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The purpose of the research was to study the content of ceruloplasmin, copper and copper-coordinating amino acids (in the molecule of the studied protein) in the milk of white-and- black cows at different periods of lactation. The research methods were: modified method for determining ceruloplasmin by Revin; copper was determined using a ZEEnit 650 P atomic adsorption spectrometer (Analytik Jena AG, Germany), amino acid analysis was performed using a highly efficient LC-20 Prominence liquid chromatography system (Shimadzu, Tokyo, Japan) equipped with a reaction module for post-column derivatization with ninhydrin ARM-1000 (Sevko & Co., Moscow, Russia). The study groups were formed depending on the timing of lactation: 1-2 months – group 1; 3-5 months – group 2; 6-7 months – group 3; 8-9 months – group 4. Each group included 8 animals. The Mann-Whitney U criterion and Pearson correlations were used for statistical processing of the obtained results. The average values of ceruloplasmin levels in the groups were 0.44-0.49 mg/ml, copper 71-83 mcg/l, histidine 0.11-013 g/100g, cysteine 0.03 g/100g and methionine 0.09-0.11 g/100g. In these 4 groups, small differences were found in the content of ceruloplasmin, copper and three amino acids, but only for the second and fourth groups of histidine, these differences were significant. Thus, the indicated intervals of ceruloplasmin and copper content can serve as important guidelines for determining the "reference intervals" for cow milk in general, while it is not necessary to take into account different lactation periods.

This research is investigation of an influence the effect of thyroid hormones on the processes of desulfuration of cysteine in animal organs and the possibility of correcting the activity of cysteine metabolism enzymes in hyper- and hypothyroidism. The aim – to investigate in an experiment the possibility of correcting the processes of cysteine metabolism and the synthesis of hydrogen sulfide, which are violated under the influence of hyper- and hypothyroidism, using folic acid, cyanocobalamin, pyridoxine and betaine. Object and methods. The work was performed on adult male rats in which the effects of hyperthyroidism (intragastric L-thyroxine for 21 days 200 µg/kg*day), hypothyroidism (thiamazole 10 kg*day) on the reactions of desulfurization cysteine in animal organs and correction were studied disturbed under the influence of L-thyroxine and thiamazole cysteine metabolism using folic acid, cyanocobalamin, pyridoxine and betaine. In the liver and kidneys, the activity of cystathionine-β-synthase (CBS), cystathionine-γ-lyase (CSE), and cysteine aminotransferase (CAT) was determined. In the brain, the activity of CBS was determined, in the myocardium – CAT, in the blood serum – the content of hydrogen sulfide (H2 S) and the concentration of total cysteine. The experimental data were compared with the control values and data from research groups of animals with hyper- and hypothyroidism. Study results. Hyperthyroidism caused an increase in the activity in the kidneys of rats of CBS and CSE (by 21 and 100%), and in the brain only of CBS (by 60%) compared with intact animals. To a greater extent, the activity of CBS increased with the administration of vitamin B6 to animals with hyperthyroidism. The activity of CSE also increased in the liver and kidneys (by 29 and 140%) when pyridoxine was used in animals with hyperthyroidism. The activity of СAT in the kidneys and brain with the introduction of vitamin B6 with thyroxine increased by 34 and 86%. Betaine caused an increase in the activity of CBS in animals with hyperthyroidism in the brain and kidneys by 24 and 69% against the control, while the activity of CSE increased in the kidneys by 105%. The combination of B9 and B12 in the kidneys led to an increase in the activity of CBS and CSE (by 33 and 115%) compared with the group of animals without correction, and in the brain to an increase in the activity of CBS by 71%. The combination of B9 , B12, B6 and betaine in animals with hyperthyroidism led to an increase in the activity of CBS and CSE, and in the kidneys of CBS, CAT and CSE. In the brain with hyperthyroidism, the combination of B9 , B12, B6 and betaine caused an increase in the activity of CBS by 94% compared with the control. L-thyroxine did not affect the level of cysteine in the blood of experimental animals. The drugs we selected and their combination did not cause significant changes in the concentration of cysteine in the blood of rats with hyperthyroidism. Administration to animals with hyperthyroidism B6 caused an increase in the level of H2 S to a greater extent – by 27% versus control. Betaine in animals with hyperthyroidism led to an increase in H2 S level by 32%, a combination of vitamins B9 and B12 – by 31%, and a combination of all mediators by 36% compared with intact animals. Thiamazole suppressed the activity of enzymes compared to intact animals: in the kidneys, the activity of CBS, CSE and CAT, respectively, by 19, 28, 39%, in the brain, the activity of CBS by 34%. Pyridoxine led to an increase in the liver activity of CBS and CSE (by 43 and 44%) compared with the group of animals in which hypothyroidism was caused, and in the kidney, the activity of CBS, CSE, and CAT. The combination of B vitamins and betaine caused an increase in the activity of CBS and CAT (by 48 and 46%) in the liver of animals with hypothyroidism. Thiamazole caused an increase (by 40%) in the serum cysteine content of animals and a decrease in H2 S level (by 24%). Only vitamin B6 led to a decrease (by 22%) in the concentration of cysteine in the blood during hypothyroidism. B6 also caused an increase (by 23%) in the level of H2 S compared with a group of animals with hypothyroidism. B9 and B12 and a combination of all mediators caused in animals with hypothyroidism an increase in the H2 S level (by 22% and 28%) compared with the group of animals with hypothyroidism. Conclusions. Hyper- and hypothyroidism cause an imbalance in the exchange of cysteine. The preparations we have chosen and their combination do not produce a reliable effect on the level of this amino acid in hyperthyroidism, but they increase the level of hydrogen sulfide in the blood serum of animals. In hypothyroidism, vitamins B6 , B12, B9 and betaine partially prevent the disruption of cysteine metabolism and lead to an increase in the level of hydrogen sulfide in serum blood, which indicates the possibility of preventing endothelial dysfunction in hypothyroidism using these drugs.

