other evidence that a significant fraction of DA WDs have such thin H layers, in spite of the expectation from canonical stellar-evolution theory of “thick” H layers with q {sub H} ≃ 10{sup −4}. The cooling age of 40 Eri B is ∼122 Myr, and its total age is ∼1.8 Gyr. We present the MRRs for 40 Eri B and three other nearby WDs in visual binaries with precise mass determinations, and show that the agreement of current theory with observations is excellent in all cases.« less

Nozzle Aerodynamic Stability During a Throat Shift

NASA Technical Reports Server (NTRS)

Kawecki, Edwin J.; Ribeiro, Gregg L.

2005-01-01

An experimental investigation was conducted on the internal aerodynamic stability of a family of two-dimensional (2-D) High Speed Civil Transport (HSCT) nozzle concepts. These nozzles function during takeoff as mixer-ejectors to meet acoustic requirements, and then convert to conventional high-performance convergent-divergent (CD) nozzles at cruise. The transition between takeoff mode and cruise mode results in the aerodynamic throat and the minimum cross-sectional area that controls the engine backpressure shifting location within the nozzle. The stability and steadiness of the nozzle aerodynamics during this so called throat shift process can directly affect the engine aerodynamic stability, and the mechanical design of the nozzle. The objective of the study was to determine if pressure spikes or other perturbations occurred during the throat shift process and, if so, identify the caused mechanisms for the perturbations. The two nozzle concepts modeled in the test program were the fixed chute (FC) and downstream mixer (DSM). These 2-D nozzles differ principally in that the FC has a large over-area between the forward throat and aft throat locations, while the DSM has an over-area of only about 10 percent. The conclusions were that engine mass flow and backpressure can be held constant simultaneously during nozzle throat shifts on this class of nozzles, and mode shifts can be accomplished at a constant mass flow and engine backpressure without upstream pressure perturbations.

Myelin management by the 18.5–kDa and 21.5–kDa classic myelin basic protein isoforms

PubMed Central

Harauz, George; Boggs, Joan M.

2013-01-01

The classic myelin basic protein (MBP) splice isoforms range in nominal molecular mass from 14 to 21.5 kDa, and arise from the gene in the oligodendrocyte lineage (Golli) in maturing oligodendrocytes. The 18.5-kDa isoform that predominates in adult myelin adheres the cytosolic surfaces of oligodendrocyte membranes together, and forms a two-dimensional molecular sieve restricting protein diffusion into compact myelin. However, this protein has additional roles including cytoskeletal assembly and membrane extension, binding to SH3-domains, participation in Fyn-mediated signaling pathways, sequestration of phosphoinositides, and maintenance of calcium homeostasis. Of the diverse post-translational modifications of this isoform, phosphorylation is the most dynamic, and modulates 18.5-kDa MBP’s protein-membrane and protein-protein interactions, indicative of a rich repertoire of functions. In developing and mature myelin, phosphorylation can result in microdomain or even nuclear targeting of the protein, supporting the conclusion that 18.5-kDa MBP has significant roles beyond membrane adhesion. The full-length, early-developmental 21.5-kDa splice isoform is predominantly karyophilic due to a non-traditional P-Y nuclear localization signal, with effects such as promotion of oligodendrocyte proliferation. We discuss in vitro and recent in vivo evidence for multifunctionality of these classic basic proteins of myelin, and argue for a systematic evaluation of the temporal and spatial distributions of these protein isoforms, and their modified variants, during oligodendrocyte differentiation. PMID:23398367

A technique for rapid source apportionment applied to ambient organic aerosol measurements from a thermal desorption aerosol gas chromatograph (TAG)

DOE PAGES

Zhang, Yaping; Williams, Brent J.; Goldstein, Allen H.; ...

