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Bukharina N.S.,RAS Institute of Chemistry | Pleshakova T.O.,RAS Institute of Chemistry | Ziborov V.S.,RAS Joint Institute for High Temperatures | Fokin D.A.,Bruker | And 2 more authors.
Science of Advanced Materials | Year: 2017

Deformation of single cytochrome CYP102A1 (P450 BM3) molecules and its influence on protein fluctuation during the catalytic cycle were explored by atomic force microscopy (AFM). A variety of AFM modes allowed to study CYP102A1 elasticity. Measurements were carried out in three AFM modes: force spectroscopy (FS mode), PeakForce QNM and tapping modes. AFM measurements in a tapping mode were conducted using standard probes (the resonance frequency of about 9 kHz in liquid) and short high-speed probes (the resonance frequency of about 850 kHz in liquid). It was shown that the relationship of a loading rate and protein deformation was nonlinear. CYP102A1 Young's modulus of 1.3-35 MPa was estimated from AFM force- distance curves. CYP102A1 deformation in a tapping mode with high-speed probes was greater compared to that in a tapping mode with standard probes. Increasing of deformation was associated with a disappearance of CYP102A1 fluctuations during AFM monitoring of enzyme catalytic cycle. © 2017 by American Scientific Publishers.


Speranskaya A.S.,RAS Engelhardt Institute of Molecular Biology | Krinitsina A.A.,Moscow State University | Kudryavtseva A.V.,RAS Engelhardt Institute of Molecular Biology | Poltronieri P.,CNR Institute of Sciences of Food Production | And 6 more authors.
Biochimie | Year: 2012

Background: The group of Kunitz-type protease inhibitors (KPI) from potato is encoded by a polymorphic family of multiple allelic and non-allelic genes. The previous explanations of the KPI variability were based on the hypothesis of random mutagenesis as a key factor of KPI polymorphism. Results: KPI-A genes from the genomes of Solanum tuberosum cv. Istrinskii and the wild species Solanum palustre were amplified by PCR with subsequent cloning in plasmids. True KPI sequences were derived from comparison of the cloned copies. "Hot spots" of recombination in KPI genes were independently identified by DnaSP 4.0 and TOPALi v2.5 software. The KPI-A sequence from potato cv. Istrinskii was found to be 100% identical to the gene from Solanum nigrum. This fact illustrates a high degree of similarity of KPI genes in the genus Solanum. Pairwise comparison of KPI A and B genes unambiguously showed a non-uniform extent of polymorphism at different nt positions. Moreover, the occurrence of substitutions was not random along the strand. Taken together, these facts contradict the traditional hypothesis of random mutagenesis as a principal source of KPI gene polymorphism. The experimentally found mosaic structure of KPI genes in both plants studied is consistent with the hypothesis suggesting recombination of ancestral genes. The same mechanism was proposed earlier for other resistance-conferring genes in the nightshade family (Solanaceae). Based on the data obtained, we searched for potential motifs of site-specific binding with plant DNA recombinases. During this work, we analyzed the sequencing data reported by the Potato Genome Sequencing Consortium (PGSC), 2011 and found considerable inconsistence of their data concerning the number, location, and orientation of KPI genes of groups A and B. Conclusions: The key role of recombination rather than random point mutagenesis in KPI polymorphism was demonstrated for the first time. © 2012 Elsevier Masson SAS. All rights reserved.


Ivanov Y.D.,RAS Institute of Chemistry | Kozlov A.F.,RAS Institute of Chemistry | Malsagova K.A.,RAS Institute of Chemistry | Pleshakova T.O.,RAS Institute of Chemistry | And 4 more authors.
Biochemistry and Biophysics Reports | Year: 2016

Monitoring of microwave emission from aqueous solution of horseradish peroxidase (HRP) in the process of the enzyme functioning was carried out. For the monitoring, a system containing HRP, luminol and n2O2 was employed. Microwave emission measurements were carried out in the 3.4-4.2 GHz frequency range using the active and passive modes (active-mode and passive-mode measurements). In the active mode, excitation of the solution in the pulsed electromagnetic field was accomplished. In the passive mode, no excitation was induced. It appears that the passive-mode measurements taken in the course of the peroxidase reaction in the enzyme system have shown a 0.5 °s increase of the microwave signal. Upon the active-mode measurements, taken in the same reaction conditions, the forced excitation of the solution has also led to the increase (by 2 °s) of the level of the microwave signal - i.e. to its 4-fold enhancement compared to the signal obtained in passive-mode measurements. © 2016 The Authors.


