共查询到20条相似文献,搜索用时 31 毫秒
1.
T. A. Shelkovenko S. A. Pikuz I. N. Tilikin A. R. Mingaleev L. Atoyan D. A. Hammer 《Plasma Physics Reports》2018,44(2):236-244
Electric explosions of flat Al, Тi, Ni, Cu, and Та foils with thicknesses of 1?16 μm, widths of 1?8 mm, and lengths of 5?11 mm were studied experimentally on the BIN, XP, and COBRA high-current generators at currents of 40?1000 kA and current densities of (5–50) × 108 A/cm2. The images of the exploded foils were taken at different angles to the foil surface by using point projection radiography with an X-pinch hot spot as the radiation source, the spatial resolution and exposure time being 3 μm and 50 ps, respectively, as well by the laser probing method with a spatial resolution of 20 μm and an exposure time of 180 ps. In the course of foil explosion, rapidly expanding objects resembling the core and corona of an exploded wire were observed. It is shown that the core of the exploded foil has a complicated time-varying structure. 相似文献
2.
The 13Cα chemical shifts for 16,299 residues from 213 conformations of four proteins (experimentally determined by X-ray crystallography
and Nuclear Magnetic Resonance methods) were computed by using a combination of approaches that includes, but is not limited
to, the use of density functional theory. Initially, a validation test of this methodology was carried out by a detailed examination
of the correlation between computed and observed 13Cα chemical shifts of 10,564 (of the 16,299) residues from 139 conformations of the human protein ubiquitin. The results of
this validation test on ubiquitin show agreement with conclusions derived from computation of the chemical shifts at the ab initio Hartree–Fock level. Further, application of this methodology to 5,735 residues from 74 conformations of the three remaining
proteins that differ in their number of amino acid residues, sequence and three-dimensional structure, together with a new
scoring function, namely the conformationally averaged root-mean-square-deviation, enables us to: (a) offer a criterion for an accurate assessment of the quality of NMR-derived protein conformations; (b) examine whether X-ray or NMR-solved structures are better representations of the
observed 13Cα chemical shifts in solution; (c) provide evidence indicating that the proposed methodology is more accurate than automated
predictors for validation of protein structures; (d) shed light as to whether the agreement between computed and observed
13Cα chemical shifts is influenced by the identity of an amino acid residue or its location in the sequence; and (e) provide evidence
confirming the presence of dynamics for proteins in solution, and hence showing that an ensemble of conformations is a better
representation of the structure in solution than any single conformation.
Electronic Supplementary Material The online version of this article (doi: ) contains supplementary material, which is available to authorized users. 相似文献
3.
The limits of resolution that can be obtained in 1H–15N 2D NMR spectroscopy of isotopically enriched nanocrystalline proteins are explored. Combinations of frequency switched Lee–Goldburg (FSLG) decoupling, fast magic angle sample spinning (MAS), and isotopic dilution via deuteration are investigated as methods for narrowing the amide 1H resonances. Heteronuclear decoupling of 15N from the 1H resonances is also studied. Using human ubiquitin as a model system, the best resolution is most easily obtained with uniformly 2H and 15N enriched protein where the amides have been exchanged in normal water, MAS at 20 kHz, and WALTZ-16 decoupling of the 15N nuclei. The combination of these techniques results in average 1H lines of only 0.26 ppm full width at half maximum. Techniques for optimizing instrument stability and 15N decoupling are described for achieving the best possible performance in these experiments. 相似文献
4.
We present two NMR experiments, (3,2)D HNHA and (3,2)D HNHB, for rapid and accurate measurement of 3J(H N-H alpha) and 3J(N-H beta) coupling constants in polypeptides based on the principle of G-matrix Fourier transform NMR spectroscopy and quantitative J-correlation. These experiments, which facilitate fast acquisition of three-dimensional data with high spectral/digital resolution and chemical shift dispersion, will provide renewed opportunities to utilize them for sequence specific resonance assignments, estimation/characterization of secondary structure with/without prior knowledge of resonance assignments, stereospecific assignment of prochiral groups and 3D structure determination, refinement and validation. Taken together, these experiments have a wide range of applications from structural genomics projects to studying structure and folding in polypeptides. 相似文献
5.
