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31.
Local polarity and hydrogen bonding inside the Sec14p phospholipid-binding cavity: high-field multi-frequency electron paramagnetic resonance studies 下载免费PDF全文
Smirnova TI Chadwick TG Voinov MA Poluektov O van Tol J Ozarowski A Schaaf G Ryan MM Bankaitis VA 《Biophysical journal》2007,92(10):3686-3695
Sec14p promotes the energy-independent transfer of either phosphatidylinositol (PtdIns) or phosphatidylcholine (PtdCho) between lipid bilayers in vitro and represents the major PtdIns/PtdCho transfer protein in the budding yeast Saccharomyces cerevisiae. Herein, we employ multi-frequency high-field electron paramagnetic resonance (EPR) to analyze the electrostatic and hydrogen-bonding microenvironments for series of doxyl-labeled PtdCho molecules bound by Sec14p in a soluble protein-PtdCho complex. A structurally similar compound, 5-doxyl stearic acid dissolved in a series of solvents, was used for experimental calibration. The experiments yielded two-component rigid limit 130- and 220-GHz EPR spectra with excellent resolution in the gx region. Those components were assigned to hydrogen-bonded and nonhydrogen-bonded nitroxide species. Partially resolved 130-GHz EPR spectra from n-doxyl-PtdCho bound to Sec14p were analyzed using this two-component model and allowed quantification of two parameters. First, the fraction of hydrogen-bonded nitroxide species for each n-doxyl-PtdCho was calculated. Second, the proticity profile along the phospholipid-binding cavity of Sec14p was characterized. The data suggest the polarity gradient inside the Sec14p cavity is a significant contributor to the driving molecular forces for extracting a phospholipid from the bilayer. Finally, the enhanced g-factor resolution of EPR at 130 and 220 GHz provides researchers with a spectroscopic tool to deconvolute two major contributions to the x-component of the nitroxide g-matrix: hydrogen-bond formation and local electrostatic effects. 相似文献
32.
To study virus propagation, we have developed a method by which the propagation of the Lambda bacteriophage can be observed and quantified. This is done by creating a fusion protein of the capsid protein gpD and the enhanced yellow fluorescent protein (EYFP). We show that this fusion allows capsid formation and that the modified viruses propagate on a surface covered with host bacteria thus forming fluorescent plaques. The intensity of fluorescence in a growing plaque determines the distribution of phages. This provides a new tool to study the propagation of infection at the microscopic level. 相似文献
33.
Calculation of the free energy of protein folding and delineation of its pre-organization are of foremost importance for understanding, predicting and designing biological macromolecules. Here, we introduce an energy smoothing variant of parallel tempering replica exchange Monte Carlo (REMS) that allows for efficient configurational sampling of flexible solutes under the conditions of molecular hydration. Its usage to calculate the thermal stability of a model globular protein, Trp cage TC5b, achieves excellent agreement with experimental measurements. We find that the stability of TC5b is attained through the coupled formation of local and non-local interactions. Remarkably, many of these structures persist at high temperature, concomitant with the origin of native-like configurations and mesostates in an otherwise macroscopically disordered unfolded state. Graph manifold learning reveals that the conversion of these mesostates to the native state is structurally heterogeneous, and that the cooperativity of their formation is encoded largely by the unfolded state ensemble. In all, these studies establish the extent of thermodynamic and structural pre-organization of folding of this model globular protein, and achieve the calculation of macromolecular stability ab initio, as required for ab initio structure prediction, genome annotation, and drug design. 相似文献
34.
