Showing papers in "Acta Naturae in 2009"
TL;DR: Data on the neuraminidase substrate specificity and different approaches in studying the activity of this enzyme are summarized, along with considerations on the mechanisms of resistance of modern influenza viruses to those antivirals.
Abstract: The structure of the influenza virus neuraminidases, the spatial organization of their active site, the mechanism of carbohydrate chains desialylation by neuraminidase, and its role in the influenza virus function at different stages of the viral infectious cycle are considered in this review. Data on the neuraminidase substrate specificity and different approaches in studying the activity of this enzyme are summarized. In addition, data on neuraminidase inhibitors (as antivirals) are provided, along with considerations on the mechanisms of resistance of modern influenza viruses to those antivirals.
159 citations
TL;DR: The main prosthetic groups and their role in the interaction of these groups and the apoenzyme in the process of catalysis are described, using pyridoxal catalysis as an example.
Abstract: This paper reviews the chemical and functional aspects of the posttranslational modifications of proteins, which are achieved by the addition of various groups to the side chain of the amino acid residue backbone of proteins. It describes the main prosthetic groups and the interaction of these groups and the apoenzyme in the process of catalysis, using pyridoxal catalysis as an example. Much attention is paid to the role of posttranslational modification of proteins in the regulation of biochemical processes in live organisms, and especially to the role of protein kinases and their respective phosphotases. Methylation and acetylation reactions and their role in the “histone code,” which regulates genome expression on the transcription level, are also reviewed. This paper also describes the modification of proteins by large hydrophobic residues and their role in the function of membrane-associated proteins. Much attention is paid to the glycosylation of proteins, which leads to the formation of glycoproteins. We also describe the main non-enzymatic protein modifications such as glycation, homocysteination, and desamidation of amide residues in dibasic acids.
92 citations
TL;DR: This review focuses on up-to-date methods of antibody engineering for diagnosing and treating various diseases, particularly on new technologies meant to refine the effector functions of therapeutic antibodies.
Abstract: The modular structure and versatility of antibodies enables one to modify natural immunoglobulins in different ways for various clinical applications. Rational design and molecular engineering make it possible to directionally modify the molecular size, affinity, specificity, and immunogenicity and effector functions of an antibody, as well as to combine them with other functional agents. This review focuses on up-to-date methods of antibody engineering for diagnosing and treating various diseases, particularly on new technologies meant to refine the effector functions of therapeutic antibodies.
66 citations
Journal Article•
TL;DR: In this study for the first time induced pluripotent cells were derived from human umbilical vein endothelial cells by genetic reprogramming and it was showed that these cells are similar to embryonic stem cells in their morphology, function, and molecular level.
Abstract: Cell biology is one of the most rapidly developing branches in modern biology. The most interesting stages in early embryonic development for cell biology are those when a large number of cells are pluripotent. Inner-cell mass of blastocyst can be cultivated in vitro, and these cells are called embryonic stem cells. They are able to differentiate into different types of cells and tissues. But the greatest interest for practical application is the return (reprogramming) of adult cells into the pluripotent state. In our study for the first time induced pluripotent cells were derived from human umbilical vein endothelial cells by genetic reprogramming. We showed that these cells are similar to embryonic stem cells in their morphology, function, and molecular level. We are the first to show that reprogramming sufficiently changes X-chromosome chromatin state, which is normally inactive in female endothelial cells, towards its activation, providing evidence that endothelial cells are reprogrammed at an epigenetic level.
47 citations
TL;DR: Non-human enzymes, phosphotriesterases and oxidases from various bacterial and eukaryotic sources could be used for external use against OP poisoning and for internal use after modifications for immunological compatibility.
Abstract: Bioscavengers are biopharmaceuticals that specifically react with toxicants thus, enzymes reacting with poisonous esters can be used as bioscavengers for neutralization of toxic molecules before they reach physiological targets Parenteral administration of bioscavengers is, therefore, intended for prophylaxis or pre-treatments, emergency and post-exposure treatments of intoxications these enzymes can also be used for application on skin, mucosa and wounds as active components of topical skin protectants and decontamination solutions Human butyrylcholinesterase is the first stoichiometric bioscavenger for safe and efficient prophylaxis of organophosphate poisoning However, huge amounts of a costly enzyme are needed for protection thus, the bioscavenger approach will be greatly improved by the use of catalytic bioscavengers catalytic bioscavengers are enzymes capable of degrading toxic esters with a turnover Suitable catalytic bioscavengers are engineered mutants of human enzymes efficient mutants of human butyrylcholinesterase have been made that hydrolyze cocaine at a high rate Mutants of human cholinesterases capable of hydrolyzing OPs have been made, but so far their activity is too low to be of medical interest Human paraoxonase a promiscuous plasma enzyme is certainly the most promising phosphotriesterase However, its biotechnology is still in its infancy Other enzymes and proteins from blood and organs, and secondary biological targets of OPs and carbamates are potential bioscavengers, in particular serum albumin that reacts with OPs and self-reactivates Lastly, non-human enzymes, phosphotriesterases and oxidases from various bacterial and eukaryotic sources could be used for external use against OP poisoning and for internal use after modifications for immunological compatibility
45 citations
TL;DR: This review is an introduction to phage display methodology, which presents recombinant antibodies display in more details, construction of phage libraries of antibody fragments and different strategies for the biopanning procedure.
