Biochemistry (Moscow), Supplement Series B: Biomedical Chemistry最新文献

Background: Obstructive sleep apnea (OSA) is characterized by recurrent apnea/hypopnea in the upper airways and oxygen desaturation accompanying respiratory events during sleep. Our study aimed to determine serum LECT2 and sclerostin levels in OSA patients. Methods: Patients who applied with the suspicion of OSA in the polysomnography unit of our hospital between June 2022 and April 2023 and who completed the polysomnography test were included in our study. Group 1: apnea-hypopnea index (AHI) < 5/h control group (n = 80), Group 2: OSA patients with AHI ≥ 5/h without comorbidity (n = 80). Results: When comparing the LECT2 and sclerostin levels of the groups, it was observed that there was a statistically significant difference, higher in the OSA group (p < 0.001 for both). When the groups were compared, it was observed that only LECT2 and sclerostin levels were higher in severe OSA patients than in mild OSA patients (p = 0.008, 0.02, respectively). A positive correlation was observed between LECT-2 level and AHI, apnea–hypopnea index during rapid eye movement (REM-AHI), and ODI levels (r = 0.55, p = 0.01, r = 0.42, p = 0.01, r = 0.61, p = 0.01). An inverse correlation was observed between LECT2 and minimum oxygen saturation (r = –0.42, p = 0.01). In the analysis performed with sclerostin level, a positive correlation (r = 0.42, p = 0.01, r = 0.28, p = 0.05, r = 0.53, p = 0.01) was observed with AHI, REM-AHI, and ODI, while an inverse correlation was observed between minimum oxygen saturation. Correlation was observed (r = –0.33, p = 0.01). Conclusion: Serum LECT2 and sclerostin levels in OSA patients can be used to determine the AHI and minimum oxygen saturation levels of individuals and their weight in OSA patients.

Resistance of bacteria to antibiotics is a serious human concern since it affects medical treatments performance against bacterial infections. Within the scope of new robust antibiotics development, we propose a heterocyclic–clay composite material. It consists of the association of 5-(2 pyridyl)-1-3-4-oxadiazoles-2-thione with pre-treated montmorillonite (MMT). Different pre-treatments were considered including acidification (H⁺-MMT) and intercalation with polar polymers facilitating the antibacterial composite material synthesis. The different composite materials that vary in terms of oxadiazole concentration were characterized in terms of structure (molecular, crystalline) using FTIR and XRD, and antibacterial properties. The obtained results showed successful intercalation of polar polymeric materials within acidified montmorillonite clay. The final composite material showed very promising antibacterial properties with reference to two well established antibiotics i.e., Penicillin and Spiramycine. The highest performance was observed for the composite containing polyvinyl alcohol intercalating the acidified montmorillonite with 50 wt % of 1,3,4-oxadiazole compound.

Kindia tick virus (KITV) is a novel, segmented, unclassified flavi-like virus of the Flaviviridae family. This virus is associated with ixodes ticks and is potentially pathogenic to humans. The main goal of this work was to search for structural motifs of viral polypeptides and to model the 3D structure of the NS3 and NS5 viral proteins of the multicomponent flavi-like KITV. Materials and methods. Genome-wide sequences of KITV, Zika, dengue, Japanese encephalitis, West Nile, and yellow fever viruses from the GenBank database were used. Bioinformatics analysis was performed using the AlphaFold2, RCSB PDB, UCSF Chimera, NCBI BLAST, MOTIF Search, Protomenal, Unipro UGENE, and ESPript software package. Results. Analysis of the VP1–VP3 structural proteins of the KITV showed that they have no analogues with currently known viral proteins. Spatial models of two viral nonstructural NS3 and NS5 proteins of the KITV have been obtained. These models had a high level of topological similarity to the NS3 and NS5 proteins of the tick-borne encephalitis and dengue viruses. The domains of methyltransferase and RNA-dependent RNA-polymerase were found in the NS5 KITV and this was also represented by subdomains of fingers, palm, and thumb and motifs A–F. The helicase domain and its main structural motifs I, Ia, II, III, IV, IVa, V, and VI were identified in NS3 KITV. However, the serine protease domain typical for NS3 flaviviruses was not detected. The highly conserved motives 3–7 amino acids in length, typical for unsegmented flaviviruses, were detected in the NS3 and NS5 KITV. Also, eight amino acid substitutions were detected for KITV/2018/1 and KITV/2018/2, five of which are localized in alpha-helix and three in loops of nonstructural proteins. Conclusions. Nonstructural proteins of segmented flavi-like KITV have structural and functional similarities with unsegmented flaviviruses. This confirms their possible evolutionary and taxonomic relationships.