Sinyov V.V.,Russian Cardiology Research and Production Complex | Chicheva M.M.,Research Institute of General Pathology and Pathophysiology | Barinova V.A.,Russian Cardiology Research and Production Complex | Ryzhkova A.I.,Skryabin Moscow State Academy of Veterinary Medicine and Biotechnology | And 6 more authors.
Russian Journal of Genetics | Year: 2016

Atherosclerosis is a polygenic socially significant disease whose risk factors include coronary heart disease, diabetes, hypertension, and myocardial infarction. According to the literature, mutations m.14846G>A (G34S), m.15762G>A (G339Q), m.15084G>A (W113Ter), and m.15059G>A (G190Ter) of cytochrome B gene (MT-CYB) are associated with mitochondrial myopathies, myoglobinuria, and exercise intolerance. Preliminary studies carried out by the authors made it possible to discover an association of certain mitochondrial genome mutations with atherosclerotic lesions of aortic intima in people who died as a result of an accident or sudden death. The most interesting seemed to be the data on the association of mutations m.14846G>A and m.15059G>A of the cytochrome B gene with lipofibrous aortic plaques, because these mutations affect the mitochondrial respiratory chain enzyme. Defects in the given chain may be the reason for the launch of pathogenic mechanisms in the human body. Owing to the fact that mutations in the mitochondrial genome are inherited by the maternal type, it was decided to analyze cytochrome B gene mutations in a sample of female volunteers from Moscow oblast. According to the findings, mutations m.14846G>A and m.15059G>A are highly significantly associated with atherosclerotic lesions of the carotid arteries: m.14846G>A is antiatherogenic and m.15059G>A is proatherogenic. © 2016, Pleiades Publishing, Inc.


Bukharina N.S.,RAS Institute of Chemistry | Ivanov Y.D.,RAS Institute of Chemistry | Pleshakova T.O.,RAS Institute of Chemistry | Frantsuzov P.A.,RAS Institute of Chemistry | And 4 more authors.
Biophysics (Russian Federation) | Year: 2015

Change in temperature is one of the factors affecting the activity of enzymes. In this work, the thermal denaturation and aggregation of cytochrome P450 BM3 were studied by atomic force microscopy. The specific temperature transitions were studied by fluorescence analysis. In the low melting temperature range (10–33°C), a decrease in the fluorescence intensity of the aromatic residues was observed simultaneously with an increase in the fluorescence intensity of the flavin groups. The protein melting in this range indicated three narrow S-shaped cooperative transitions at 16, 22, and 29°C. Atomic force microscopy analysis in this temperature range showed that the BM3 molecules retained a globular shape as compact objects (heights, h < 7 nm; lateral dimensions, d < 50 nm), but the protein oligomeric state changed. The first two transitions were accompanied by a decrease in the degree of oligomerization and the third one by its increase. © 2015, Pleiades Publishing, Inc.


Guseva M.A.,Skryabin Moscow State Academy of Veterinary Medicine and Biotechnology | Epova E.Y.,Skryabin Moscow State Academy of Veterinary Medicine and Biotechnology | Kovalev L.I.,RAS A.N. Bach Institute of Biochemistry | Shevelev A.B.,RAS A.N. Bach Institute of Biochemistry
Applied Biochemistry and Microbiology | Year: 2010

Using proteomic technologies-two-dimensional electrophoresis in denaturing conditions in combination with mass spectroscopy of MALDI-TOF proteins-we demonstrated, for the first time, that the most noticeable alteration of protein composition of a Yarrowia lipolytica cell during adaptation to alkaline conditions was an increase of mitochondrial proteins relatively to proteins of cytoplasm. © Pleiades Publishing, Inc., 2010.


