Once you have missed the first button …, you'll never manage to button up
Johann Wolfgang von Goethe
Formate oxidation is a final step of methanol oxidation in methylotrophic prokaryotes and is ...important for detoxification of formate in other organisms. The structural mechanism of the formate dehydrogenase (FDH) of Pseudomonas sp. 101 has been studied for about 30 years. In the active center of FDH, the oxidation of formic acid into carbon dioxide in a NAD+-dependent way takes place. Residues that form the active center of that enzyme, as well as those that form the so-called substrate channel, are engaged in the catalytic cycle. Our study allowed to characterize a new residue, Tyr102, involved in the work of the enzyme. This residue is located in the outer neck of the substrate channel (at the beginning of the path of the substrate to the active center) and acts as a “button” which connects two enzyme domains into an active, “buttoned up” conformation. Our study of the kinetic parameters of mutant enzymes has shown that Tyr102Phe substitution leads to an approximately 80-fold increase of the Michaelis constant relative to the native enzyme, unlike Phe311Trp and Phe311Tyr substitution of neighboring residue Phe311. Our analysis of the Tyr102Phe mutant in the open conformation by X-ray crystallography has shown that its overall fold remains almost the same as that of the native enzyme. Molecular dynamics simulations of the ternary complexes of the native FDH enzyme and its Tyr102Phe mutant showed that Tyr102Phe substitution results in the loss of an interdomain hydrogen bond between the Tyr102 and Gln313 residues, which, in turn, destabilizes the closed conformation and affects the isolation of the FDH active site from water molecules. Our structural investigations have shown that Tyr102Phe replacement also leads to the destruction of interdomain contacts of Phe102 with Phe311, Pro312 residues, and decreases the stability of the Leu103-Val127 beta bridge. Phylogenetic analysis also confirmed the importance of the Tyr102 residue for enzymes from the FDH family, in which it is absolutely conserved.
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•Tyr102 is strongly conserved in the FDH family.•Tyr102Phe substitution leads to an 80-fold decrease in the Michaelis constant.•Tyr102Phe substitution decreases stability of the Leu103-Val127 beta bridge.•Tyr102Phe substitution decreases interdomain contacts.•Tyr102 is necessary for the correct transition of FDH into the closed state.
Using the effective mass model for an electron and the dielectric continuum model, analytical solutions of the self-consistent Schrödinger-Poisson system of equations are obtained. Quantum mechanical ...theory of electronic stationary states, the oscillator strengths of quantum transitions and a method of potential profile calculation is developed for the experimentally constructed three-well resonance-tunneling structure — a separate cascade of quantum cascade detector. For the proposed method, a comparison with the results of other methods and with the results of the experiment was carried out. A good agreement between the calculated value of the detected energy and its experimental value has been obtained, the difference being no more than 2.5%.
We studied powder samples of europium titanates Eu1−x2+Eux3+TiO3+x/2 prepared by sol-gel and coprecipitation methods with subsequent thermal treatment of precursors at different conditions (reducing ...or oxidizing atmosphere, temperature, exposure time). We have revealed a radical rearrangement in the local, electronic and crystal structure of europium titanates depending on synthesis conditions. The combination of synchrotron X-ray diffraction (s-XRD), X-ray absorption (XANES and EXAFS), infrared and Raman spectroscopies supplemented by photoluminescence and simultaneous thermal analysis was used in our study. It is shown that the combined XANES and thermogravimetric analysis gives clear evidence of a change in the oxidation state of europium cations from Eu2+ to Eu3+. The comparison of s-XRD, XANES, and EXAFS data allowed us to explain the local structure rearrangement during the phase transition from cubic perovskite Eu2+TiO3 (space group Pm3¯m) to pyrochlore phase Eu23+Ti2O7 (space group Fd3¯m) through intermediate layered perovskite Eu23+Ti2O7 with monoclinic structure (space group P21) under annealing in oxidizing atmosphere. The Raman spectroscopy study pointed to significant changes in the local structure of the anionic sublattice upon the observed phase transitions. The analysis of luminescence spectra has shown that Eu3+ cations occupy positions with low local site symmetry in all synthesized compounds regardless of the type of the crystal structure.