2016-11-25

Here, we present a rapid method for apportioning the sources of atmospheric organic aerosol composition measured by gas chromatography–mass spectrometry methods. Here, we specifically apply this new analysis method to data acquired on a thermal desorption aerosol gas chromatograph (TAG) system. Gas chromatograms are divided by retention time into evenly spaced bins, within which the mass spectra are summed. A previous chromatogram binning method was introduced for the purpose of chromatogram structure deconvolution (e.g., major compound classes) (Zhang et al., 2014). Here we extend the method development for the specific purpose of determining aerosol samples' sources. Chromatogram bins are arrangedmore » into an input data matrix for positive matrix factorization (PMF), where the sample number is the row dimension and the mass-spectra-resolved eluting time intervals (bins) are the column dimension. Then two-dimensional PMF can effectively do three-dimensional factorization on the three-dimensional TAG mass spectra data. The retention time shift of the chromatogram is corrected by applying the median values of the different peaks' shifts. Bin width affects chemical resolution but does not affect PMF retrieval of the sources' time variations for low-factor solutions. A bin width smaller than the maximum retention shift among all samples requires retention time shift correction. A six-factor PMF comparison among aerosol mass spectrometry (AMS), TAG binning, and conventional TAG compound integration methods shows that the TAG binning method performs similarly to the integration method. However, the new binning method incorporates the entirety of the data set and requires significantly less pre-processing of the data than conventional single compound identification and integration. In addition, while a fraction of the most oxygenated aerosol does not elute through an underivatized TAG analysis, the TAG binning method does have the ability to achieve molecular level resolution on other bulk aerosol components commonly observed by the AMS.« less

A technique for rapid source apportionment applied to ambient organic aerosol measurements from a thermal desorption aerosol gas chromatograph (TAG)

NASA Astrophysics Data System (ADS)

Zhang, Yaping; Williams, Brent J.; Goldstein, Allen H.; Docherty, Kenneth S.; Jimenez, Jose L.

2016-11-01

We present a rapid method for apportioning the sources of atmospheric organic aerosol composition measured by gas chromatography-mass spectrometry methods. Here, we specifically apply this new analysis method to data acquired on a thermal desorption aerosol gas chromatograph (TAG) system. Gas chromatograms are divided by retention time into evenly spaced bins, within which the mass spectra are summed. A previous chromatogram binning method was introduced for the purpose of chromatogram structure deconvolution (e.g., major compound classes) (Zhang et al., 2014). Here we extend the method development for the specific purpose of determining aerosol samples' sources. Chromatogram bins are arranged into an input data matrix for positive matrix factorization (PMF), where the sample number is the row dimension and the mass-spectra-resolved eluting time intervals (bins) are the column dimension. Then two-dimensional PMF can effectively do three-dimensional factorization on the three-dimensional TAG mass spectra data. The retention time shift of the chromatogram is corrected by applying the median values of the different peaks' shifts. Bin width affects chemical resolution but does not affect PMF retrieval of the sources' time variations for low-factor solutions. A bin width smaller than the maximum retention shift among all samples requires retention time shift correction. A six-factor PMF comparison among aerosol mass spectrometry (AMS), TAG binning, and conventional TAG compound integration methods shows that the TAG binning method performs similarly to the integration method. However, the new binning method incorporates the entirety of the data set and requires significantly less pre-processing of the data than conventional single compound identification and integration. In addition, while a fraction of the most oxygenated aerosol does not elute through an underivatized TAG analysis, the TAG binning method does have the ability to achieve molecular level resolution on other bulk aerosol components commonly observed by the AMS.

REVEAL: Reconstruction, Enhancement, Visualization, and Ergonomic Assessment for Laparoscopy

DTIC Science & Technology

2008-08-01

measurable disparity shift. Such an endoscope can be used to generate a stereoscopic view for a surgeon, as with the DaVinci robot in use today...training or surgery. We are working on the user interface issues of incorporating this measurement capability into the standard set of tools during...scope use, and in structuring a set of tasks around the use of through-the-scope measurement in order to determine how this tool can affect efficiency

Cultivar variability of patatin biochemical characteristics: table versus processing potatoes (Solanum tuberosum L.).

PubMed

Bárta, Jan; Bártová, Veronika; Zdráhal, Zbyněk; Sedo, Ondrej

2012-05-02

Biochemical characteristics of patatin proteins purified by ion-exchange and affinity chromatography from tubers of 20 potato cultivars were studied to evaluate their genotype differences with respect to utility groups, table potato cultivars (TPCs) and processing potato cultivars (PPCs). Both groups of cultivars showed similar values of protein content in dry matter (3.98-7.39%) and of patatin relative abundance (5.40-35.40%). Three mass levels (∼40.6, 41.8, and 42.9 kDa) of purified patatins were found by MALDI-TOF MS within all cultivars. Differences among mass levels corresponding with the mass of sugar antenna (∼1.2 kDa) confirmed the previous concept of different glycosylation extentsin patatin proteins. It was showed that the individual types of patatin varying in their masses occur in the patatin family in a ratio specific for each of the cultivars, with the lowest mass type being the major one. Electrophoretic analyses demonstrated wide cultivar variability in number of patatin forms. Especially 2D-PAGE showed 17-23 detected protein spots independently on the utility group. Specific lipid acyl hydrolase (LAH) activity of purified patatins from the individual tested cultivars varied between 0.92 and 5.46 μmol/(min mg). Patatin samples within most of the TPCs exhibited higher values of specific LAH activity than samples of PPCs. It may be supposed that individual patatin forms do not have similar physiological roles.