Homocystinuria is a genetically heterogeneous hereditary disease from the group of aminoacidopathies caused by a metabolic disorder of sulphur-containing amino acids, primarily methionine. The article presents the etiopathogenetic, diagnostic and therapeutic aspects of this disease and covers modern opportunities of biochemical and molecular diagnostics. The approach to dietary and pharmacological correction of metabolic disorders in homocystinuria and the general strategy of patients’ management are described in detail. Important information is given for physicians of various disciplines and parents of patients.

Diabetes mellitus remains one of the most pressing problems in the 21st century. The research objective was to study the level of glycotoxins and oxidation-reduction potential of a serum/cell at the complicated and uncomplicated course of diabetes mellitus type 2 was and find the ways to correct the disorder. Materials and methods. There were three groups of patients aged from 55 up to 70 years, a total of 80 people. The main group (group No. 3) was presented by 40 men suffering from a diabetes mellitus type 2 with a macrovascular disease of the lower extremities. Depending on the chosen remedy and the carried-out therapy, patients of the first group were sectioned into subgroups on 20 people, in the 2nd subgroup additionally N-acetylcysteine solution was introduced in a daily dose of 600 mg once, intravenously, within 7 days. Definition of the studied indicators was made by HPLS methods, prior to therapy, for the third and seventh day. Results. Obtained data confirm activation of polyol pathway of glucose disintegration. Antioxidant potential of the serum and cells decreases. Using N-acetylcysteini in complex therapy reduces implications of carbonyl and oxidative stresses, due to multiple decrease of the final products of glycation. Conclusions. When identifying the presented indicators in patients with diabetic macroangiopathy of the lower extremities, the administration of N-acetylcysteine, intravenously in a daily dose of 600 mg (7 days course) is recommended.

We studied the pharmacokinetic parameters of N-acetylcysteine at a single oral and intravenous administration of healthy volunteers. Elimination half-life (t1/2) within 2 hours and the maximum concentration in blood (Cmax) after 1 hour after administration was more than 10 times higher when intravenous on a comparison with oral administration. Effect of exchange rate on intravenous administration of N-acetylcysteine (1 200 mg/day for one week) the concentration of endogenous aminothiols in patients with diseases that are accompanied by oxidative stress (ischemic stroke and vascular complications of diabetes mellitus) are also studied. In ischemic stroke example it shows that the concentration of cysteine in plasma was increased by 1,6 times, and homocysteine levels did not change, as the ratio of reduced and oxidized cysteine (CysSH / CysSSCys). The content of total and free glutathione in red blood cells was not significantly changed, but significantly increased GSH / GSSG ratio, characterizing the antioxidant capacity of cells. We conclude that the inclusion in the scheme of N-acetylcysteine therapy of these diseases, accompanied by a more favorable clinical outcome due to a significant increase in antioxidant capacity of cells and tissues.

Counteraction of the origin and distribution of multidrug-resistant pathogens responsible for intra-hospital infections is a worldwide issue in medicine. In this brief review, we discuss the results of our recent investigations, which argue that many antibiotics, along with inactivation of their traditional biochemical targets, can induce oxidative stress (ROS production), thus resulting in increased bactericidal efficiency. As we previously showed, hydrogen sulfide, which is produced in the cells of different pathogens protects them not only against oxidative stress but also against bactericidal antibiotics. Next, we clarified the interplay of oxidative stress, cysteine metabolism, and hydrogen sulfide production. Finally, demonstrated that small molecules, which inhibit a bacterial enzyme involved in hydrogen sulfide production, potentiate bactericidal antibiotics including quinolones, beta-lactams, and aminoglycosides against bacterial pathogens in in vitro and in mouse models of infection. These inhibitors also suppress bacterial tolerance to antibiotics by disrupting the biofilm formation and substantially reducing the number of persister bacteria, which survive the antibiotic treatment. We hypothesise that agents which limit hydrogen sulfide biosynthesis are effective tools to counteract the origin and distribution of multidrug-resistant pathogens.