Early loss of Xist RNA expression and inactive X chromosome associated chromatin modification in developing primordial germ cells 总被引:1,自引:0,他引:1
BACKGROUND: The inactive X chromosome characteristic of female somatic lineages is reactivated during development of the female germ cell lineage. In mouse, analysis of protein products of X-linked genes and/or transgenes located on the X chromosome has indicated that reactivation occurs after primordial germ cells reach the genital ridges. PRINCIPAL FINDINGS/METHODOLOGY: We present evidence that the epigenetic reprogramming of the inactive X-chromosome is initiated earlier than was previously thought, around the time that primordial germ cells (PGCs) migrate through the hindgut. Specifically, we find that Xist RNA expression, the primary signal for establishment of chromosome silencing, is extinguished in migrating PGCs. This is accompanied by displacement of Polycomb-group repressor proteins Eed and Suz(12), and loss of the inactive X associated histone modification, methylation of histone H3 lysine 27. CONCLUSIONS/SIGNIFICANCE: We conclude that X reactivation in primordial germ cells occurs progressively, initiated by extinction of Xist RNA around the time that germ cells migrate through the hindgut to the genital ridges. The events that we observe are reminiscent of X reactivation of the paternal X chromosome in inner cell mass cells of mouse pre-implantation embryos and suggest a unified model in which execution of the pluripotency program represses Xist RNA thereby triggering progressive reversal of epigenetic silencing of the X chromosome. 相似文献
35.
Voznesenskaya T Makogon N Bryzgina T Sukhina V Grushka N Alexeyeva I 《Reproductive biology》2007,7(3):207-220
Experimental immune ovarian failure induced in CBA mice by either administration of xenogenic anti-ovarian antibodies or immunization with allogenic ovarian extracts impaired the meiotic maturation of oocytes and increased apoptosis of follicular cells. Immunization was accompanied with the inflammation and active immune reaction, as shown by the enlargement of regional lymph nodes, the increase of apoptosis in cultured lymph node cells and the increase of band and segmented neutrophil percentage in the blood. Triple injections of melatonin (5 mg/kg of the body weight) an hour before antibodies administration restored the meiotic maturation of oocytes and supported the survival of follicular and lymph node cells. In contrast, melatonin application upon immunization was not effective to prevent the ovary impairment and cell death. It is concluded that melatonin protects against immune ovary failure induced by xenogenic anti-ovarian antibodies. 相似文献
36.
Akunevich Anastasia Aleksandrovna Khrustalev Vladislav Victorovich Khrustaleva Tatyana Aleksandrovna Poboinev Victor Vitoldovich Shalygo Nikolai Vladimirovich Stojarov Aleksander Nicolaevich Arutyunyan Alexander Migranovich Kordyukova Larisa Valentinovna Sapon Yehor Gennadyevich 《The protein journal》2022,41(2):245-259
The Protein Journal - An interplay between monomeric and dimeric forms of human epidermal growth factor (EGF) affecting its interaction with EGF receptor (EGFR) is poorly understood. While EGF... 相似文献
37.
Soo Hyun Im Tatyana A. Klochkova Da Jeoung Lee Claire M. M. Gachon Gwang Hoon Kim 《Journal of phycology》2019,55(4):801-815
Disease outbreaks devastate Pyropia aquaculture farms every year. The three most common and serious diseases are Olpidiopsis‐blight and red‐rot disease caused by oomycete pathogens and green‐spot disease caused by the PyroV1 virus. We hypothesized that a basic genetic profile of molecular defenses will be revealed by comparing and analyzing the genetic response of Pyropia tenera against the above three pathogens. RNAs isolated from infected thalli were hybridized onto an oligochip containing 15,115 primers designed from P. tenera expressed sequence tags (EST)s. Microarray profiles of the three diseases were compared and interpreted together with histochemical observation. Massive amounts of reactive oxygen species accumulated in P. tenera cells exposed to oomycete pathogens. Heat shock genes and serine proteases were the most highly up‐regulated genes in all infection experiments. Genes involved in RNA metabolism, ribosomal proteins and antioxidant metabolism were also highly up‐regulated. Genetic profiles of P. tenera in response to pathogens were most similar between the two biotrophic pathogens, Olpidiopsis pyropiae and PyroV1 virus. A group of plant resistance genes were specifically regulated against each pathogen. Our results suggested that disease response in P. tenera consists of a general constitutive defense and a genetic toolkit against specific pathogens. 相似文献
38.