Abstract: The display of peptides and proteins on the surface of filamentous bacteriophage is a powerful methodology for selection of peptides and protein domains, including antibodies. An advantage of this methodology is the direct physical link between the phenotype and the genotype, as an analyzed polypeptide and its encoding DNA fragment exist in one phage particle. Development of phage display antibody libraries provides repertoires of phage particles exposing antibody fragments of great diversity. The biopanning procedure facilitates selection of antibodies with high affinity and specificity for almost any target. This review is an introduction to phage display methodology. It presents recombinant antibodies display in more details:, construction of phage libraries of antibody fragments and different strategies for the biopanning procedure.
40 citations
TL;DR: Tests showed that the genotoxic effects of the SNP solution are associated with the presence of AOT rather than SNP, and both were significantly higher than those found with AgNO3 and in control mice.
Abstract: The toxic and genotoxic effects of silver nanoparticles were studied on injected mice (BALB/c line) in vivo. A water solution of silver nanoparticles (SNP) with particle sizes of 9±6 nm was obtained by means of the original method of biochemical synthesis. The effect of the SNP solution was compared to those of AOT (anionic surfactant used as SNP stabilizer) and silver nitrate (i.e. Ag+ ions) introduced as water solutions. In studies of the toxic effects, the death of mice was registered 12-24 hours after injection only at two maximum dozes of SNP (equivalent to 0.54 and 0.36 gAg/l). It is shown that the toxic effect decreases in the sequence SNP>AOT>>AgNO3. The LE50/30 values for SNP and AOT are equal to 0.30±0.07 gAg/l and 13.3±2.1 gAg/l, respectively. Genotoxic effects were assessed by the abnormal sperm heads test and neutral Comet assay. The frequencies of abnormal sperm heads (ASHs) did not differ after treatment by SNP and AOT, but both were significantly higher than those found with AgNO3 and in control mice. Comet assay showed an increase of the DNA percentage in the comet tail in spleen cells after the injection of SNP and AOT in concentrations of ≤ LE50/30. Tail DNA % was 32.8±1.3 and 26.3±1.7%, respectively, vs 16.2±0.7% for the untreated control. To sum up, these tests showed that the genotoxic effects of the SNP solution are associated with the presence of AOT rather than SNP.
39 citations
TL;DR: The investigation of marine natural products (low molecular weight bioregulators) is a rapidly developing scientific field at the intersection of biology and chemistry that has led to the discovery of 20,000 new substances.
Abstract: The investigation of marine natural products (low molecular weight bioregulators) is a rapidly developing scientific field at the intersection of biology and chemistry. Investigations aimed at detecting, identifying, and understanding the structure of marine natural products have led to the discovery of 20,000 new substances, including those characterized by an extremely high physiological activity. Some results and prospects of works aimed at creating new drugs on the basis of marine natural products are discussed herein.
30 citations
TL;DR: Putative stem/progenitor cells from AF are capable of long-term proliferation in vitro and the profile of gene expression led us to speculate that they have greater differentiation potential than mesenchymal stem cells and may be useful for cell therapy.
Abstract: Stem cells capable of long-term proliferation and differentiation into different cell types may be a promising source of cells for regenerative medicine. Recently, much attention has been paid to fetal stem cells, among which are cells from amniotic fluid (AF). We have isolated amniotic stem cells from 3 AF samples. Flow cytometry, RT -PCR and immunohistochemistry have shown that these cells express mesenchymal (CD90, CD73, CD105, CD13, CD29, CD44, and CD146), neural (≤3-tubulin, Nestin, and Pax6), epithelial (keratin 19 and p63) markers and also markers of pluripotency (Oct4, Nanog, and Rex-1). Transplantation of the cells to nude mice does not lead to tumor formation. Thus, putative stem/progenitor cells from AF are capable of long-term proliferation in vitro and the profile of gene expression led us to speculate that they have greater differentiation potential than mesenchymal stem cells and may be useful for cell therapy.