E. Yu. Aleshina N. V. Pyndyk A. A. Moisa M. A. Sanzhakov O. N. Kharybin E. N. Nikolaev E. F. Kolesanova 《Biochemistry (Moscow) Supplemental Series B: Biomedical Chemistry》2008,2(3):288-292
The peptide RHDSGY, a fragment of the human β-amyloid Zn-binding site, and its isomers RH(D-Asp)SGY and RH(β-Asp)SGY have been obtained as amides by means of solid-phase synthesis and analyzed by HPLC and various mass spectrometric methods. The problem of low yield of the RHDSGY peptide and its isomers attributed to 9-fluorenylmethoxycarbonyl (Fmoc)-amino acids and/or formation of such side-products as RH(β-Asp)SGY (or RHDSGY during synthesis of RH(β-Asp)SGY) and RH(Asp-imide) SGY was solved via selection of individual reagents for removal of Fmoc groups from α-amino groups of the growing peptide chain. 相似文献
6.
Khandekar Jishan Bari Shrikant Sharma Kandala V. R. Chary 《Biomolecular NMR assignments》2018,12(1):51-55
γS-crystallin is a major structural component of the human eye lens, which maintains its stability over the lifetime of an organism with negligible turnover. The G57W mutant of human γS-crystallin (abbreviated hereafter as γS-G57W) is associated with dominant congenital cataracts. In order to provide a structural basis for the ability of γS-G57W causing cataract, we have cloned, overexpressed, isolated and purified the protein. The 2D [15N–1H]-HSQC spectrum recorded with uniformly 13C/15N-labelled γS-G57W was highly dispersed indicating the protein to adopt an ordered conformation. In this paper, we report almost complete sequence-specific 1H, 13C and 15N resonance assignments of γS-G57W using a suite of heteronuclear 3D NMR experiments. 相似文献
7.
NMR relaxation of arginine (Arg) 15Nε nuclei is useful for studying side-chain dynamics of proteins. In this work, we studied the impact of two geminal 15N–15N scalar couplings on measurements of transverse relaxation rates (R 2 ) for Arg side-chain 15Nε nuclei. For 12 Arg side chains of the DNA-binding domain of the Antp protein, we measured the geminal 15N–15N couplings ( 2 J NN ) of the 15Nε nuclei and found that the magnitudes of the 2 J NN coupling constants were virtually uniform with an average of 1.2 Hz. Our simulations, assuming ideal 180° rotations for all 15N nuclei, suggested that the two 2 J NN couplings of this magnitude could in principle cause significant modulation in signal intensities during the Carr–Purcell-Meiboom–Gill (CPMG) scheme for Arg 15Nε R 2 measurements. However, our experimental data show that the expected modulation via two 2 J NN couplings vanishes during the 15N CPMG scheme. This quenching of J modulation can be explained by the mechanism described in Dittmer and Bodenhausen (Chemphyschem 7:831–836, 2006). This effect allows for accurate measurements of R 2 relaxation rates for Arg side-chain 15Nε nuclei despite the presence of two 2 J NN couplings. Although the so-called recoupling conditions may cause overestimate of R 2 rates for very mobile Arg side chains, such conditions can readily be avoided through appropriate experimental settings. 相似文献
8.