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•Local, electronic and crystal structure rearrangement in Eu titanates was studied.•s-XRD, XAFS, IR, Raman spectroscopy, TGA and PL methods were used.•XANES and TGA are effective tools for determining the Eu2+/Eu3+ cations ratio.•Eu3+ cations occupy low local site symmetry positions in synthesized compounds.
An optimized method for analysis of free amino acids using a modified lithium-citrate buffer system with a Hitachi L-8800 amino acid analyzer is described. It demonstrates clear advantages over the ...sodium-citrate buffer system commonly used for the analysis of protein hydrolysates. A sample pretreatment technique for amino acid analysis of brain extracts is also discussed. The focus has been placed on the possibility of quantitative determination of the reduced form of glutathione (GSH) with simultaneous analysis of all other amino acids in brain extracts. The method was validated and calibration coefficient (
K
GSH
) was determined. Examples of chromatographic separation of free amino acids in extracts derived from different parts of the brain are presented.
IntroductionThe results of the study of psychological factors of hostility in depression are presented. The topicality of the study is due to hostility considered, on the one hand, as a property of ...depression, and on the other hand, as a risk factor, associated with the likelihood of auto-aggressive behavior.ObjectivesThe aim of the study was to analyze the relationship between hostility and attachment disorders in endogenous depression.MethodsThe study involved 49 patients with depressive disorder (mean age 19,8±4,5). All patients were assessed using the Hamilton Depression Rating Scale (HDRS-17 mean 21,03±6,02). All completed the following methods: Revised Experiences in Close Relationships (ECR-R); Symptom Check List-90-Revised (SCL-90R); Aggression Questionnaire by Buss and Perry (BPAQ); I-structural test by G. Ammon (ISTA). According to the “depression” parameter of the SCL-90R, the group was divided into subgroups with high and medium severity of depression. Analysis of variance (ANOVA) or Mann-Whitney test were. Correlation analysis (Spearman) and stepwise multiple regression analysis were also used.ResultsAt high levels of depression, the indicators of “hostility”, “destructive” and “deficit aggression” are statistically significantly higher. The severity of depression significantly correlates with the severity of “anxiety” in attachment (close relationships), as well as with pathological “narcissism”, “destructive external self-delimitation”, “deficient internal self-delimitation”.For the measure of depression, regression analysis showed that the regression model explained more than 76% of the variance, with the measures of “interpersonal sensitivity”, “deficit narcissism”, and “avoidance” in attachment making significant contributions. For the “hostility” the regression model explains about 62% of the variance, while a significant contribution is made, as in the analysis of “depression”, by the indicators of “interpersonal sensitivity” and “avoidance”, however, unlike “depression”, the contribution of the “destructive narcissism” is noted in contrast to the “deficit narcissism”.ConclusionsWith severe depressive symptoms, indicators of hostility are increased. Hostility in depression is associated with factors caused by a violation of early interpersonal relationships (anxious attachment), which causes increased sensitivity in relations with others, “building a barrier” between oneself and the external environment perceived as hostile in the narcissistic pathology, problems in emotional regulation. One of the targets of psychotherapeutic work may be the ambivalence between desire for symbiotic dependence and the experienced hostility.Disclosure of InterestNone Declared
Unlike the
OGDH
-encoded 2-oxoglutarate dehydrogenase (OGDH), which is an essential enzyme present in all animal tissues, expression of the
DHTKD1
-encoded isoenzyme, 2-oxoadipate dehydrogenase ...(OADH), depends on a number of factors, and mutant
DHTKD1
phenotypes are rarely manifested. Physiological significance of OADH is also obscured by the fact that both isoenzymes transform 2-oxoglutarate and 2-oxoadipate. By analogy with other members of the 2-oxo acid dehydrogenases family, OADH is assumed to be a component of the multienzyme complex that catalyzes oxidative decarboxylation of 2-oxoadipate. This study aims at molecular characterization of OADH from animal tissues. Phylogenetic analysis of 2-oxo acid dehydrogenases reveals OADH only in animals and
Dictyostelium discoideum
slime mold, within a common branch with bacterial OGDH. Examination of partially purified animal OADH by immunoblotting and mass spectrometry identifies two OADH isoforms with molecular weights of about 130 and 70 kDa. These isoforms are not observed upon the expression of human
DHTKD1
protein in either bacterial or yeast system, where the synthesized OADH is of expected molecular weight (about 100 kDa). Thus, the OADH isoforms present in animal tissues, may result from the animal-specific regulation of the
DHTKD1
expression and/or posttranslational modifications of the encoded protein. Mapping of the peptides identified in the OADH preparations, onto the protein structure suggests that the 70-kDa isoform is truncated at the N-terminus, but retains the active site. Since the N-terminal domain of OGDH is required for the formation of the multienzyme complex, it is possible that the 70-kDa isoform catalyzes non-oxidative transformation of dicarboxylic 2-oxo acids that does not require the multienzyme structure. In this case, the ratio of the OADH isoforms in animal tissues may correspond to the ratio between the oxidative and non-oxidative decarboxylation of 2-oxoadipate.