CMB seen through random Swiss Cheese

NASA Astrophysics Data System (ADS)

Lavinto, Mikko; Räsänen, Syksy

2015-10-01

We consider a Swiss Cheese model with a random arrangement of Lemaȋtre-Tolman-Bondi holes in ΛCDM cheese. We study two kinds of holes with radius rb=50 h-1 Mpc, with either an underdense or an overdense centre, called the open and closed case, respectively. We calculate the effect of the holes on the temperature, angular diameter distance and, for the first time in Swiss Cheese models, shear of the CMB . We quantify the systematic shift of the mean and the statistical scatter, and calculate the power spectra. In the open case, the temperature power spectrum is three orders of magnitude below the linear ISW spectrum. It is sensitive to the details of the hole, in the closed case the amplitude is two orders of magnitude smaller. In contrast, the power spectra of the distance and shear are more robust, and agree with perturbation theory and previous Swiss Cheese results. We do not find a statistically significant mean shift in the sky average of the angular diameter distance, and obtain the 95% limit |Δ DA/bar DA|lesssim 10-4. We consider the argument that areas of spherical surfaces are nearly unaffected by perturbations, which is often invoked in light propagation calculations. The closed case is consistent with this at 1σ, whereas in the open case the probability is only 1.4%.

General contrast effects in speech perception: effect of preceding liquid on stop consonant identification.

PubMed

Lotto, A J; Kluender, K R

1998-05-01

When members of a series of synthesized stop consonants varying acoustically in F3 characteristics and varying perceptually from /da/ to /ga/ are preceded by /al/, subjects report hearing more /ga/ syllables relative to when each member is preceded by /ar/ (Mann, 1980). It has been suggested that this result demonstrates the existence of a mechanism that compensates for coarticulation via tacit knowledge of articulatory dynamics and constraints, or through perceptual recovery of vocal-tract dynamics. The present study was designed to assess the degree to which these perceptual effects are specific to qualities of human articulatory sources. In three experiments, series of consonant-vowel (CV) stimuli varying in F3-onset frequency (/da/-/ga/) were preceded by speech versions or nonspeech analogues of /al/ and /ar/. The effect of liquid identity on stop consonant labeling remained when the preceding VC was produced by a female speaker and the CV syllable was modeled after a male speaker's productions. Labeling boundaries also shifted when the CV was preceded by a sine wave glide modeled after F3 characteristics of /al/ and /ar/. Identifications shifted even when the preceding sine wave was of constant frequency equal to the offset frequency of F3 from a natural production. These results suggest an explanation in terms of general auditory processes as opposed to recovery of or knowledge of specific articulatory dynamics.

Molecular weight dependent charge carrier mobility in poly(3,3' '-dioctyl-2,2':5',2' '-terthiophene).

PubMed

Verilhac, Jean-Marie; Pokrop, Rafal; LeBlevennec, Gilles; Kulszewicz-Bajer, Irena; Buga, Katarzyna; Zagorska, Malgorzata; Sadki, Said; Pron, Adam

2006-07-13

Poly(3,3' '-dioctyl-2,2':5',2' '-terthiophene), a polymer recently used for the fabrication of organic field effect transistors, has been fractionated into five fractions distinctly differing in their molecular weights (Mn), with the goal of determining the influence of the degree of polymerization (DPn) on its principal physicochemical parameters. It has been demonstrated that within the Mn range studied (from 1.5 kDa to 10.5 kDa by SEC), corresponding to DPn from 10 to 38, the polymer band gap steadily decreases with growing molecular weight, which is clearly manifested by an increasing bathochromic shift of the band originating from the pi-pi* transition. The same trend is observed for the HOMO level, determined from the onset of the p-doping in cyclic voltammetry, which shifts from -5.10 eV to -4.90 eV for the lowest and the highest molecular weight fractions, respectively. The most pronounced influence of DPn has been found for the charge carriers' mobility-one of the most important parameters of field effect transistors (FETs) fabricated from this polymer. A fourfold increase in DPn results in an increase of the carriers' mobility by more than 3 orders of magnitude. Comparison of these results with those obtained for fractionated regioregular poly(3-hexylthiophene) shows a strikingly similar behavior of both polymers with respect to the molecular weight.

