Your browser doesn't support javascript.
loading
Show: 20 | 50 | 100
Results 1 - 20 de 52
Filter
Add more filters










Publication year range
1.
Curr Top Microbiol Immunol ; 435: 1-31, 2022.
Article in English | MEDLINE | ID: mdl-30599050

ABSTRACT

Chikungunya virus (CHIKV) was discovered more than six decades ago, but has remained poorly investigated. However, after a recent outbreak of CHIK fever in both hemispheres and viral adaptation to new species of mosquitoes, it has attracted a lot of attention. The currently available experimental data suggest that molecular mechanisms of CHIKV replication in vertebrate and mosquito cells are similar to those of other New and Old World alphaviruses. However, this virus exhibits a number of unique characteristics that distinguish it from the other, better studied members of the alphavirus genus. This review is an attempt to summarize the data accumulated thus far regarding the molecular mechanisms of alphavirus RNA replication and interaction with host cells. Emphasis was placed on demonstrating the distinct features of CHIKV in utilizing host factors to build replication complexes and modify the intracellular environment for efficient viral replication and inhibition of the innate immune response. The available data suggest that our knowledge about alphavirus replication contains numerous gaps that potentially hamper the development of new therapeutic means against CHIKV and other pathogenic alphaviruses.


Subject(s)
Chikungunya Fever , Chikungunya virus , Viruses , Animals , Chikungunya virus/genetics , Virus Replication
2.
bioRxiv ; 2021 Jul 27.
Article in English | MEDLINE | ID: mdl-34341791

ABSTRACT

The ongoing world-wide Severe Acute Respiratory Syndrome coronavirus 2 (SARS-CoV-2) pandemic shows the need for new sensing and therapeutic means against the CoV viruses. The SARS-CoV-2 nsp1 protein is important, both for replication and pathogenesis, making it an attractive target for intervention. In recent years nanoparticles have been shown to interact with peptides, ranging in size from single amino acids up to proteins. These nanoparticles can be tailor-made with specific functions and properties including bioavailability. To the best of our knowledge, in this study we show for the first time that a tailored titanium oxide nanoparticle interacts specifically with a unique site of the full-length SARS-CoV-2 nsp1 protein. This can be developed potentially into a tool for selective control of viral protein functions.

3.
Mol Biol (Mosk) ; 51(2): 251-261, 2017.
Article in Russian | MEDLINE | ID: mdl-28537232

ABSTRACT

Recently, a number of new highly efficient antibody-based anticancer therapeutics have emerged. These receptor-binding antibodies have beneficial toxicity profiles associated with relatively mild side effects. Therefore, the search for novel surface proteins that are present on cancer cells and play important metabolic or defensive roles has intensified. Additionally, the therapeutic stimulation of patient's immune system in order to aim its components, specifically, phagocytes and cytotoxic T-lymphocytes, at tumor cells is gaining traction. This review is focused on the CD47 surface receptor, a ubiquitously expressed molecule, which could nevertheless serve as a therapeutic target due to its ability to simultaneously stimulate both natural and adaptive immune response.


Subject(s)
Antigens, Neoplasm/immunology , CD47 Antigen/immunology , Immunity, Cellular , Immunity, Innate , Neoplasms , Phagocytes/immunology , T-Lymphocytes, Cytotoxic/immunology , Animals , Humans , Neoplasms/immunology , Neoplasms/therapy
4.
Mol Biol (Mosk) ; 50(1): 69-79, 2016.
Article in Russian | MEDLINE | ID: mdl-27028812

ABSTRACT

High heterogeneity is characteristic of oncology diseases, often complicating the choice of optimal anticancer treatment. One cancer type may combine tumors differing in histogenesis, genetic lesions, and mechanism of cell transformation. Differences in the mechanism of cell malignant transformation result in specifics of cancer cell metabolism and sensitivity to various agents, including anticancer treatments. Hence, the molecular subtype of a tumor is essential to know for choosing the optimal therapeutic strategy. The review considers the role actin-associated proteins and tyrosine kinases, in particular, PDLIM4 and Src kinase, play in the formation of pathological signaling pathways.