V. A. Vodeneev E. K. Akinchits L. A. Orlova V. S. Sukhov 《Russian Journal of Plant Physiology》2011,58(6):974-981
The contributions of Ca2+, H+, and Cl− in generation of variation potentials (VP) in 3- to 4-week-old pumpkin (Cucurbita pepo L., cv. Mozoleevskaya) plants were assessed. During VP generation, transient alkalinization of the medium around the stem
was recorded with a potentiometric method. The pH changes were kinetically similar to the electric potential changes and were
apparently due to temporal suppression of the plasma-membrane electrogenic H+ pump. These data and the observed inhibition of VP in the stem zone treated locally with a metabolic inhibitor (NaN3) indicate that the VP generation is related to the reversible suppression of the H+-pump. The anion channel blocker (ethacrynic acid) decelerated significantly the front slope of VP and reduced the VP amplitude.
A short-term increase in external Cl− concentration around the stem was observed during potential transients representing the VP front slope and the pulses integrated
into VP. The removal of Ca2+ from extracellular medium inhibited the VP generation. It is proposed that Ca2+ plays a role in activation of anion channels and in the H+-pump inactivation. The VP generation is probably determined by a complex mechanism, with contributions from passive ion fluxes
(Ca2+, Cl−) moving along the electrochemical gradients and from changes in the electrogenic pump activity. 相似文献
9.
Microarrays are a powerful tool for comparison and understanding of gene expression levels in healthy and diseased states. The method relies upon the assumption that signals from microarray features are a reflection of relative gene expression levels of the cell types under investigation. It has previously been reported that the classical fluorescent dyes used for microarray technology, Cy3 and Cy5, are not ideal due to the decreased stability and fluorescence intensity of the Cy5 dye relative to the Cy3, such that dye bias is an accepted phenomena necessitating dye swap experimental protocols and analysis of differential dye affects. The incentive to find new fluorophores is based on alleviating the problem of dye bias through synonymous performance between counterpart dyes. Alexa Fluor 555 and Alexa Fluor 647 are increasingly promoted as replacements for CyDye in microarray experiments. Performance relates to the molecular and steric similarities, which will vary for each new pair of dyes as well as the spectral integrity for the specific application required. Comparative analysis of the performance of these two competitive dye pairs in practical microarray applications is warranted towards this end. The findings of our study showed that both dye pairs were comparable but that conventional CyDye resulted in significantly higher signal intensities (P < 0.05) and signal minus background levels (P < 0.05) with no significant difference in background values (P > 0.05). This translated to greater levels of differential gene expression with CyDye than with the Alexa Fluor counterparts. However, CyDye fluorophores and in particular Cy5, were found to be less photostable over time and following repeated scans in microarray experiments. These results suggest that precautions against potential dye affects will continue to be necessary and that no one dye pair negates this need. 相似文献
10.
Roxanne P. Smith Biswaranjan Mohanty Martin L. Williams Martin J. Scanlon Begoña Heras 《Biomolecular NMR assignments》2017,11(2):181-186
DsbD is a disulfide bond reductase present in the inner membrane of many Gamma-Proteobacteria. In the human pathogen Neisseria meningitidis, DsbD is required for viability and represents a potential target for the development of antibiotics. Here we report the chemical shift assignments (HN, N, Cα and Cβ) for the reduced and oxidized forms of the two periplasmic domains of N. meningitidis DsbD, n-NmDsbD and c-NmDsbD. The backbone amide resonances in all four forms were completely assigned, and the secondary structures for the core regions of the proteins were calculated using 13Cαβ shifts. The reduced and oxidized forms of each domain have similar secondary shifts suggesting they retain the same fold. We anticipate that these data will provide an important basis for studying the interaction between n-NmDsbD and c-NmDsbD, which is required for electron transfer across the bacterial cytoplasmic membrane. 相似文献
11.