Biodiversity of Exopolysaccharides Produced by Streptococcus thermophilus Strains Is Reflected in Their Production and Their Molecular and Functional Characteristics

PubMed Central

Vaningelgem, Frederik; Zamfir, Medana; Mozzi, Fernanda; Adriany, Tom; Vancanneyt, Marc; Swings, Jean; De Vuyst, Luc

2004-01-01

Twenty-six lactic acid bacterium strains isolated from European dairy products were identified as Streptococcus thermophilus and characterized by bacterial growth and exopolysaccharide (EPS)-producing capacity in milk and enriched milk medium. In addition, the acidification rates of the different strains were compared with their milk clotting behaviors. The majority of the strains grew better when yeast extract and peptone were added to the milk medium, although the presence of interfering glucomannans was shown, making this medium unsuitable for EPS screening. EPS production was found to be strain dependent, with the majority of the strains producing between 20 and 100 mg of polymer dry mass per liter of fermented milk medium. Furthermore, no straightforward relationship between the apparent viscosity and EPS production could be detected in fermented milk medium. An analysis of the molecular masses of the isolated EPS by gel permeation chromatography revealed a large variety, ranging from 10 to >2,000 kDa. A distinction could be made between high-molecular-mass EPS (>1,000 kDa) and low-molecular-mass EPS (<1,000 kDa). Based on the molecular size of the EPS, three groups of EPS-producing strains were distinguished. Monomer analysis of the EPS by high-performance anion-exchange chromatography with amperometric detection was demonstrated to be a fast and simple method. All of the EPS from the S. thermophilus strains tested were classified into six groups according to their monomer compositions. Apart from galactose and glucose, other monomers, such as (N-acetyl)galactosamine, (N-acetyl)glucosamine, and rhamnose, were also found as repeating unit constituents. Three strains were found to produce EPS containing (N-acetyl)glucosamine, which to our knowledge was never found before in an EPS from S. thermophilus. Furthermore, within each group, differences in monomer ratios were observed, indicating possible novel EPS structures. Finally, large differences between the consistencies of EPS solutions from five different strains were assigned to differences in their molecular masses and structures. PMID:14766570

Venoms of Centruroides and Tityus species from Panama and their main toxic fractions.

PubMed

Salazar, Marcos H; Arenas, Iván; Corrales-García, Ligia L; Miranda, Roberto; Vélez, Sara; Sánchez, Jairo; Mendoza, Karla; Cleghorn, John; Zamudio, Fernando Z; Castillo, Adolfo; Possani, Lourival D; Corzo, Gerardo; Acosta, Hildaura

2018-01-01

The scorpionism in Panama is notorious for the confluence and coexistence of buthid scorpions from the genera Centruroides and Tityus. This communication describes an overview of the larger representative toxic venom fractions from eight dangerous buthid scorpion species of Panama: Centruroides (C. granosus, C. bicolor, C. limbatus and C. panamensis) and Tityus (T. (A.) asthenes, T. (A.) festae, T. (T.) cerroazul and T. (A.) pachyurus). Their venoms were separated by HPLC and the corresponding sub-fractions were tested for lethality effects on mice and insects. Many fractions toxic to either mice or insects, or both, were found and have had their molecular masses determined by mass spectrometry analysis. The great majority of the lethal components had a molecular mass close to 7000 Da, assumed to be peptides that recognize Na + -channels, responsible for the toxicity symptoms observed in other buthids scorpion venoms. A toxic peptide isolated from the venom of T. pachyurus was sequenced by Edman degradation, allowing the synthesis of nucleotide probe for cloning the correspondent gene. The mature toxin based on the cDNA sequencing has the C-terminal residue amidated, contains 62 amino acid packed by 4 disulfide linkages, with molecular mass of 7099.1 Da. This same toxic peptide seems to be present in scorpions of the species T. pachyurus collected in 5 different regions of Panama, although the overall HPLC profile is quite different. The most diverse neurotoxic venom components from the genus Centruroides were found in the species C. panamensis, whereas T. cerroazul was the one from the genus Tityus. The most common neurotoxins were observed in the venoms of T. festae, T. asthenes and T. pachyurus with closely related molecular masses of 7099.1 and 7332 Da. The information reported here is considered very important for future generation of a neutralizing antivenom against scorpions from Panama. Furthermore, it will contribute to the growing interest in using bioactive toxins from scorpions for drug discovery purposes. Copyright © 2017 Elsevier Ltd. All rights reserved.