25 citations
TL;DR: The gene expression profile under specific conditions during reversible transition of M. tuberculosis cells to the "non-culturable" (NC) state in a prolonged stationary phase concludes that NC cell formation is an active process involved in the regulation of many genes of different pathways.
Abstract: We analyzed the gene expression profile under specific conditions during reversible transition of M. tuberculosis cells to the "non-culturable" (NC) state in a prolonged stationary phase. More than 500 genes were differentially regulated, while 238 genes were upregulated over all time points during NC cell formation. Approximately a quarter of these upregulated genes belong to insertion and phage sequences indicating a possible high intensity of genome modification processes taking place under transition to the NC state. Besides the high proportion of hypothetical/conserved hypothetical genes in the cohort of upregulated genes, there was a significant number of genes belonging to intermediary metabolism, respiration, information pathways, cell wall and cell processes, and genes encoding regulatory proteins. We conclude that NC cell formation is an active process involved in the regulation of many genes of different pathways. A more detailed analysis of the experimental data will help to understand the precise molecular mechanisms of dormancy/latency/persistence of M. tuberculosis in the future. The list of upregulated genes obtained in this study includes many genes found to be upregulated in other models of M. tuberculosis persistence. Thirteen upregulated genes, which are common for different models, can be considered as potential targets for the development of new anti-tuberculosis drugs directed mainly against latent tuberculosis.
23 citations
TL;DR: This review presents in this review the most interesting results of the molecular modeling of the structural parameters and optical and vibrational spectra of the chromophorecontaining domains of fluorescent proteins by methods of quantum chemistry, molecular dynamics, and combined quantum-mechanical-molecular-MEchanical approaches.
Abstract: Fluorescent proteins from the family of green fluorescent proteins are intensively used as biomarkers in living systems. The chromophore group based on the hydroxybenzylidene-imidazoline molecule, which is formed in nature from three amino-acid residues inside the protein globule and well shielded from external media, is responsible for light absorption and fluorescence. Along with the intense experimental studies of the properties of fluorescent proteins and their chromophores by biochemical, X-ray, and spectroscopic tools, in recent years, computer modeling has been used to characterize their properties and spectra. We present in this review the most interesting results of the molecular modeling of the structural parameters and optical and vibrational spectra of the chromophorecontaining domains of fluorescent proteins by methods of quantum chemistry, molecular dynamics, and combined quantum-mechanical-molecular-mechanical approaches. The main emphasis is on the correlation of theoretical and experimental data and on the predictive power of modeling, which may be useful for creating new, efficient biomarkers.
TL;DR: In the atomic resolution structure, the covalent bonds in the nicotinamide group are somewhat changed in agreitalicent with the results of quantum mechanical calculations, providing evidence that the cofactor acquires a bipolar form in the transition state of the enzymatic reaction.
Abstract: The crystal structure of the ternary complex of NAD+-dependent formate dehydrogenase from the methylotrophic bacterium Moraxella sp. C-1 with the cofactor (NAD+) and the inhibitor (azide ion) was established at 1.1 A resolution. The complex mimics the structure of the transition state of the enzymatic reaction. The structure was refined with anisotropic displacitalicents parameters for non-hydrogen atoms to a R factor of 13.4%. Most of the nitrogen, oxygen, and carbon atoms were distinguished based on the analysis of the titalicperature factors and electron density peaks, with the result that side-chain rotamers of histidine residues and most of asparagine and glutamine residues were unambiguously determined. A comparative analysis of the structure of the ternary complex determined at the atomic resolution and the structure of this complex at 1.95 A resolution was performed. In the atomic resolution structure, the covalent bonds in the nicotinamide group are somewhat changed in agreitalicent with the results of quantum mechanical calculations, providing evidence that the cofactor acquires a bipolar form in the transition state of the enzymatic reaction.
TL;DR: Stacking interactions are studied, which usually are not properly taken into account in widely used scoring functions and may include either components of molecular mechanics force fields or empirical terms, e.g. hydrogen bonds described by their geometrical parameters.
Abstract: The biological function of proteins is closely connected to interactions with their ligands and substrates. Proteins acting as receptors and enzymes bind these small molecules. Knowledge of the molecular mechanisms of protein-ligand interactions, particularly in the spatial structure of the protein-ligand complex, is a prerequisite for understanding the structure-functional properties of proteins and their role in biochemical pathways in the living cell. The availability of such a structure serves as a basis for rational drug design projects and greatly assists the search for new inhibitors (the ligands of certain protein-targets in an organism).