John E. Linley Stefan H. Boese Nicholas L. Simmons Michael A. Gray 《The Journal of membrane biology》2009,230(2):57-68
We have previously shown that the membrane conductance of mIMCD-3 cells at a holding potential of 0 mV is dominated by a Ca2+-dependent Cl− current (ICLCA). Here we report that ICLCA activity is also voltage dependent and that this dependence on voltage is linked to the opening of a novel Al3+-sensitive, voltage-dependent, Ca2+ influx pathway. Using whole-cell patch-clamp recordings at a physiological holding potential (−60 mV), ICLCA was found to be inactive and resting currents were predominantly K+ selective. However, membrane depolarization to 0 mV resulted in a slow, sigmoidal, activation of ICLCA (T
0.5 ~ 500 s), while repolarization in turn resulted in a monoexponential decay in ICLCA (T
0.5 ~ 100 s). The activation of ICLCA by depolarization was reduced by lowering extracellular Ca2+ and completely inhibited by buffering cytosolic Ca2+ with EGTA, suggesting a role for Ca2+ influx in the activation of ICLCA. However, raising bulk cytosolic Ca2+ at −60 mV did not produce sustained ICLCA activity. Therefore ICLCA is dependent on both an increase in intracellular Ca2+ and depolarization to be active. We further show that membrane depolarization is coupled to opening of a Ca2+ influx pathway that displays equal permeability to Ca2+ and Ba2+ ions and that is blocked by extracellular Al3+ and La3+. Furthermore, Al3+ completely and reversibly inhibited depolarization-induced activation of ICLCA, thereby directly linking Ca2+ influx to activation of ICLCA. We speculate that during sustained membrane depolarization, calcium influx activates ICLCA which functions to modulate NaCl transport across the apical membrane of IMCD cells. 相似文献
12.
Luke W. Arbogast Frank Delaglio Joel R. Tolman John P. Marino 《Journal of biomolecular NMR》2018,72(3-4):149-161
While the use of 1H–13C methyl correlated NMR spectroscopy at natural isotopic abundance has been demonstrated as feasible on protein therapeutics as large as monoclonal antibodies, spectral interference from aliphatic excipients remains a significant obstacle to its widespread application. These signals can cause large baseline artifacts, obscure protein resonances, and cause dynamic range suppression of weak peaks in non-uniform sampling applications, thus hampering both traditional peak-based spectral analyses as well as emerging chemometric methods of analysis. Here we detail modifications to the 2D 1H–13C gradient-selected HSQC experiment that make use of selective pulsing techniques for targeted removal of interfering excipient signals in spectra of the NISTmAb prepared in several different formulations. This approach is demonstrated to selectively reduce interfering excipient signals while still yielding 2D spectra with only modest losses in protein signal. Furthermore, it is shown that spectral modeling based on the SMILE algorithm can be used to simulate and subtract any residual excipient signals and their attendant artifacts from the resulting 2D NMR spectra. 相似文献
13.
Imino 1H–15N residual dipolar couplings (RDCs) provide additional structural information that complements standard 1H–1H NOEs leading to improvements in both the local and global structure of RNAs. Here, we report measurement of imino 1H–1H RDCs for the Iron Responsive Element (IRE) RNA and native E. coli tRNAVal using a BEST-Jcomp-HMQC2 experiment. 1H–1H RDCs are observed between the imino protons in G–U wobble base pairs and between imino protons on neighboring base pairs
in both RNAs. These imino 1H–1H RDCs complement standard 1H–15N RDCs because the 1H–1H vectors generally point along the helical axis, roughly perpendicular to 1H–15N RDCs. The use of longitudinal relaxation enhancement increased the signal-to-noise of the spectra by ~3.5-fold over the
standard experiment. The ability to measure imino 1H–1H RDCs offers a new restraint, which can be used in NMR domain orientation and structural studies of RNAs.
Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. 相似文献
14.
