Biochemistry (Moscow), Supplement Series B: Biomedical Chemistry (v.7, #3)

Metabolic profiling of human blood by O. P. Trifonova; P. G. Lokhov; A. I. Archakov (179-186).
Metabolomics, the youngest branch of “omics” sciences, intended for studying a comprehensive set of low molecular weight substances (metabolites) of various biological objects. Metabolite profiles represent a molecular phenotype of biological systems and reflect information encoded at the genome level and realized at the transcriptome and proteome levels. Analysis of the human blood metabolic profile is a universal and promising tool for clinical applications because it is a sensitive measure of both endogenous and exogenous (environmental) factors influencing the human body. In this review we discuss technical implementation of blood metabolic profiling methods and statistic analysis of metabolite profiles for effective diagnostics and risk assessments of diseases.
Keywords: metabolomics; metabolite profile; mass spectrometry; blood plasma/serum

The SPR-based biosensor test system for analysis of small compounds interaction with human cytochrome P450 51A1 (CYP51A1) by O. V. Gnedenko; L. A. Kaluzhskiy; A. A. Molnar; A. V. Yantsevich; D. V. Mukha; A. A. Gilep; S. A. Usanov; V. A. Stonik; A. S. Ivanov; A. V. Lisitsa; A. I. Archakov (187-195).
The SPR-based test for human cytochrome P450 51A1 (CYP51A1) ligand screening has been developed. Applicability of this system has been validated with known azole inhibitors of cytochromes P450. The studied azoles selectively interacted with human cytochrome P450 51A1, which showed the highest affinity towards ketoconazole. The efficiency of the SPR based assay has been tested using 19 steroid and triterpene compounds, which have not been investigated as potential ligands of CYP51A1.
Keywords: human cytochrome P450 51A1 (CYP51A1); SPR-based test-system assay; CYP51A1 inhibitors; azoles; interaction screening; atherosclerosis

Molecular modeling of the interaction of 17(20)Z- and 17(20)E-pregna-5,17(20)-dien-21-oyl amides with the nuclear receptor LXRβ by I. V. Fedyushkina; S. V. Stulov; N. O. Dugin; A. Yu. Misharin; A. R. Mehtiev; G. E. Morozevich; A. V. Veselovsky (196-201).
Eight isomeric 17(20)Z- and 17(20)E-pregna-5,17(20)-dien-21-oyl amides, conformationally rigid oxysterol analogues, differing in the structure of the amide moiety have been analyzed. Analysis of low energy conformers revealed that all 17(20)E-isomers had three main energy minima (corresponding to the values of the dihedral angle θ20,21 (C17=C20-C21=O) about ∼0°, ∼120°, and ∼240°); the most occupied minimum corresponded to θ20,21 about ∼0°. 17(20) Z-Isomers had either one or two pools of stable low energy conformations. Molecular docking of these compounds to the ligand-binding site of the nuclear receptor LXRβ (a potential target) demonstrated high probability of binding of E-isomers but not Z-isomers with this target. Results of the molecular modeling were confirmed by an experiment in which stimulation of triglyceride biosynthesis in Hep G2 cells in the presence of 17(20)E-3β-hydroxypregna-5,17(20)-dien-21-oyl (hydroxyethyl)amide was demonstrated.
Keywords: pregna-5,17(20)-dien-21-oyl amides; docking; LXR; nuclear receptor; triglyceride biosynthesis; Hep G2

Glycine receptors in the nervous tissue and their functional role by V. N. Nikandrov; T. V. Balashevich (202-211).
The literature data on glycine metabolism in neural tissue, the mitochondrial Gly-cleaving system, and the Gly uptake system in neural and glial cells are summarized. Brief description of localization and abundance of glycine receptors and specific binding sites in the mammalian nervous tissue is given. Four types of glycine binding receptors are described. These include own specific glycine receptor (GlyR), ionotropic glutamate receptor, which selectively binds N-methyl-D-aspartate selectively (NMDAR), and ionotropic receptors of γ-aminobutyrate (GABAAR, GABACR).
Keywords: glycine; metabolism; receptors; neuroprotective action; nervous tissue cells; regulatory effects