Subject(s)
Breast Neoplasms/metabolism , Breast Neoplasms/pathology , DNA-Binding Proteins/metabolism , Disease Progression , LIM Domain Proteins/metabolism , Signal Transduction , src-Family Kinases/metabolism , Breast Neoplasms/enzymology , Humans , Phosphorylation
5.
Mol Biol (Mosk) ; 49(2): 264-78, 2015.
Article in Russian | MEDLINE | ID: mdl-26065254

ABSTRACT

Cancer therapeutics based on protein biomolecules that exhibit selective toxic of inhibiting effects towards tumor cells without affecting normal tissue, are gaining extensive attention in cancer research. This heterogenous group of proteins consists of several subgroups, among them, are engineered cancer antigen-specific antibodies that suppress tumor growth by blocking proliferation-inducing receptors, or by direct action of a covalently attached toxin. Another subgroup of anticancer proteins that also represents promising potential therapeutic agents is oncotoxic proteins that can selectively trigger proapoptotic signaling in cancer cells. The oncotoxic proteins target such commonly disturbed processes in tumor calls as enhanced cell proliferation, altered cell-cycle control, deficient apoptotic response, inhibited mitochondrial respiration and activated glycolysis. The introduction of oncotoxic proteins to the clinic might substantially widen and upgrade modern arsenal of anticancer therapeutics.


Subject(s)
Antibodies, Neoplasm/therapeutic use , Antibodies, Neutralizing/therapeutic use , Antineoplastic Agents/therapeutic use , Neoplasms , Animals , Antibodies, Neoplasm/genetics , Antibodies, Neutralizing/genetics , Apoptosis/drug effects , Cell Proliferation/drug effects , Humans , Neoplasms/drug therapy , Neoplasms/metabolism , Neoplasms/pathology , Protein Engineering , Signal Transduction/drug effects
6.
Proc Natl Acad Sci U S A ; 105(17): 6302-7, 2008 Apr 29.
Article in English | MEDLINE | ID: mdl-18424558

ABSTRACT

Identification of unique features of cancer cells is important for defining specific and efficient therapeutic targets. Mutant p53 is present in nearly half of all cancer cases, forming a promising target for pharmacological reactivation. In addition to being defective for the tumor-suppressor function, mutant p53 contributes to malignancy by blocking a p53 family member p73. Here, we describe a small-molecule RETRA that activates a set of p53-regulated genes and specifically suppresses mutant p53-bearing tumor cells in vitro and in mouse xenografts. Although the effect is strictly limited to the cells expressing mutant p53, it is abrogated by inhibition with RNAi to p73. Treatment of mutant p53-expressing cancer cells with RETRA results in a substantial increase in the expression level of p73, and a release of p73 from the blocking complex with mutant p53, which produces tumor-suppressor effects similar to the functional reactivation of p53. RETRA is active against tumor cells expressing a variety of p53 mutants and does not affect normal cells. The results validate the mutant p53-p73 complex as a promising and highly specific potential target for cancer therapy.


Subject(s)
Antineoplastic Agents/pharmacology , Catechols/pharmacology , DNA-Binding Proteins/metabolism , Mutant Proteins/metabolism , Neoplasms/pathology , Nuclear Proteins/metabolism , Small Molecule Libraries/pharmacology , Thiazoles/pharmacology , Tumor Suppressor Protein p53/metabolism , Tumor Suppressor Proteins/metabolism , Animals , Antineoplastic Agents/chemistry , Catechols/chemistry , Cell Line, Tumor , DNA-Binding Proteins/genetics , Drug Screening Assays, Antitumor , Gene Expression Regulation, Neoplastic/drug effects , Genes, Reporter , Humans , Mice , Neoplasm Proteins/genetics , Neoplasm Proteins/metabolism , Nuclear Proteins/genetics , RNA, Messenger/genetics , RNA, Messenger/metabolism , Small Molecule Libraries/chemistry , Thiazoles/chemistry , Transcription, Genetic/drug effects , Tumor Suppressor Protein p53/genetics , Tumor Suppressor Proteins/genetics
7.
Mol Biol (Mosk) ; 42(6): 1004-11, 2008.
Article in Russian | MEDLINE | ID: mdl-19140320