NAD
+
-dependent formate dehydrogenase from
Staphylococcus aureus
(SauFDH) is one of the key enzymes responsible for the survival of this pathogen in the form of biofilms. 3D structure of the enzyme ...might be helpful in the search for highly specific SauFDH inhibitors that can be used as antibacterial agents exactly against
S. aureus
biofilms. Here, we prepared a recombinant SauFDH in
Escherichia coli
cells with a yield of 1 g target protein per liter medium. The developed procedure for the enzyme purification allowed to obtain 400 mg of homogenous enzyme with 61% yield. The specific activity of the purified recombinant SauFDH was 20 U per mg protein, which was 2 times higher than the previously reported activities of formate dehydrogenases. We also found crystallization conditions in the course of two rounds of optimization and obtained 200- and 40-µm crystals for the SauFDH apo- and holoenzymes, respectively. X-ray analysis using synchrotron X-ray sources produced diffraction data sufficient for solving the three-dimensional structures of the apo- and holoenzymes with the resolution of 2.2 and 2.7 Å, respectively. Crystals of the apo- and holoforms of SauFDH had different crystal space groups, which suggest coenzyme binding in the SauFDH holoenzyme.
The problem of general non-linear stochastic optimal control with small Wiener noise is studied. The problem is approximated by a Markov Decision Process. Bellman Equation is solved using Value ...Iteration (VI) algorithm in the low rank Tensor Train format (TT-VI). In this paper a modification of the TT-VI algorithm called TT-Q-Iteration (TT-QI) is proposed by authors. In it, the nonlinear Bellman Optimality Operator is iteratively applied to the solution as a composition of internal Tensor Train algebraic operations and TT-CROSS algorithm. We show that it has lower asymptotic complexity per iteration than the method existing in the literature, provided that TT-ranks of transition probabilities are small. In test examples of an underpowered inverted pendulum and Dubins cars our method shows up to 3–10 times faster convergence in terms of wall clock time compared with the original method.
Molecular mechanisms of long-term changes in brain metabolism after thiamine administration (single i.p. injection, 400 mg/kg) were investigated. Protocols for discrimination of the activities of the ...thiamine diphosphate (ThDP)-dependent 2-oxoglutarate and 2-oxoadipate dehydrogenases were developed to characterize specific regulation of the multienzyme complexes of the 2-oxoglutarate (OGDHC) and 2-oxoadipate (OADHC) dehydrogenases by thiamine. The thiamine-induced changes depended on the brain-region-specific expression of the ThDP-dependent dehydrogenases. In the cerebral cortex, the original levels of OGDHC and OADHC were relatively high and not increased by thiamine, whereas in the cerebellum thiamine upregulated the OGDHC and OADHC activities, whose original levels were relatively low. The effects of thiamine on each of the complexes were different and associated with metabolic rearrangements, which included (i) the brain-region-specific alterations of glutamine synthase and/or glutamate dehydrogenase and NADP
+
-dependent malic enzyme, (ii) the brain-region-specific changes of the amino acid profiles, and (iii) decreased levels of a number of amino acids in blood plasma. Along with the assays of enzymatic activities and average levels of amino acids in the blood and brain, the thiamine-induced metabolic rearrangements were assessed by analysis of correlations between the levels of amino acids. The set and parameters of the correlations were tissue-specific, and their responses to the thiamine treatment provided additional information on metabolic changes, compared to that gained from the average levels of amino acids. Taken together, the data suggest that thiamine decreases catabolism of amino acids by means of a complex and long-term regulation of metabolic flux through the tricarboxylic acid cycle, which includes coupled changes in activities of the ThDP-dependent dehydrogenases of 2-oxoglutarate and 2-oxoadipate and adjacent enzymes.
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