Ongoing studies carried out worldwide using banks of tumor and non-tumor samples give evidence that perspective markers predicting response of individual patients to intended radiation therapy may be found among some apoptotic indexes, spectrum of specialized proteins, and DNA-based microarray molecular profiling analysis as well determination of single nucleotide polymorphisms in genome of the patients. In the last years there is increasing interest in radiogenomics and characterization of gene expression profiles by DNA (micro)array technique that can predict radioresponse of tumor and non-tumor tissues. So far there are only a few reliable molecular markers predicting response of tumor and non-tumor tissues to radiation.
Keywords: tumor and non-tumor tissues; radiation therapy efficacy; predictive biotesting; biochemical markers

Is cobalt-binding capacity of serum a new perspective diagnostic marker? by E. A. Litus; V. G. Zaitsev; V. E. Verovsky; L. V. Goncharova; G. P. Dudchenko; O. V. Ostrovskij (222-225).
The aim of this study was to determine the reference interval of serum cobalt-binding capacity (CoBC), and to estimate the effect of factors unrelated to oxidative modification of serum albumin on this diagnostic marker. Healthy volunteers (n = 194), patients with autoimmune diseases (n = 44) and patients with type 2 diabetes mellitus (n = 50) were enrolled in the study. The regional reference interval was found to be 0,462–0,744 mmol Co2+/L of serum. The study of serum CoBC in patients with autoimmune pathology and type 2 diabetes mellitus has shown that the CoBC level strongly depends on the serum protein profile. Therefore, this test may be used to diagnose the ischemia, although other pathologies associated with changes in the ratio of blood protein fractions can also influence the serum CoBC level.
Keywords: cobalt-binding capacity; type 2 diabetes mellitus; autoimmune diseases; blood protein fractions

Comparative studies of the genotoxic activity of a new palladium (II) acidocomplex and cisplatin in human blood lymphocytes in vitro by A. K. Grekhova; L. B. Gorbacheva; N. A. Ivanova; I. A. Efimenko; A. N. Osipov (226-230).
Comparative studies on the genotoxic activity of cisplatin and morfozol, the first representative of a new class of cation-anion complexes of palladium [AH]2[PdCl4] (where A—methylmorpholine), have been performed using human lymphocytes in vitro. The results of the DNA-DNA cross-linking activity investigations showed that both compounds studied exhibited biphasic dose-effect relationship: a linear decrease of the DNA percent in the comet tail and a plateau region. However, in the plateau region, morfozol caused a 6-fold reduction of the DNA percent in the comet tail while cisplatin caused a 2-fold decrease only. Morfozol, like cisplatin, inducing DNA-protein cross-linking and generating reactive oxygen species, was more effective than cisplatin.
Keywords: anticancer drugs; cisplatin; palladium compounds; morfozol; DNA damage; oxidative stress

The effect of food supplementation with chromium (CrCl3 · 6H2O) on intensity of peroxide processes and activity of antioxidant enzymes has been investigated in some rat tissues. Food supplementation with 200 μg/kg CrCl3 · 6H2O for 30 days resulted in the increase of tissue chromium. The tissue chromium content of chromium-treated rats decreased in the following order: spleen, heart, kidney, lung, brain, liver, skeletal muscles.All organs and tissues (except skeletal muscles) of chromium-treated rats were characterized by decreased content of lipid peroxidation (LPO) products: hydroperoxides and thiobarbituric acid reactive substances (TBARS). The maximal reduction in LPO products was observed in spleen, kidney, liver, and lung. Treatment with chromium also caused an increase in the activity of glutathione peroxidase, glutathione reductase, and calatase in all tissues and organs studied. In the brain and kidney an increase in the content of reduced glutathione was observed. Superoxide dismutase activity was higher in myocardium and skeletal muscles, basically equal in lung and liver, while in other organs (brain, kidney, spleen) of experimental animals it was lower than in control animals. Results of this study suggest that chromium exhibits tissue/organ-specific regulatory effects on enzymes of the antioxidant defense
Keywords: chromium; TBARS; glutathione peroxidase; glutathione reductase; catalase; reduced glutathione