Connecting optical and X-ray tracers of galaxy cluster relaxation

NASA Astrophysics Data System (ADS)

Roberts, Ian D.; Parker, Laura C.; Hlavacek-Larrondo, Julie

2018-04-01

Substantial effort has been devoted in determining the ideal proxy for quantifying the morphology of the hot intracluster medium in clusters of galaxies. These proxies, based on X-ray emission, typically require expensive, high-quality X-ray observations making them difficult to apply to large surveys of groups and clusters. Here, we compare optical relaxation proxies with X-ray asymmetries and centroid shifts for a sample of Sloan Digital Sky Survey clusters with high-quality, archival X-ray data from Chandra and XMM-Newton. The three optical relaxation measures considered are the shape of the member-galaxy projected velocity distribution - measured by the Anderson-Darling (AD) statistic, the stellar mass gap between the most-massive and second-most-massive cluster galaxy, and the offset between the most-massive galaxy (MMG) position and the luminosity-weighted cluster centre. The AD statistic and stellar mass gap correlate significantly with X-ray relaxation proxies, with the AD statistic being the stronger correlator. Conversely, we find no evidence for a correlation between X-ray asymmetry or centroid shift and the MMG offset. High-mass clusters (Mhalo > 1014.5 M⊙) in this sample have X-ray asymmetries, centroid shifts, and Anderson-Darling statistics which are systematically larger than for low-mass systems. Finally, considering the dichotomy of Gaussian and non-Gaussian clusters (measured by the AD test), we show that the probability of being a non-Gaussian cluster correlates significantly with X-ray asymmetry but only shows a marginal correlation with centroid shift. These results confirm the shape of the radial velocity distribution as a useful proxy for cluster relaxation, which can then be applied to large redshift surveys lacking extensive X-ray coverage.

CHARACTERIZATION OF THE HEMOLYSIN, FROM STACHYBOTRYS CHARTARUM

EPA Science Inventory

Stachybotrys chartarum is a toxigenic fungus that has been associated with human health concerns, including pulmonary hemorrhage and hemosiderosis. This fungus produces a hemolysin, stachylysin, which in its apparent monomeric form has a molecular mass of 11,920
Da as determ...

Structural characterization and identification of iridoid glycosides, saponins, phenolic acids and flavonoids in Flos Lonicerae Japonicae by a fast liquid chromatography method with diode-array detection and time-of-flight mass spectrometry.

PubMed

Qi, Lian-Wen; Chen, Chun-Yun; Li, Ping

2009-10-01

A fast liquid chromatography method with diode-array detection (DAD) and time-of-flight mass spectrometry (TOF-MS) has been developed for analysis of constituents in Flos Lonicerae Japonicae (FLJ), a traditional Chinese medicine derived from the flower bud of Lonicera japonica. The chromatographic analytical time decreased to 25 min without sacrificing resolution using a column packed with 1.8-microm porous particles (4.6 x 50 mm), three times faster than the performance of conventional 5.0-microm columns (4.6 x 150 mm). Four major groups of compounds previously isolated from FLJ were structurally characterized by DAD-TOF-MS: iridoid glycosides showed maximum UV absorption at 240 nm; phenolic acids at 217, 242, and 326 nm; flavonoids at 255 and 355 nm; while saponins had no absorption. In electrospray ionization (ESI)-TOF-MS experiments, elimination of a glucose unit (162 Da), and successive losses of H(2)O, CH(3)OH and CO, were generally observed in iridoid glycosides; saponins were characterized by a series of identical aglycone ions; phenolic acids typically generated a base peak at [M-H-caffeoyl](-) by loss of a caffeic acid unit (162 Da) and several marked quinic acid moiety ions; cleavage of the glycosidic bond (loss of 162 or 308 Da), subsequent losses of H(2)O, CO, RDA and C-ring fragmentation were the most possible fragmentation pathways for flavonoids. By accurate mass measurements within 4 ppm error for each molecular ion and subsequent fragment ions, as well as the 'full mass spectral' information of TOF-MS, a total of 41 compounds including 13 iridoid glycosides, 11 phenolic acids, 7 saponins, and 10 flavonoids were identified in a methanolic extract of FLJ. Copyright (c) 2009 John Wiley & Sons, Ltd.

Analysis of large-scale gene expression data.

PubMed

Sherlock, G

2000-04-01

The advent of cDNA and oligonucleotide microarray technologies has led to a paradigm shift in biological investigation, such that the bottleneck in research is shifting from data generation to data analysis. Hierarchical clustering, divisive clustering, self-organizing maps and k-means clustering have all been recently used to make sense of this mass of data.