Cysteine cathepsins (Cts) also known as thiol proteinases belong to the superfamily of cysteine proteinases (EC 3.4.22). Cts are known as lysosomal proteases responsible for the intracellular proteins degradation. All Cts are synthesized as zymogens, activation of which occurs autocatalytically. Their activity is regulated by endogenous inhibitors. Cts can be secreted into the extracellular environment, which is of particular importance in tumor progression. Extracellular Cts not only hydrolyze extracellular matrix (ECM) proteins, but also contribute to ECM remodeling, processing and/or release of cell adhesion molecules, growth factors, cytokines and chemokines. In cancer, the expression and activity of Cts sharply increase both in cell lysosomes and in the intercellular space, which correlates with neoplastic transformation, invasion, metastasis and leads to further tumor progression. It has been shown that Cts expression depends on the cells type, therefore, their role in the tumor development differs depending on their cellular origin. The mechanism of Cts action in cancer is not limited only by their proteolytic action. The Cts influence on signal transduction pathways associated with cancer development, including the pathway involving growth factors, which is mediated through receptors tyrosine kinases (RTK) and various signaling mitogen-activated protein kinases (MAPK), has been proven. In addition, Cts are able to promote the epithelial-mesenchymal transition (EMT) by activating signal transduction pathways such as Wnt, Notch, and the pathway involving TGF-β. So, Ctc perform specific both destructive and regulatory functions, carrying out proteolysis, both inside and outside the cell.

Disulfide bridges are essential for maintaining the structure and function of proteins. Traditionally, studies of the disulfide bonds require expensive equipment and high purity of the protein sample, therefore, the development of simpler techniques is warranted. Here, were present a novel protocol for the detection of disulfide bonds in proteins, which is based on the labeling reduced disulfide bridges with a high molecular weight (HMW) maleimide derivative. After irreversible blocking of free thiol groups of proteins, the labeling of new thiols released from disulfide bridges with a high-molecular-weight (HMW) maleimide derivative is performed. To confirm localization of cysteines involved in the formation of disulfide bonds, cysteine mutagenesis was conducted. For validation, aquaporin 5 (AQP5) and transient receptor potential cation channel subfamily V member 4 (TRPV4) proteins were tagged with FLAG (DYKDDDDK) on N-termini. Increase in MW of the target proteins from immunoblot indicated the presence of disulfide bonds. No bands with increased MW were detected in AQP5, while TPRV4 cysteines at disulfide bridges-constituting positions 639, 645, 652, 660, 770 were detected and confirmed by cysteine mutagenesis. These data indicate that the proposed technique is feasible and effective for the detection of protein disulfide bonds.

Mass spectrometric proteomic analysis at the sample preparation stage involves the artificial reduction of disulfide bonds in proteins formed between cysteine residues. Such bonds, when preserved in their native state, complicate subsequent enzymatic hydrolysis and interpretation of the research results. To prevent the re-formation of the disulfide bonds, cysteine residues are protected by special groups, most often by alkylation. In this review, we consider the methods used to modify cysteine residues during sample preparation, as well as possible artifacts of this stage. Particularly, adverse reactions of the alkylating agents with other amino acid residues are described. The most common alkylating compound used to protect cysteine residues in mass spectrometric proteomic analysis is iodoacetamide. However, an analysis of the literature in this area indicates that this reagent causes more adverse reactions than other agents used, such as chloroacetamide and acrylamide. The latter can be recommended for wider use. In the review we also discuss the features of the cysteine residue modifications and their influence on the efficiency of the search for post-translational modifications and protein products of single nucleotide substitutions.

The superoxide-generating reaction of adrenaline autoxidation in an alkaline medium, used in vitro to identify the antioxidant properties of various compounds, simulates the complex multistep process of quinoid oxidation of catecholamines (CA) in the body. Sulfur-containing cysteine (Cys) and reduced glutathione (GSH), as well as oxidized glutathione (GSSG), have been shown to inhibit this process. The studied substances were considered as inhibitors of quinoid oxidation and are evaluated as antioxidants. The IC50 values for Cys and GSH were close to 7.5 mM. Inhibition by GSSG was weaker; represented approximately 50-70% of Cys and GSH. Other sulfur-containing compounds that differ in chemical structure, the amino acids taurine and methionine were ineffective. The interest in this model and the search for effective compounds acting on this reaction is associated with one of the mechanisms of the etiopathogenesis of Parkinson's disease (PD) discussed in the literature, which occurs when the biochemical transformations of dopamine CA and its quinoid oxidation process are violated. Cys, GSH and GSSG in the model system inhibit quinoid oxidation of adrenaline, as a result of which the formation of superoxide (O2 ·-) is also inhibited. Experiments with the superoxide-generating enzymatic reaction xanthine xanthioxidase, the chemistry of which is different and not related to formation of quinoid metabolites, showed that the studied substances did not inhibit O2 ·- formation in this model. Thus, it was established that the biologically active sulfur-containing compounds Cys, GSH and GSSG are specific inhibitors of quinoid oxidation of CA, and are likely to be able to play the role of a neuroprotector. It is proposed to use these compounds in the treatment and prevention of PD by activating their biosynthesis in the body.