Experimental tools for determining the spatial structure of proteins and their complexes with ligands (such as X-ray crystallography or NMR spectroscopy) have particular limitations. Even if the structure of a protein is available, determining the structure of its complex with ligands may be experimentally demanding. Problems with purification and crystallization become especially difficult in studies of transmembrane proteins, which include a biologically important class of G-protein coupled receptors. However, recent successes in determining the structure of beta-adrenergic and adenosine receptors [1] are cause for optimism.
The technical difficulties restraining experimental methods stimulated computational molecular modeling. One of them (molecular docking) is a method aimed at predicting the spatial structure of a protein-ligand complex by docking a ligand molecule into the known atomic-resolution structure of a protein-binding site and estimating the reliability of the results. Nowadays, molecular docking has become an integral part of both fundamental studies aimed at understanding the structure-functional role of protein amino acids and applied drug-design programs [2, 3].
Docking approaches are further improved by implementing new algorithms of the conformational search and new scoring functions (methods to estimate the free energy of ligand binding). Scoring functions may include either components of molecular mechanics force fields [2] or empirical terms, e.g. hydrogen bonds described by their geometrical parameters [4]. In this work we studied stacking interactions, which usually are not properly taken into account in widely used scoring functions.
TL;DR: In the present paper, syntropy/dystropy is defined as a natural generic nonrandom phenomenon with an evolutionary-genetic basis that can be used in order to identify syntropic genes, as will be shown as examples in an analysis of cardiovascular diseases.
Abstract: In clinical medicine, the phenomenon of polypathy, as a particular object of investigation, was first put forth by French clinicians at the end of the 19 th century through the "arthritismus" doctrine. In the first half of the 20 th century, German paediatricians singled out "syntropias," which are combinations of diseases with common pathophysiological mechanisms, and "dystropias," which are diseases that rarely co-occur in one individual. In the present paper, syntropy/dystropy is defined as a natural generic nonrandom phenomenon with an evolutionary-genetic basis. t he genes involved in the development of syntropy are called "syntropic genes," whereas the genes that co-participate in pathophysiological mechanisms and prevent the co-occurrence of particular phenotypes are called "dystropic genes." Prospects for studying the genetic basis of this phenomenon are highlighted. the publicly available database HuGenet can be used in order to identify syntropic genes, as will be shown as examples in an analysis of c ardiovascular diseases.
TL;DR: The whole genome sequencing of a human (Russian man) was performed using two technologies currently present on the market - Sequencing by Oligonucleotide Ligation and Detection (SOLiD™) and sequencing technologies of molecular clusters using fluorescently labeled precursors (Illumina).
Abstract: At present, the new technologies of DNA sequencing are rapidly developing allowing quick and efficient characterisation of organisms at the level of the genome structure. In this study, the whole genome sequencing of a human (Russian man) was performed using two technologies currently present on the market - Sequencing by Oligonucleotide Ligation and Detection (SOLiD™) (Applied Biosystems) and sequencing technologies of molecular clusters using fluorescently labeled precursors (Illumina). The total number of generated data resulted in 108.3 billion base pairs (60.2 billion from Illumina technology and 48.1 billion from SOLiD technology). Statistics performed on reads generated by GAII and SOLiD showed that they covered 75% and 96% of the genome respectively. Short polymorphic regions were detected with comparable accuracy however, the absolute amount of them revealed by SOLiD was several times less than by GAII. Optimal algorithm for using the latest methods of sequencing was established for the analysis of individual human genomes. The study is the first Russian effort towards whole human genome sequencing.
TL;DR: The concept of a genetic pass as an individual DNA databank reflecting inherited human predisposition to different complex and monogenic disorders, with special emphasis on its present state, and the numerous difficulties related to the practical implementation of personalized medicine are outlined.