ABSTRACT

An enzymatic assay system is described that allows quantitative localization within different cellular structures of recombinant proteins. The system is based on alpha-complementation of beta-galactosidase. The large omega-fragment of beta-galactosidase is expressed in predefined cellular structures with the aid of attached protein localization signals. The obtained reporter cell lines are used for the introduction of a second construct that expresses a protein of study fused with a shorter alpha-fragment of beta-galactosidase. Physical proximity of the two recombinant proteins carrying beta-galactosidase fragments results in reconstitution of an active enzyme, and the activity can be measured in a plate reader. The recombinant constructs are based on lentiviral vectors, which allows rapid and efficient introduction of recombinant proteins into cells by infection with stocks of lentiviral particles. The efficiency of the system is demonstrated with transcriptional factor FOXO3A, which is shuttling between cytoplasm and nuclei in model colon carcinoma cell line RKO.


Subject(s)
Cell Nucleus/metabolism , Cytoplasm/metabolism , Forkhead Transcription Factors/metabolism , Genetic Vectors , Lentivirus , Recombinant Proteins/metabolism , Active Transport, Cell Nucleus/physiology , Cell Line, Tumor , Cell Nucleus/genetics , Cytoplasm/genetics , Forkhead Box Protein O3 , Forkhead Transcription Factors/genetics , Humans , Recombinant Proteins/genetics
8.
Mol Biol (Mosk) ; 41(3): 515-23, 2007.
Article in Russian | MEDLINE | ID: mdl-17685229

ABSTRACT

Inactivation of tumor suppressor p53 accompanies the majority of malignant diseases in humans. Restoration of p53 functions in tumor results in death of cancer cells, which can be used in cancer therapy. In cervical cancer a product of E6 gene of the human papilloma virus promotes accelerated degradation of p53 in proteasome system. Therefore, one of the approaches to reactivation of p53 in cervical carcinoma cells could be the use of small molecules that inhibit functions of viral proteins. By using as a test system human cervical carcinoma cells (HeLa cell line bearing human papilloma virus type 18, HPV-18) with introduced reporter construct that expresses beta-galactosidase under control of a p53-dependent promoter we carried out screening of a library of small molecules to select small molecules capable of reactivating transcriptional activity of p53. We then characterized the effects of two most active compounds in cell lines that differ in the status of p53-dependent signaling pathway. Both of the compounds caused specific activation of p53 in the cell lines expressing HPV-18, to a lesser extent--HPV-16, and do not cause any effect in control p53 negative cells, or in the cells with undisrupted p53 pathway. Activation of p53 in cervical carcinoma cells was accompanied by the induction of the p53-dependent gene CDKN1 (p21), by inhibition of proliferation, and by the induction of apoptosis. Both of the compounds were capable of deep inhibition of transcription from the HPV genome, which apparently was the cause for p53 reactivation in response to decreased expression of the E6 protein. The observed low toxicity for normal cells allows considering these chemical compounds as prototypes for future anticancer drugs.