Taras Antal Ivan Konyukhov Alena Volgusheva Tatyana Plyusnina Sergei Khruschev Galina Kukarskikh Sergey Goryachev Andrey Rubin 《Physiologia plantarum》2019,165(3):476-486
The development of high‐performance photobioreactors equipped with automatic systems for non‐invasive real‐time monitoring of cultivation conditions and photosynthetic parameters is a challenge in algae biotechnology. Therefore, we developed a chlorophyll (Chl) fluorescence measuring system for the online recording of the light‐induced fluorescence rise and the dark relaxation of the flash‐induced fluorescence yield (Qa? ? re‐oxidation kinetics) in photobioreactors. This system provides automatic measurements in a broad range of Chl concentrations at high frequency of gas‐tight sampling, and advanced data analysis. The performance of this new technique was tested on the green microalgae Chlamydomonas reinhardtii subjected to a sulfur deficiency stress and to long‐term dark anaerobic conditions. More than thousand fluorescence kinetic curves were recorded and analyzed during aerobic and anaerobic stages of incubation. Lifetime and amplitude values of kinetic components were determined, and their dynamics plotted on heatmaps. Out of these data, stress‐sensitive kinetic parameters were specified. This implemented apparatus can therefore be useful for the continuous real‐time monitoring of algal photosynthesis in photobioreactors. 相似文献
39.
Natalia Mokshina Olga Makshakova Alsu Nazipova Oleg Gorshkov Tatyana Gorshkova 《Physiologia plantarum》2019,167(2):173-187
Rhamnogalacturonan lyases (RGLs; EC 4.2.2.23) degrade the rhamnogalacturonan I (RG‐I) backbone of pectins present in the plant cell wall. These enzymes belong to polysaccharide lyase family 4, members of which are mainly from plants and plant pathogens. RGLs are investigated, as a rule, as pathogen ‘weapons’ for plant cell wall degradation and subsequent infection. Despite the presence of genes annotated as RGLs in plant genomes and the presence of substrates for enzyme activity in plant cells, evidence supporting the involvement of this enzyme in certain processes is limited. The differential expression of some RGL genes in flax (Linum usitatissimum L.) tissues, revealed in our previous work, prompted us to carry out a total revision (phylogenetic analysis, analysis of expression and protein structure modeling) of all the sequences of flax predicted as coding for RGLs. Comparison of the expressions of LusRGL in various tissues of flax stem revealed that LusRGLs belong to distinct phylogenetic clades, which correspond to two co‐expression groups. One of these groups comprised LusRGL6‐A and LusRGL6‐B genes and was specifically upregulated in flax fibers during deposition of the tertiary cell wall, which has complex RG‐I as a key noncellulosic component. The results of homology modeling and docking demonstrated that the topology of the LusRGL6‐A catalytic site allowed binding to the RG‐I ligand. These findings lead us to suggest the presence of RGL activity in planta and the involvement of special isoforms of RGLs in the modification of RG‐I of the tertiary cell wall in plant fibers. 相似文献
40.
Alexander S. Kashin Tatyana A. Kritskaya Alena S. Parkhomenko Ivan A. Schanzer 《Nordic Journal of Botany》2019,37(11)
We studied genetic diversity in 54 populations of nine sexual and apomictic species of the genus Chondrilla (C. acantholepis, C. ambigua, C. brevirostris, C. canescens, C. graminea, C. juncea, C. laticoronata, C. latifolia and C. pauciflora) in SE European Russia and neighboring territories of NW Kazakhstan. We analysed the trnT–trnF region of plastid DNA and the internal transcribed spacer of ribosomal DNA (ITS1–5.8S–ITS2) using statistical parsimony, maximum likelihood and neighbor net methods. Two major evolutionary lineages, roughly corresponding to the two subgenera traditionally recognized in the region, were revealed. Within the first evolutionary lineage (subgenus Brachyrhynchus), the sexual diploid C. ambigua and its putatively hybrid apomictic derivatives C. brevirostris, C. laticoronata and C. pauciflora could be recognized. Their identity was also confirmed by analyses of ISSR markers. The second evolutionary lineage (subgenus Chondrilla) comprises C. juncea, C. acantholepis, C. canescens, C. graminea and C. latifolia in European Russia, but analyses of morphological variability and the genealogy of plastid and nuclear markers favor their treatment as the single facultatively apomictic species C. juncea. The results demonstrate that an apomictic mode of reproduction does not necessarily result in the formation of genetically separated microspecies. 相似文献