Pharmacological activity of echinochrome a alone and in the biologically active additive Timarin by A. A. Artyukov; A. M. Popov; A. V. Tsybulsky; O. N. Krivoshapko; N. V. Polyakova (237-242).
Pharmacological activity of echinochrome A (EchA) alone and in the biologically active additives (BAA) Timarin, administered per os has been investigated in volunteers. Blood hematological, immunological, and biochemical parameters were investigated before and after administration of the substances used. EchA decreased serum glutatione (GSH) and increased catalase activity 1 h after treatment. Later (3 h after administration) catalase activity normalized, while GSH exceeded the initial level. Changes in blood lipids suggest decreased risk of atherogenesis. Changes found in blood sex hormone levels indicate that Timarin may influence sex gland functioning. Changes of hematological and immunological parameters have been interpreted as the result of a mild stressor effect of both EchA alone and in the BAA Timarin increasing adaptation reactivity of the body.
Keywords: echinochrome A; 1,4-naphtoquinone; pharmacological activity; mechanism of action

Peptidase activity of the fetoplacental complex in the norm and pathology by O. P. Petrushova; N. I. Mikulyak (243-246).
The activity of angiotensin converting enzyme (ACE), carboxypeptidase N (CPN), and leucine aminopeptidase (LAP) has been investigated in the fetoplacental complex (FPC) in norm and fetoplacental insufficiency (FPI). ACE and LAP activities were significantly higher in the placental tissue than in maternal serum and umbilical vein serum. CPN activity was significantly lower in umbilical vein serum as compared to that of women in childbirth. Probably, the studied enzymes are involved in formation of reduced vascular sensitivity of FPC during physiological pregnancy. In cases of placental insufficiency a significant increase of LAP activity was found in the placental tissue and umbilical vein serum. In addition, the pathological course of pregnancy caused a significant increase of CPN activity in serum of pregnant women in comparison to the norm. The obtained data suggest that during FPI proteolytic enzymes participate in the formation of compensatory-adaptive reactions in the FPC. Results of this study are interesting in the context of development of methods for prevention and correction of metabolic disorders in pathologies of pregnancy.
Keywords: placenta; fetoplacental complex; leucine aminopeptidase; carboxypeptidase N; angiotensin-converting enzyme

Using the L-phenylalanine secreting strain of Gram-negative aerobic facultative methylotrophic bacteria Brevibacterium methylicum, assimilating methanol via the ribulose-5-monophosphate (RuMP) cycle of carbon assimilation, as an example, we have continued studies on the use of methylotrophic bacteria for the preparative microbial synthesis of amino acids labeled with stable isotopes, including deuterium (2H), suitable for biomedical applications and clinical diagnostics. Here we demonstrate the data on adaptation of the methylotrophic bacterium B. methylicum to the maximal concentration of deuterium in the growth medium with 98% (v/v) 2H2O and 2% (v/v) [2H]MeOH, and biosynthesis of deuterium labeled L-phenylalanine with different levels of isotopic enrichment. The strain was adapted to 2H2O by means of plating of initial cells on solid (2% agarose) minimal growth media M9 with an increasing gradient of 2H2O concentration from 0, 24.5, 49.0, 73.5 up to 98% (v/v) 2H2O and subsequent selection of individual colonies stable to the action of 2H2O, which were capable to produce L-phenylalanine. L-phenylalanine was extracted from the growth medium with isopropanol followed by subsequent crystallization in ethanol (output 0.65 g/L). Using the developed method of microbial synthesis it is possible to obtain deuterated L-phenylalanine with different levels of isotopic enrichment, depending on concentration of 2H2O in growth media, from 17% (the growth medium with 24.5% (v/v) 2H2O) right up to 75% (the growth medium with 98% (v/v) 2H2O) of deuterium as evidenced by results of the electron impact (EI) mass-spectrometry analysis of methyl ethers of N-dimethylamino(naphthalene)-5-sulfonyl chloride (dansyl) phenylalanine isolated from growth media under different experimental conditions.
Keywords: Brevibacterium methylicum ; L-phenylalanine; biosynthesis; heavy water; electron impact mass spectrometry