Murakami D Suzuki MF da Silva Dias M Okazaki K 《Radiation and environmental biophysics》2004,43(2):91-99
Among various types of ionizing radiation, the beta emitter radionuclides are involved in many sectors of human activity, such as nuclear medicine, nuclear industries and biomedicine, with a consequently increased risk of accidental, occupational or therapeutic exposure. Despite their recognized importance, there is little information about the effect of beta particles at the cellular level when compared to other types of ionizing radiation. Thus, the objective of the present study was to evaluate the genotoxic and cytotoxic effects of 90Sr/90Y—a pure, highly energetic beta source—on Chinese hamster ovary (CHO) cells and to compare them with data obtained with 60Co. CHO cells irradiated with different doses of 60Co (0.34 Gy min–1) and 90Sr/90Y (0.23 Gy min–1) were processed for analysis of clonogenic death, induction of micronuclei (MN) and interphase death. The survival curves obtained for both types of radiation were fitted by the exponential quadratic model and were found to be similar. Also, the cytogenetic results showed similar frequencies of radio-induced MN between gamma and beta radiations and the MN distribution pattern among cells did not follow the expected Poisson probability pattern. The relative variance values were significantly higher in cells irradiated with 90Sr/90Y than with 60Co in all exposure doses. The irradiated cells showed more necrotic cells 72 h and 96 h after exposure to beta than to gamma radiation. In general, the 90Sr/90Y -radiation was more damaging than 60Co -rays. The data obtained also demonstrated the need to use several parameters for a better estimate of cellular sensitivity to the action of genotoxic agents, which would be important in terms of radiobiology, oncology and therapeutics. 相似文献
15.
V. A. Belyaev D. A. Kozlov A. A. Terent’ev A. E. Trenin 《Plasma Physics Reports》2017,43(10):1039-1041
A method for determining the lifetime of unstable ions is described. The method is based on measuring the decrease in the ion beam current onto a fixed detector with increasing path length of the ion beam from the ion source to the detector. The measurements performed for D? 2 and HD? molecular ions have shown that their lifetimes are 3.5 ± 0.1 and 4.4 ± 0.1 μs, respectively. 相似文献
16.
A confined aquifer in the Malm Karst of the Franconian Alb, South Germany was investigated in order to understand the role of the vadose zone in denitrifiaction processes. The concentrations of chemical tracers Sr2+ and Cl– and concentrations of stable isotope 18O were measured in spring water and precipitation during storm events. Based on these measurements a conceptual model for runoff was constructed. The results indicate that pre-event water, already stored in the system at the beginning of the event, flows downslope on vertical and lateral preferential flow paths. Chemical tracers used in a mixing model for hydrograph separation have shown that the pre-event water contribution is up to 30%. Applying this information to a conceptual runoff generation model, the values of 15N and 18O in nitrate could be calculated. Field observations showed the occurence of significant microbial denitrification processes above the soil/bedrock interface before nitrate percolates through to the deeper horizon of the vadose zone. The source of nitrate could be determined and denitrification processes were calculated. Assuming that the nitrate reduction follows a Rayleigh process one could approximate a nitrate input concentration of about 170 mg/l and a residual nitrate concentration of only about 15%. The results of the chemical and isotopic tracers postulate fertilizers as nitrate source with some influence of atmospheric nitrate. The combined application of hydrograph separation and determination of isotope values in 15N and 18O of nitrate lead to an improved understanding of microbial processes (nitrification, denitrification) in dynamic systems. 相似文献
17.