Optimized Negative Staining: a High-throughput Protocol for Examining Small and Asymmetric Protein Structure by Electron Microscopy

DOE PAGES

Rames, Matthew; Yu, Yadong; Ren, Gang

2014-08-15

Structural determination of proteins is rather challenging for proteins with molecular masses between 40 - 200 kDa. Considering that more than half of natural proteins have a molecular mass between 40 - 200 kDa, a robust and high-throughput method with a nanometer resolution capability is needed. Negative staining (NS) electron microscopy (EM) is an easy, rapid, and qualitative approach which has frequently been used in research laboratories to examine protein structure and protein-protein interactions. Unfortunately, conventional NS protocols often generate structural artifacts on proteins, especially with lipoproteins that usually form presenting rouleaux artifacts. By using images of lipoproteins from cryo-electronmore » microscopy (cryo-EM) as a standard, the key parameters in NS specimen preparation conditions were recently screened and reported as the optimized NS protocol (OpNS), a modified conventional NS protocol. Artifacts like rouleaux can be greatly limited by OpNS, additionally providing high contrast along with reasonably high-resolution (near 1 nm) images of small and asymmetric proteins. These high-resolution and high contrast images are even favorable for an individual protein (a single object, no average) 3D reconstruction, such as a 160 kDa antibody, through the method of electron tomography. Moreover, OpNS can be a high-throughput tool to examine hundreds of samples of small proteins. For example, the previously published mechanism of 53 kDa cholesteryl ester transfer protein (CETP) involved the screening and imaging of hundreds of samples. Considering cryo-EM rarely successfully images proteins less than 200 kDa has yet to publish any study involving screening over one hundred sample conditions, it is fair to call OpNS a high-throughput method for studying small proteins. Hopefully the OpNS protocol presented here can be a useful tool to push the boundaries of EM and accelerate EM studies into small protein structure, dynamics and mechanisms.« less

Significant role of UV and carbonate radical on the degradation of oxytetracycline in UV-AOPs: Kinetics and mechanism.

PubMed

Liu, Yiqing; He, Xuexiang; Duan, Xiaodi; Fu, Yongsheng; Fatta-Kassinos, Despo; Dionysiou, Dionysios D

2016-05-15

Carbonate radical (CO3(•-)), a selective oxidant, reacts readily with electron-rich compounds through electron transfer and/or hydrogen abstraction. In this study, the role of CO3(•-) in degrading oxytetracycline (OTC) by UV only, UV/H2O2 and UV/persulfate (UV/PS) advanced oxidation processes (AOPs) in the presence of HCO3(-) or CO3(2-) was investigated. For UV only process, the presence of photosensitizers, i.e., nitrate (NO3(-)) and natural organic matter (NOM), had different impacts on OTC degradation, i.e., an enhancing effect by NO3(-) due to the generation of HO(•) and a slight inhibiting effect by NOM possibly due to a light scattering effect. Differently for UV/H2O2 and UV/PS processes, the presence of NO3(-) hardly influenced the destruction of OTC. Generation of CO3(•-) presented a positive role on OTC degradation by UV/NO3(-)/HCO3(-). Such influence was also observed in the two studied AOPs in the presence of both bicarbonate and other natural water constituents. When various natural water samples from different sources were used as reaction matrices, UV only and UV/H2O2 showed an inhibiting effect while UV/PS demonstrated a comparable or even promoting effect in OTC decomposition. After elucidating the potential contribution of UV direct photolysis via excited state OTC* at an elevated reaction pH condition, putative OTC transformation byproducts via CO3(•-) reaction were identified by ultra-high definition accurate-mass quadrupole time-of-flight tandem mass spectrometry (QTOF/MS). Five different reaction pathways were subsequently proposed, including hydroxylation (+16 Da), quinonization (+14 Da), demethylation (-14 Da), decarbonylation (-28 Da) and dehydration (-18 Da). The significant role of UV at high pH and CO3(•-) on OTC removal from contaminated water was therefore demonstrated both kinetically and mechanistically. Copyright © 2016 Elsevier Ltd. All rights reserved.

Mass balance of arsenic fluxes in rivers impacted by gold mining activities in Paracatu (Minas Gerais State, Brazil).