MicroRNAs are a class of small noncoding RNAs that are 18 to 25 nucleotides in length and are found in most eukaryotic organisms. MicroRNAs can play an important role in epigenetic mechanisms of genome regulation, including DNA methylation and RNA and histone modification. Current methods for detecting and quantifying miRNAs rely heavily on cloning, Northern blotting, or primer extension, but each requires a pure preparation of the RNA type being analyzed. The standard method of RNA isolation, based on phenol−chloroform extraction with specific coprecipitants of nucleic acids, allows one to obtain preparations of total cellular RNA with a predominance of high-molecular types of ribonucleic acids. This greatly complicates the identification and quantification of microRNAs in sample preparations. Modification of the method of phenol−chloroform extraction of RNA, based on its precipitation of DNA with a specific precipitant, such as lithium chloride, showed that the use of polyethylene glycol 1500 using 2.5 M LiCl as a precipitant in the presence of 96% ethanol provides high yield and high-quality extraction of microRNA, which can be used for further analytical studies. Carrying out PCR to assess the quality of the isolated microRNA with specific primers for miR165a showed the presence of one amplification product approximately 80 bp in size, which corresponds to the theoretical values calculated on the basis of the developed probe for this microRNA. A positive PCR result indicates the presence of the analyzed microRNA in the matrix used. Consequently, the use of a modified RNA isolation technique using polyethylene glycol 1500 (PEG 1500) as an element for separating high- and low-molecular weight nucleic acids made it possible to obtain microRNA preparations that can be used for further analytical studies.

It is known that amino acid compositions for parenteral nutrition exhibit an immunomodulatory effect on T and B lymphocytes and phagocytes. The aim of the study was to evaluate the effect of amino acid compositions on the antibacterial functions of human peripheral blood neutrophilic granulocytes (neutrophils) under various experimental conditions.Materials and methods. Neutrophils were preincubated with amino acid compositions Amin or Vamin 14, then phorbol-myristate-acetate (PMA) was added and the respiratory burst of neutrophils was evaluated by flow cytometry. In another model, neutrophils were incubated with bacteria (S. aureus) at a ratio of 1 : 10 or 10 : 1, washed, incubated with amino acid compositions, washed again, lysed, and inoculated on meat-peptone agar to account for the colonies formed by surviving bacteria.Results. Amin and Vamin 14 had a weak immunostimulating effect on the respiratory burst of neutrophils activated by PMA. The addition of amino acid compositions to neutrophils, which phagocytized bacteria at a neutrophil to bacteria ratio of 1 : 10, led to an increase in the number of colonies formed by the surviving bacteria. With a neutrophil/bacteria ratio of 10 : 1, the studied amino acid compositions enhanced the bactericidal activity of neutrophils, causing a decrease in the number of surviving bacteria forming colonies. Direct addition of amino acid compositions enhanced colony formation by bacteria.Conclusions. The amino acid compositions Amin and Vamin 14 enhance the growth of bacteria, including those phagocytized by neutrophils at a cell to bacteria ratio of 1 : 10 but stimulate the bactericidal activity of neutrophils that phagocytized S. aureus at a cell to bacteria ratio of 10 : 1 as well as in the respiratory burst induction test in neutrophils activated by PMA.

The review provides an analysis of the literature data on the use of various modern molecular genetic methods for the indication and identification of Yersinia pestis strains with different properties and degrees of virulence, which is due to the diverse natural conditions in which they circulate. The methods are also considered from the perspective of their application at three levels of organizations forming the system of laboratory diagnostics of infectious diseases of the Russian Federation (territorial, regional, and federal) to solve the problem of maintaining the sanitary and epidemiological well-being of the country’s population. The main conditional groups of methods are considered: based on the analysis of the lengths of restriction fragments (ribo- and IS-typing, pulse gel electrophoresis); based on the analysis of specific fragments (DFR typing, VNTR typing); based on sequencing (MLST, CRISPR analysis, SNP analysis); PCR methods (including IPCR, SPA); isothermal amplification methods (LAMP, HDA, RPA, SEA, PCA, SHERLOCK); DNA microarray; methods using aptamer technology; bio- and nanosensors; DNA origami; and methods based on neural networks. As a result of the analysis, it can be concluded that there is rapid development of molecular diagnostics and genetics, which is aimed at increasing efficiency, multifactority, and simplification of application with no need for expensive equipment and highly qualified personnel for analysis. At all levels of the organizations forming the system of laboratory diagnostics of infectious diseases of the Russian Federation, it is possible to use methods based on PCR, isothermal amplification, SHERLOCK, biosensors, and small-sized sequencing devices. At the territorial level, at antiplague stations, the use of immuno-PCR and SPA for the indication of Y. pestis is promising. At the regional level, the introduction of technologies based on the use of aptamers and DNA microarray looks promising. At the federal level, the use of DNA origami methods and new technologies of whole genome sequencing is promising in the framework of advanced identification, molecular typing, and sequencing of the genomes of plague pathogen strains.