Abstract: The review is devoted to the impact of human genome research on progress in modern medicine. Basic achievements in genome research have resulted in the deciphering of the human genome and creation of a molecular landmarks map of the human haploid genome (HapMap Project), which has made a tremendous contribution to our understanding of common genetic and multifactorial (complex) disorders. Current genome studies mainly focus on genetic testing and gene association studies of multifactorial (complex) diseases, with the purpose of their efficient diagnostics and prevention . Identification of candidate ("predisposition") genes participating in the functional genetic modules underlying each common disorder and the use of this genetic background to elaborate sophisticated measures to efficiently prevent them constitutes a major goal in personalized molecular medicine. The concept of a genetic pass as an individual DNA databank reflecting inherited human predisposition to different complex and monogenic disorders, with special emphasis on its present state, and the numerous difficulties related to the practical implementation of personalized medicine are outlined. The problems related to the uncertainness of the results of genetic testing could be overcome at least partly by means of new technological achievements in genome research methods, such as genome-wide association studies (GWAS), massive parallel DNA sequencing, and genetic and epigenetic profiling. The basic tasks of genomic today could be determined as the need to properly estimate the clinical value of genetic testing and its applicability in clinical practice. Feasible ways towards the gradual implementation of personal genetic data, in line with routine laboratory tests, for the benefit of clinical practice are discussed.
TL;DR: The last three years were especially productive in the development of new methods of DNA purification and analysis, and a new evolutionary hypothesis and reconstruction of the biota history have been re-created from recent ancient DNA data.
Abstract: Studies of ancient DNA specimens started 25 years ago. At that time short mitochondrial DNA (mtDNA) fragments were the main targets in ancient DNA studies. The last three years were especially productive in the development of new methods of DNA purification and analysis. Complete mtDNA molecules and relatively large fragments of nuclear DNA are the targets of ancient DNA studies today. Ancient DNA studies allowed us to study organisms that went extinct more than ten thousand years ago, to reconstruct their phenotypic traits and evolution. Ancient DNA analyses can help understand the development of ancient human populations and how they migrated. A new evolutionary hypothesis and reconstruction of the biota history have been re-created from recent ancient DNA data. Some peculiarities and problems specific to the study of ancient DNA were revealed, such as very limited amounts of DNA available for study, the short length of the DNA fragments, breaks and chemical modifications in DNA molecules that result in “postmortem” mutations or complete blockage of DNA replication in vitro. The same specific features of DNA analysis were revealed for specimens from complicated forensic cases that result in the lack of experimental data or interpretation problems.. Here, we list the specific features of ancient DNA methodology and describe some achievements in fundamental and applied research of ancient DNA, including our own work in the field.
TL;DR: Psoriasis was used as a model to analyze the pathogenetic pathways of immune-mediated inflammatory diseases, and the results of bioinformatic, molecular-genetic and proteomic studies are provided.
Abstract: Psoriasis was used as a model to analyze the pathogenetic pathways of immune-mediated inflammatory diseases, and the results of bioinformatic, molecular-genetic and proteomic studies are provided. Cell mechanisms, common for the pathogenesis of psoriasis, as well as Crohn's disease, are identified. New approaches for immune-mediated diseases are discussed.
TL;DR: A technological scheme is developed which is suitable for the study of membrane localization in E.coli cells of recombinant proteins containing voltage-gated eukaryotic potassium channels as the functional active site and involves both biochemical and fluorescent methods for detecting target proteins in the cytoplasmic membrane.
Abstract: The effective expression of recombinant membrane proteins in E.coli depends upon the targeting and insertion of proteins into the cellular membrane, as well as on those proteins adopting the correct spatial structure. A significant technological problem involves the design of approaches for detecting the location of target proteins within a host cell. Using a hybrid potassium channel KcsA-Kv1.3 as a model, we developed a technological scheme which is suitable for the study of membrane localization in E.coli cells of recombinant proteins containing voltage-gated eukaryotic potassium channels as the functional active site. The scheme involves both biochemical and fluorescent methods for detecting target proteins in the cytoplasmic membrane of E.coli, as well as the study of the ligand-binding activity of membrane-embedded proteins.
TL;DR: The reliable slowing-down of the embriod-body formation by the embryonic SC transfected by theGDNF gene has been shown and no significant influence of the expression of the GDNF gene on the morphology and the proliferative activity of the transfecting embryonic SCs has been found when compared with the control ones.
Abstract: In 1961 Hayflick and Moorhead showed that a somatic cell culture has a limited life span [1]. In 1973 Olovnikov suggested that shortening the chromosomal ends (telomeres) determines the potential number of cell divisions. [2]. Telomeres protect the cellular genome from degradation; they participate in the chromosomal pairing during meiosis and in the gene expression regulation in the telomeres region [3]. In immortal cells that can divide infinitely, this should be the mechanism for compensating the chromosomal shortening. In 1975 Blackburn and Greider discovered the enzyme telomerase that elongates chromosomes [4].