Zaitseva V.V.,Ukrainian Academy of Sciences | Zaitseva V.V.,Skryabin Moscow State Academy of Veterinary Medicine and Biotechnology | Tyurina T.G.,Ukrainian Academy of Sciences | Zaitsev S.Y.,Skryabin Moscow State Academy of Veterinary Medicine and Biotechnology
Russian Journal of Organic Chemistry | Year: 2015

Schemes of complex formation and decomposition in monomers copolymerization in the absence of initiator (313 K) are suggested. The energy of formation of π-H- and H-complexes of N-vinylpyrrolidone with maleic anhydride, methyl methacrylate, or two latter with each other is 9.9-18.0 kJ mol-1, the equilibrium constant is 0.58-0.19 L mol-1 (CCl4). In the case of N-vinylpyrrolidone complexes with maleic anhydride we found the proceeding of a parallel radical process of formation of dimers, oligomers, and copolymers till ~10 wt % conversion. The copolymerization rate of these monomers is higher than the rate of their homopolymerization. © 2015 Pleiades Publishing, Ltd.


Starodubova E.S.,RAS Engelhardt Institute of Molecular Biology | Preobrazhenskaia O.V.,RAS Engelhardt Institute of Molecular Biology | Kuzmenko Y.V.,RAS Engelhardt Institute of Molecular Biology | Latanova A.A.,RAS Engelhardt Institute of Molecular Biology | And 2 more authors.
Molecular Biology | Year: 2015

Rabies is an infectious disease among humans and animals that remains incurable, despite its longstanding research history. The only way to prevent the disease is prompt treatment, including vaccination as an obligatory component and administration of antirabies immunoglobulin as a supplement. Since the first antirabies vaccination performed in the 19th century, a large number of different rabies vaccines have been developed. Progress in molecular biology and biotechnology enabled the development of effective and safe technologies of vaccine production. Currently, new-generation vaccines are being developed based on recombinant rabies virus strains or on the production of an individual recombinant rabies antigen—glycoprotein (G protein), either as a component of nonpathogenic viruses, or in plants, or in the form of DNA vaccines. In this review, the main modern trends in the development of rabies vaccines have been discussed. © 2015, Pleiades Publishing, Inc.


PubMed | Moscow State University and Skryabin Moscow State Academy of Veterinary Medicine and Biotechnology
Type: Journal Article | Journal: Biomeditsinskaia khimiia | Year: 2016

Perioperative corneal abrasion is an ophthalmic complication commonly found in patients underwent general anesthesia. In this study, correlations between development of corneal injury and proteomic changes in tear film during general anesthesia were examined using an animal (rabbit) model. Being started after 1-h anesthesia, the process of accumulation of pathological changes in the cornea unequivocally led clinically significant abrasions following 3-6 h of the narcosis. The corneal damage was associated with alterations in profiles of major proteins of the tear film. Analysis of the tear proteome pointed to depression of lachrymal glands function, and suggested serotransferrin, serum albumin and annexin A1 as potential tear markers of the complication. The tear film alterations included fast drop of total antioxidant activity and activity of superoxide dismutase, and decrease in interleukin-4 and increase in interleukin-6 content indicating development of oxidative and pro-inflammatory responses. These findings suggest antioxidant and anti-inflammatory therapy as prospective approach for prevention/treatment of perioperative corneal abrasions. The observed anesthesia-induced effects should be considered in any study of ocular surface diseases employing anesthetized animals.


PubMed | RAS Engelhardt Institute of Molecular Biology and Skryabin Moscow State Academy of Veterinary Medicine and Biotechnology
Type: Journal Article | Journal: Molekuliarnaia biologiia | Year: 2015

Rabies is an infectious disease among humans and animals that remains incurable, despite its longstanding research history. The only way to prevent the disease is prompt treatment, including vaccination as an obligatory component and administration of antirabies immunoglobulin as a supplement. Since the first antirabies vaccination performed in the 19th century, a large number of different rabies vaccines have been developed. Progress in molecular biology and biotechnology enabled the development of effective and safe technologies of vaccine production. Currently, new-generation vaccines are being developed based on recombinant rabies virus strains or on the production of an individual recombinant rabies antigen-glycoprotein (G protein), either as a component of nonpathogenic viruses, or in plants, or in the form of DNA vaccines. In this review, the main modern trends in the development of rabies vaccines have been discussed.

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