Fructan-modifying enzymes are divided into fructan-producing enzymes (fructosyl transferases) and fructan hydrolyzing enzymes (invertases, inulinases, levanases). Fructosyl transferases break the glycosidic bond of sucrose and use the energy of this bond to attach the resulting fructosyl to another sucrose molecule or other acceptor, increasing the fructan chain. Invertases hydrolyze sucrose and small fructooligosaccharides. Oligo- and polyfructans are cleaved by inulinases and levanases. A difference of only three amino acid residues affects the ability of glycoside hydrolases to cleave various substrates, in particular inulin and levan, or to exhibit transfructosylating activity. In this regard, the aim of the work was to carry out a comparative analysis of the primary structures of glycoside hydrolases of various origins. The paper presents the results of a comparative analysis of the amino acid sequences of glycoside hydrolases from the NCBI database (https://www.ncbi.nlm.nih.gov/). The overlap percentage (Query cover) of the sequences and their identity (Ident) were calculated using the Blast program (https://blast.ncbi.nlm.nih.gov/Blast.cgi). It was found that the affinity of endoinulinase from Aspergillus ficuum with 6- and 1-fructan exohydrolases from Arabidopsis thaliana and Arabidopsis lyrata subsp. Lyrata was higher (89% overlap and 24% identity) than exoinulinase from Kluyveromyces marxianus (38 and 57% overlap, 29 and 26% identity, respectively). Fructan 1-exohydrolase I from Cichorium intybus was also closer in primary structure to fungal endoinulinase (90% overlap and 25% identity) than to yeast exoinulinase (51% overlap and 27% identity). From the results obtained, the following conclusion can be drawn: the mechanism of substrate hydrolysis does not in all cases determine the degree of homology of glycoside hydrolases and related enzymes. It is possible that some glycoside hydrolases, including inulinases, can act both as endo and exo-enzymes, i.e., possess both types of catalytic activity towards fructans.

We studied the interaction of the antitumor agent abiraterone and its pharmacologically active metabolite D4A, which is promising for use as an agent for the treatment of prostate cancer, with cytochrome P450 2C9 (CYP2C9). Using the absorption spectroscopy, it has been shown that both compounds under study cause spectral changes of CYP2C9, indicating the interaction of the nitrogen atom of the pyridine ring of the ligand with the heme iron ion of the active site of the enzyme. However, the ligand–enzyme interaction, which is mediated by water bound to the heme iron ion, is possible. Based on the spectral changes, the values of dissociation constants (K_S) of the complexes of abiraterone and D4A with CYP2C9 were determined, which amounted to 1.73 ± 0.14 µM and 3.95 ± 0.16 µM, respectively. Both compounds inhibited the O-demethylase activity of CYP2C9 toward the substrate of this enzyme, naproxen. At a naproxen concentration of 100 µM, the concentrations of abiraterone, D4A, and sulfaphenazole, which inhibit CYP2C9 activity by 50% (IC₅₀), were determined as 13.9 µM, 40 µM, and 41 µM, respectively. The data obtained can be used to predict drug-drug interactions at the CYP2C9 level when using abiraterone or D4A as an antitumor agent for the treatment of prostate cancer in complex pharmacotherapy.

The analysis of cytochrome P450 transcripts was carried out by the nanopore sequencing in liver tissue samples of three donors and HepG2 line cells. It was demonstrated that direct mRNA sequencing with a MinION nanopore sequencer (Oxford Nanopore Technologies) makes it possible to obtain quantitative profiles for transcripts (and their splice variants) of cytochrome P450 superfamily genes encoding isoforms involved in metabolism of the majority (~80%) of drugs. The splice variant profiles substantially differ for donors. The cytochrome P450 gene expression at the transcript level is significantly weaker in cells of the HepG2 line compared with that in the normal liver tissue. This limits the capability of the direct mRNA nanopore sequencing for studying alternative splicing of cytochrome P450 transcripts in HepG2 cells. Both quantitative and qualitative profiles of the cytochrome P450 gene expression at the transcript level notably differ in human liver tissue and HepG2 cells.