PubMed

Bidone, Edison; Cesar, Ricardo; Santos, Maria Carla; Sierpe, Ricardo; Silva-Filho, Emmanuel Vieira; Kutter, Vinicius; Dias da Silva, Lílian I; Castilhos, Zuleica

2018-03-01

Arsenic (As) is a dangerous and carcinogenic element and drinking water is its main pathway of human exposure. Gold mines are widely recognized as important sources of As pollution. This work proposes the assessment of As distribution along watersheds surrounding "Morro do Ouro" gold mine (Paracatu, southeastern Brazil). A balance approach between filtered As fluxes (As < 0.45 μm) and suspended particulate material (AsSPM) in different river segments was applied. Ultrafiltration procedure was used to categorize As into the following classes: particulate > 0.1 μm, colloidal < 0.1 μm to > 10 kDa, dissolved < 10 kDa to > 1 kDa, and truly dissolved < 1 kDa. By applying this approach, arsenic contributions from mining facilities were quantified in order to identify critical fluvial segments and support decision makers in actions of remediation. The mass balance indicated the occurrence of a decreasing gradient from upstream to downstream: (i) of the As concentrations higher than the limit established by Brazilian law (10 μg L -1 ); (ii) of the ratio between specific fluxes (g As km -2  day -1 ) and those determined using an uncontaminated watershed (a proxy for estimating the anthropic contribution), from 103 to 101; (iii) of the specific fluxes As < 0.45 μm and AsSPM from 102 to 100; and (iv) of the negative balance output minus input for each river segment that suggests As accumulation in sediments along the rivers in both urban and rural areas, mainly due to SPM sedimentation and sorption by Fe oxyhydroxides. Ultrafiltration shattering showed concentrations of decreasing As with particle size; the SPM load (> 0.1 μm) was almost one order higher to dissolved load (< 1 kDa).

Isotope shifts in francium isotopes Fr 206 - 213 and Fr 221

DOE PAGES

Collister, R.; Gwinner, G.; Tandecki, M.; ...

2014-11-07

We present the isotope shifts of the 7s 1/2 to 7p 1/2 transition for francium isotopes ²⁰⁶⁻²¹³Fr with reference to ²²¹Fr collected from two experimental periods. The shifts are measured on a sample of atoms prepared within a magneto-optical trap by a fast sweep of radio-frequency sidebands applied to a carrier laser. King plot analysis, which includes literature values for 7s 1/2 to 7p 3/2 isotope shifts, provides a field shift constant ratio of 1.0520(10) and a difference between the specific mass shift constants of 170(100) GHz amu between the D₁ and D₂ transitions, of sufficient precision to differentiate betweenmore » ab initio calculations.« less

Miniature Piezoelectric Macro-Mass Balance

NASA Technical Reports Server (NTRS)

Sherrit, Stewart; Trebi-Ollennu, Ashitey; Bonitz, Robert G.; Bar-Cohen, Yoseph

2010-01-01

Mass balances usually use a strain gauge that requires an impedance measurement and is susceptible to noise and thermal drift. A piezoelectric balance can be used to measure mass directly by monitoring the voltage developed across the piezoelectric balance, which is linear with weight or it can be used in resonance to produce a frequency change proportional to the mass change (see figure). The piezoelectric actuator/balance is swept in frequency through its fundamental resonance. If a small mass is added to the balance, the resonance frequency shifts down in proportion to the mass. By monitoring the frequency shift, the mass can be determined. This design allows for two independent measurements of mass. Additionally, more than one sample can be verified because this invention allows for each sample to be transported away from the measuring device upon completion of the measurement, if required. A piezoelectric actuator, or many piezoelectric actuators, was placed between the collection plate of the sampling system and the support structure. As the sample mass is added to the plate, the piezoelectrics are stressed, causing them to produce a voltage that is proportional to the mass and acceleration. In addition, a change in mass delta m produces a change in the resonance frequency with delta f proportional to delta m. In a microgravity environment, the spacecraft could be accelerated to produce a force on the piezoelectric actuator that would produce a voltage proportional to the mass and acceleration. Alternatively, the acceleration could be used to force the mass on the plate, and the inertial effects of the mass on the plate would produce a shift in the resonance frequency with the change in frequency related to the mass change. Three prototypes of the mass balance mechanism were developed. These macro-mass balances each consist of a solid base and an APA 60 Cedrat flextensional piezoelectric actuator supporting a measuring plate. A similar structure with 3 APA 120 Cedrat flextensional piezoelectric actuators spaced equidistantly at 120 degrees supporting the plate and a softer macro balance with an APA 150 actuator/sensor were developed. These flextensional actuators were chosen because they increase the sensitivity of the actuator to stress, allow the piezoelectric to be pre-stressed, and the piezoelectric element is a stacked multilayer actuator, which has a considerably lower input impedance than a monolithic element that allows for common instruments (e.g., input impedance of 10 megohms) to measure the voltage without rapidly discharging the charge/voltage on the piezoelectric actuator.