Telomerase is a ribonucleoprotein complex that consists of components that are absolutely required for its activity: the RNA molecule and Telomerase Reverse Transcriptase TERT [5]; also, optionally several telomerase-associated proteins could be included in the telomerase complex. TR is also a template for TERT when telomerase elongates telomeres. Telomerase exists in human cells as dimers and contains two subunits of reverse transcriptase and two RNA molecules [6]. In human telomerase, p23/p90-shaperone, which is responsible for the complex assembling/configuration, binds 14-3-3, which is responsible for nuclear localization, and TP1 with an unknown function. Proteins hGAR1, Dyskerin/NAP57, hNHP2, and C1/C2, which are responsible for the stability, maturation, and localisation of RNA, bind to the hTR; La and hStau, which are supposedly responsible for the binding to telomeres; L22, which acts in processing and nuclear localisation; and hNOP10, A1/UP1, and TP1 with an unknown function [7]; TCAB1, which is responsible for the localisation of hTR in Cajal bodies and binding with telomers [8]. The enzymatic activity of human telomerase in the rabbit reticulocytes lysate is detected by adding hTR and hTERT [9, 10]. Note that telomerase functioning in vivo is not always consistent with the telomerase activity that was measured in vitro. For example, adding the Hemagglutinin epitope to the C-end of hTERT stops telomere alongation but does not suppress telomerase activity [11].
Telomerase activity detected in vitro appears in leucocytes in the G1 phase (Fig. 1) [12]. On the other hand, telomeres are replicated in vivo during the S-phase (Fig. 1) [13, 14]. During most of the cell cycle, TCAB1 helps the hTR accumulation in Cajal bodies [8], and in S-phase it is combined with telomeres in a cell. During the S-phase of the cell cycle, hTERT also moves to telomeres [15, 16]. This means that there is regulation at the level of the spatial localisation of active telomerase (enzyme) and telomere (substrate). A correlation between the telomerase activity and the length of telomeres is not always possible to obtain. For example, there is no dependency between the length of telomeres and telomerase activity in leukaemia [17].
Fig. 1.
Cell cycle scheme. The appearance of telomerase activity in vitro happens in the G1-phase, but it works at the S-phase.
The two-step hypothesis of cell aging and immortalisation M1/M2 theory describes the activity of telomerase dependency on the number of cell divisions very well (Fig. 2). In embryonic cell lines, telomerase is active and the length of telomeres is constant. In stem cells, the activity of polymerase is lower and it only partially compensates the telomeres shortening. In somatic cells, telomerase in not active. The shortening of telomeres leads to the moment of M1, i.e., the achievement of the Hayflick limit (point A on Fig 1) and the transition of cells to the senescence (aging) condition, which could be rescued by the inactivation or deletion of pRB/p16 or p53. Cells that pass through M1 continue their cell division and achieve the condition of crisis M2 (point B on Fig 2), which leads to massive cellular death. Cells that survive begin their transformation into cancer cells. Cancer cells have the ability for unlimited cell division and support the length of telomeres (usually due to telomerase activity). In the case of the transfection of hTR-expressing somatic cells by the hTERT gene (point C on Fig. 2) before the moment of M2, they, similarly to cancer cells, demonstrate elongation and stabilization of telomeres [7].
Fig. 2.
Dependence of telomeres length on the cell cycle number in different types of cells: embryonic cell lines, somatic cells, and hTERT-transfected cells. (A) achieving the Hayflick limit by cells, (B) crisis with the following cell death and transformation ...
Telomerase activity, with rare exceptions, does not occur in human somatic cells and tissues. Its activity was shown in reproductive tissues, as well as in intensively renewing tissues, such as some types of blood cells, the intestinal epithelium, and the layer of skin cells [18]; however, the level of telomerase activity in the somatic cells with active polymerase is lower than in cancer cells [19].
TL;DR: It was demonstrated that MBP (non-ubiquitinated) is a good substrate for 20S and for the 26S proteasome, and substantial differences in the degradation pattern of this neuroantigen were found, which could indicate the better presentation MBP parts on MHC molecules in the case of mice predisposed to the development of experimental autoimmune encephalomyelitis.
Abstract: The proteasome is a high molecular protein complex whose purpose is specific protein degradation in eukaryotic cells. One of the proteasome functions is to produce peptides, which will then be presented on the outer cell membrane using main histocompatibility complex (MHC) molecules of the first or second class. There are definite reasons to believe that proteasome directly takes part in the specific degradation of myelin basic protein (MBP), which make up to 30% of all proteins in the myelin sheath of neuronal axons. The details of the proteasomal degradation of MBP are still unclear. In this work, the features of specific MBP degradation by proteasome were studied.It was demonstrated that MBP (non-ubiquitinated) is a good substrate for 20S and for the 26S proteasome. This is the first work on detecting the sites of MBP proteolysis by proteasome from brains of SJL/J/J and Balb/C mice's lines. Substantial differences in the degradation pattern of this neuroantigen were found, which could indicate the better presentation MBP parts on MHC molecules in the case of mice predisposed to the development of experimental autoimmune encephalomyelitis.