Seasonal oscillations in the carbon (δ13C) and nitrogen (δ15N) isotope signatures of aquatic algae can cause seasonal enrichment–depletion cycles in the isotopic composition of planktonic
invertebrates (e.g., copepods). Yet, there is growing evidence that seasonal enrichment–depletion cycles also occur in the
isotope signatures of larger invertebrate consumers, taxa used to define reference points in isotope-based trophic models
(e.g., trophic baselines). To evaluate the general assumption of temporal stability in non-zooplankton aquatic invertebrates,
δ13C and δ15N time series data from the literature were analyzed for seasonality and the influence of biotic (feeding group) and abiotic
(trophic state, climate regime) factors on isotope temporal patterns. The amplitude of δ13C and δ15N enrichment–depletion cycles was negatively related to body size, although all size-classes of invertebrates displayed a
winter-to-summer enrichment in δ13C and depletion in δ15N. Among feeding groups, periphytic grazers were more variable and displayed larger temporal changes in δ13C than detritivores. For nitrogen, temporal variability and magnitude of directional change of δ15N was most strongly related to ecosystem trophic state (eutrophic > mesotrophic, oligotrophic). This study provides evidence
of seasonality in the isotopic composition of aquatic invertebrates across very broad geographical and ecological gradients
as well as identifying factors that are likely to modulate the strength and variability of seasonality. These results emphasize
the need for researchers to recognize the likelihood of temporal changes in non-zooplankton aquatic invertebrate consumers
at time scales relevant to seasonal studies and, if present, to account for temporal dynamics in isotope trophic models. 相似文献
18.
Jaroniec CP Boisbouvier J Tworowska I Nikonowicz EP Bax A 《Journal of biomolecular NMR》2005,31(3):231-241
New 3D HCN quantitative J (QJ) pulse schemes are presented for the precise and accurate measurement of one-bond 15N1/9–13C1, 15N1/9–13C6/8, and 15N1/9–13C2/4 residual dipolar couplings (RDCs) in weakly aligned nucleic acids. The methods employ 1H–13C multiple quantum (MQ) coherence or TROSY-type pulse sequences for optimal resolution and sensitivity. RDCs are obtained from the intensity ratio of H1–C1–N1/9 (MQ-HCN-QJ) or H6/8–C6/8–N1/9 (TROSY-HCN-QJ) correlations in two interleaved 3D NMR spectra, with dephasing intervals of zero (reference spectrum) and 1/(2JNC) (attenuated spectrum). The different types of 15N–13C couplings can be obtained by using either the 3D MQ-HCN-QJ or TROSY-HCN-QJ pulse scheme, with the appropriate setting of the duration of the constant-time 15N evolution period and the offset of two frequency-selective 13C pulses. The methods are demonstrated for a uniformly 13C, 15N-enriched 24-nucleotide stem-loop RNA sequence, helix-35, aligned in the magnetic field using phage Pf1. For measurements of RDCs systematic errors are found to be negligible, and experiments performed on a 1.5 mM helix-35 sample result in an estimated precision of ca. 0.07 Hz for 1DNC, indicating the utility of the measured RDCs in structure validation and refinement. Indeed, for a complete set of 15N1/9–13C1, 15N1/9–13C6/8, and 15N1/9–13C2/4 dipolar couplings obtained for the stem nucleotides, the measured RDCs are in excellent agreement with those predicted for an NMR structure of helix-35, refined using independently measured observables, including 13C–1H, 13C–13C and 1H–1H dipolar couplings.Supplementary material to this paper is available in electronic form at
http://dx.doi.org/10.1007/s10858-005-0646-2. 相似文献
19.
Shucha Zhang Lingyan Liu Debora Steffen Tao Ye Daniel Raftery 《Metabolomics : Official journal of the Metabolomic Society》2012,8(2):323-334
This study aims to investigate the metabolic difference between male and female healthy adults using a combination of GC–MS
and NMR metabolomics techniques. While metabolomics has shown wide applications in characterizing the status and progression
of many diseases, physiological factors such as gender often contribute high levels of variability that can hinder the detection
of biomarkers of interest, such as in disease detection. We carried out a detailed exploration of gender related metabolic
profiling of human urine using a Headspace-SPME/GC–MS approach and detected over two hundred peaks. Fifty-nine metabolites
were identified using the NIST library. 1H NMR spectroscopy was also utilized, and resulted in the identification of eighteen metabolites. We find that both GC–MS
and NMR are able to capture human gender metabolic differences, and their combination allows a significantly better understanding
of this difference. Subtle differences between genders are found to be related to the metabolism of fats, amino acids, and
TCA cycle intermediates. 相似文献
20.