Differential expressed protein in developing stages of Nepenthes gracilis Korth. pitcher.

PubMed

Pinthong, Krit; Chaveerach, Arunrat; Tanee, Tawatchai; Sudmoon, Runglawan; Mokkamul, Piya

2009-03-15

Nepenthes gracilis Korth. is a member of carnivorous plants in family Nepenthaceae. The plants have beautiful and economically important pitchers. It is interesting to study the protein(s) correlated with the pitcher. Crude proteins were extracted from leaf, leaf with developing pitcher and developed pitcher of the same plant and analyzed by Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis (SDS-PAGE). Two protein bands with molecular weights of 42.7 and 38 kDa were obtained from young leaf and leaf with developing pitcher, respectively. The 42.7 kDa protein was identified as phosphoglycerate kinase (PGK) by Liquid Chromatography Mass Spectrometry (LC-MS/MS), but the 38 kDa band is an unknown protein. Both proteins were differentially expressed in each developing stage of the pitcher, thus may be powerful candidates play role in development pathway of leaf and pitcher.

Multi-element analysis of water decoction of medicine food homology plants using inductively coupled plasma-tandem mass spectrometry

NASA Astrophysics Data System (ADS)

Fu, Liang; Shi, Shu-Yun; Chen, Xiao-Qing

2017-07-01

The concentration of twelve trace elements in the water decoction of medicine food homology plants (MFHP) was determined by inductively coupled plasma-tandem mass spectrometry (ICP-MS/MS). Water decoctions of MFHP were analyzed directly using the MS/MS mode after acidification by 1% (v/v) nitric acid. The polyatomic interferences were eliminated by oxygen mass shift, oxygen on-mass, and ammonia mass shift. The accuracy of the method was verified by analysis of standard reference materials. This method was utilized to investigate the water decoction composition of 16 common Chinese MFHPs. The trace elements in the water decoctions of different MFHPs presented significantly different dissolution ratios. The dissolution ratio of V was the lowest (4.21%-14.86%), whereas Zn showed the highest dissolution ratio (24.87%-86.80%). In addition, the dissolution ratio of heavy metallic elements in most MFHP was equal to or was lower than 30%. Therefore, consumption of MHFP decoction could decrease the heavy metal intake associated with MFHP use and reduce the risk of heavy metal poisoning.

Long Work Hours and Adiposity Among Police Officers in a US Northeast City

PubMed Central

Gu, Ja K.; Charles, Luenda E.; Burchfiel, Cecil M.; Fekedulegn, Desta; Sarkisian, Khachatur; Andrew, Michael E.; Ma, Claudia; Violanti, John M.

2015-01-01

Objective To investigate the associations between long work hours and adiposity measures in police officers. Methods Participants included 408 officers from the Buffalo Cardio-Metabolic Occupational Police Stress study who were examined between 2004 and 2009. Total work hours were abstracted from payroll records and questionnaires. Analysis of variance and covariance models were used. Results Among male officers who worked the midnight shift, mean values of waist circumference and body mass index increased with longer work hours after adjustment for age, physical activity, energy intake, sleep duration, smoking status, police rank, activities after work (eg, child/family care, sports), and household income. Adiposity measures were not associated with work hours among women on any shift. Conclusion Working longer hours was significantly associated with larger waist circumferences and higher body mass index among male police officers working the midnight shift. PMID:23013913

Air-mass flux measurement system using Doppler-shifted filtered Rayleigh scattering

NASA Technical Reports Server (NTRS)

Shirley, John A.; Winter, Michael

1993-01-01

An optical system has been investigated to measure mass flux distributions in the inlet of a high speed air-breathing propulsion system. Rayleigh scattered light from air is proportional to the number density of molecules and hence can be used to ascertain the gas density in a calibrated system. Velocity field measurements are achieved by spectrally filtering the elastically-scattered Doppler-shifted light with an absorbing molecular filter. A novel anamorphic optical collection system is used which allows optical rays from different scattering angles, that have different Doppler shifts, to be recorded separately. This is shown to obviate the need to tune the laser through the absorption to determine velocities, while retaining the ability to make spatially-resolved measurements along a line. By properly selecting the laser tuning and filter parameters, simultaneous density measurements can be made. These properties are discussed in the paper and experiments demonstrating the velocimetry capability are described.