TL;DR: It is shown that the gene-targeted oligonucleotides show considerable variations in their effect on tumor tissue, depending on the target gene in question, and not all the antisense drugs targeting the same region of the mRNA of the target-gene were equally effective.
Abstract: Currently, the main way to fight cancer is still chemotherapy. This method of treatment is at the height of its capacity, so, setting aside the need for further improvements in traditional treatments for neoplasia, it is vital to develop now approaches toward treating malignant tumors. This paper reviews innovational experimental approaches to treating malignant malformations based on the use of gene-targeted drugs, such as antisense oligonucleotides (asON), small interfering RNA (siRNA), ribozymes, and DNAzymes, which can all inhibit oncogene expression. The target genes for these drugs are thoroughly characterized, and the main results from pre-clinical and first-step clinical trials of these drugs are presented. It is shown that the gene-targeted oligonucleotides show considerable variations in their effect on tumor tissue, depending on the target gene in question. The effects range from slowing and stopping the proliferation of tumor cells to suppressing their invasive capabilities. Despite their similarity, not all the antisense drugs targeting the same region of the mRNA of the target-gene were equally effective. The result is determined by the combination of the drug type used and the region of the target-gene mRNA that it complements.
TL;DR: It is suggested that the variations in the pub gene expression in the embryonic stem cells influence significantly the mesodermal and ectodermal differentiation of these cells.
Abstract: The influence of low and high pub gene expression on the initial stages of the differentiation of mouse embryonic stem cells into derivatives of ecto-, meso-, and endoderm in vitro was investigated. As follows from the results of a RT -PCR analysis, the expression of the vimentin, somatostatin, GATA 4, and GATA 6 genes, being the markers of endodermal differentiation, does not vary in both the cells with high pub gene expression and the cells with low pub gene expression, as well as in the corresponding control lines. The cells with high pub gene expression are characterized by an increase in the expression of mesodermal differentiation gene-markers (trI card, trI skel, c-kit, and IL-7), whereas the cells with low pub gene expression are specified by a decrease in their expression. According to the analyses carried out, the reverse is characteristic of the expression of ectodermal differentiation gene-markers (nestin, ≤-III tubulin, gfap, and th). Expression of these genes decreases in cell lines with high pub gene expression, whereas their expression increases with the decrease in pub gene expression. Hence, it is suggested that the variations in the pub gene expression in the embryonic stem cells influence significantly the mesodermal and ectodermal differentiation of these cells.
TL;DR: The catalytic properties of human foamy virus integrase, which belongs to the Retroviridae family, were studied and a considerable similarity in the activity of the two enzymes was observed.
Abstract: Due to their ability to integrate into the host cell's genome, retroviruses represent an optimal basis for the creation of gene therapy vectors. The integration reaction is carried out by a viral enzyme integrase: thus, a detailed research of this enzyme is required. In this work, the catalytic properties of human foamy virus integrase were studied. This virus belongs to the Retroviridae family. The dissociation constant was determined, together with the kinetics of integrase catalytic activity. The data obtained were compared to those for the human immunodeficiency virus integrase and a considerable similarity in the activity of the two enzymes was observed.
TL;DR: Data indicate GDP-D-Arap transport in vivo by the LPG2 multispecific nucleotide sugar transporter into the Leishmania Golgi apparatus, in which it can be used by glycosyltransferases as a donor substrate for Glycosylation.
Abstract: In this work we describe an unusual enzyme from Leishmania major (Arabinokinase/Pyrophosphorylase) that catalyzes the synthesis of GDP-D-arabinopyranose (GDP-D-Arap) via a D-arabinose-1-phosphate intermediate in the presence of ATP and GTP. Our data indicate GDP-D-Arap transport in vivo by the LPG2 multispecific nucleotide sugar transporter into the Leishmania Golgi apparatus, in which it can be used by glycosyltransferases as a donor substrate for glycosylation.
TL;DR: The synthetic capacity of penicillin acylase with 6-aminopenicillanic acid as an external nucleophile was especially sensitive to mutation of the β71 residue in contrast to the synthesis with 7-aminodeacetoxycephalosporanic acid.
Abstract: Residue phenylalanine 71 of the β-chain of penicillin acylase from E. coli is involved in substrate binding and chiral discrimination of its enantiomers. Different amino acid residues have been introduced at position βF71, and the mutants were studied with respect to their enantioselectivity and substrate specificity. Some mutants demonstrated remarkably improved catalytic activity. Moreover, mutation of βF71 residue allowed to enhance penicillin acylase enantioselectivity. The catalytic activity to the specific substrates was improved up to 36 times, most notably for K, R, and L mutants. Increased activity to a D-phenylglycine derivative - a valuable specificity improvement for biocatalytic synthesis of new penicillins and cephalosporins - was shown for βF71R and βF71L mutants. The synthetic capacity of penicillin acylase with 6-aminopenicillanic acid as an external nucleophile was especially sensitive to mutation of the β71 residue in contrast to the synthesis with 7-aminodeacetoxycephalosporanic acid.
TL;DR: In this study for the first time induced pluripotent cells were derived from human umbilical vein endothelial cells by genetic reprogramming, and it was showed that these cells are similar to embryonic stem cells in their morphology, function, and molecular level.
Abstract: Cell biology is one of the most rapidly developing branches in modern biology. The most interesting stages in early embryonic development for cell biology are those when a large number of cells are pluripotent. Inner-cell mass of blastocyst can be cultivated in vitro, and these cells are called embryonic stem cells. They are able to differentiate into different types of cells and tissues. But the greatest interest for practical application is the return (reprogramming) of adult cells into the pluripotent state. In our study for the first time induced pluripotent cells were derived from human umbilical vein endothelial cells by genetic reprogramming. We showed that these cells are similar to embryonic stem cells in their morphology, function, and molecular level. We are the first to show that reprogramming sufficiently changes X-chromosome chromatin state, which is normally inactive in female endothelial cells, towards its activation, providing evidence that endothelial cells are reprogrammed at an epigenetic level.
TL;DR: The suggested direct MALDI mass-spectrometry profiling strategy, coupled with special analysis software, seems promising for the species identification of N. gonorrhoeae but is assumed insufficient for H. pylori species determination.
Abstract: This study examines the features and limitations of direct Matrix-Assisted Laser Desorption-Ionisation (MALDI) mass-spectrometry profiling of bacterial cells for investigating a microbial population. The optimal laboratory protocol, including crude bacteria lyses by a solution of 50% acetonitrile, 2.5% trifluoroacetic acid, and using α-cyano-4-hydroxy cinnamic acid as the MALDI matrix, has been developed. Two different bacteria species were under investigation, and representative mass spectra from 278 strains of Neisseria gonorrhoeae and 22 strains of Helicobacter pylori have been analyzed. It's known that both bacteria demonstrate a variable degree of polymorphism. For N. gonorrhoeae, the MALDI mass spectra that was collected possessed about 70 peaks, 20 of which were good reproducible ones. In spite of the fact that three peaks were found with differing spectra in some strains, little diversity in the N. gonorrhoeae population was revealed. This fact indicates the prospects in using direct MALDI mass-spectrometry profiling for gonococcus identification. In the case of H. pylori strains, the variety in the collected mass-spectra was shown to be essential. Only five peaks were present in more than 70% of strains, and a single mass value was common for all spectra. While these data call into question the possibility of the reliable species identification of H. pylori using this approach, the intraspecies differentiation of strains was offered. Good association between MALDI profile distributions and the region of strain isolation have been found. Thus, the suggested direct MALDI mass-spectrometry profiling strategy, coupled with special analysis software, seems promising for the species identification of N. gonorrhoeae but is assumed insufficient for H. pylori species determination. At the same time, this would create a very good chance for an epidemiological study of such variable bacteria as H. pylori.
TL;DR: The results of comparison of inhibition of expression of oncogene AML-ET O by synthetic siRNA and by recombinant lentiviruses coding for corresponding shRNA are presented.
Abstract: RNA-interference is an effective natural mechanism of post-transcriptional modulation of gene expression. RNA-interference mechanism exist as in high eukaryotes both animals and plants as well in lower eukaryotes and viruses. RNA-interference is now used as a powerful tool in study of functional gene activity and many essential for fundamental biology results was obtained with this approach. Also it's widely believed that RNA-interference could be used in working out of new therapeutic medicine against malignant, infectious and hereditary diseases. One of the main problems of these developments is search of effective methods of siRNA transfer into the target cells. At present time for these purpose different sorts of transfect ions or viral transduction are used. At present article the results of comparison of inhibition of expression of oncogene AML-ET O by synthetic siRNA and by recombinant lentiviruses coding for corresponding shRNA are presented.