Subject(s)
Antineoplastic Agents/pharmacology , DNA-Binding Proteins/metabolism , Human papillomavirus 18/drug effects , Oncogene Proteins, Viral/metabolism , Tumor Suppressor Protein p53/metabolism , Antineoplastic Agents/chemistry , Apoptosis , Benzodioxoles/chemistry , Benzodioxoles/pharmacology , Benzopyrans/chemistry , Benzopyrans/pharmacology , Cell Line, Tumor , Cell Proliferation/drug effects , Cyclin-Dependent Kinase Inhibitor p21/metabolism , Drug Screening Assays, Antitumor , Female , Genes, Reporter , HeLa Cells , Human papillomavirus 16/drug effects , Human papillomavirus 16/metabolism , Human papillomavirus 18/genetics , Human papillomavirus 18/metabolism , Humans , Promoter Regions, Genetic , Pyrans/chemistry , Pyrans/pharmacology , Quinolines/chemistry , Quinolines/pharmacology , Transcription, Genetic , Uterine Cervical Neoplasms , beta-Galactosidase/metabolism
9.
Mol Biol (Mosk) ; 40(3): 448-59, 2006.
Article in Russian | MEDLINE | ID: mdl-16813164

ABSTRACT

We have optimized lentiviral vector constructs and cassettes for expression of short hairpin RNAs (shRNAs) in order to create genome-wide library capable of inhibition of full variety of human mRNAs. The vector optimization has resulted in 15-20-fold improvement in virus stock titers. We found that in the context of lentiviral vector the most effective structure for the shRNA is simple hairpin with 21 nucleotide stem. The shRNA-expressing lentiviral constructs contain choice of puro(R), copGFP or H-2K(k) selective markers. The efficiency of the optimized library was evaluated in experiments on screening of shRNAs that reactivate oncosuppressor p53 in HeLa cells. The cells contained reporter construct with p53-dependent expression of a fluorescent protein, which allows cytofluorimetric isolation of cell population with reactivated p53.


Subject(s)
Gene Library , Genome, Human/genetics , Lentivirus , MicroRNAs/genetics , RNA Interference , Gene Expression Regulation , Genes, p53/genetics , Genetic Markers , Genetic Vectors , HeLa Cells , Humans , Nucleic Acid Conformation
10.
Mol Biol ; 40(3): 396-405, 2006 May 01.
Article in English | MEDLINE | ID: mdl-19777129

ABSTRACT

To obtain a whole genome library that suppresses the total diversity of human mRNAs, lentiviral vector constructs and a short hairpin RNA (shRNA) expression cassette were optimized. The optimization of the vector increased the virus titer in preparations by 15-20 times. A simple shRNA structure with a 21-bp stem proved to be the most effective. Lentivector-based shRNA expression constructs were obtained by using puro(R), copGFP, or H-2K(k) as a selectable marker. The efficiency of the optimized library was demonstrated when screening for shRNAs reactivating the tumor suppressor p53 in HeLa cells. Cells carried a reporter construct ensuring p53-responsive synthesis of a fluorescent protein, which allowed selection of cells with reactivated p53 by flow cytometry.

11.
Mol Biol (Mosk) ; 39(5): 905-9, 2005.
Article in Russian | MEDLINE | ID: mdl-16240724

ABSTRACT

Structure and dynamics of actin cytoskeleton play a role ih regulation of cell adhesion, spreading and migration. TRIP6 is a LIM domain-containing protein interacting with many actin-associated proteins and in addition modulating activity of certain transcription factors. To study functions of TRIP6 we inhibited its expression in A549 and A431 cells by short interfering RNAs (siRNAs). The TRIP6 knock-down lead to the increased number and length of stress fibers and to the induction of locomotive phenotype. There was observed decreased number and reorganization of focal adhesions revealed by staining for paxillin, and loss of cell to cell adhesions revealed by staining for E-cadherin. The above changes in cell morphology were accompanied by 2-fold increase in the cell motility rate assessed by the wound healing assay. Thus, down-regulation of TRIP6 in the cell lines used results in increase in the features characteristic to malignant transformation of epithelial cells. Possible mechanisms for the observed effects are discussed.


Subject(s)
Actin Cytoskeleton/ultrastructure , Adaptor Proteins, Signal Transducing/physiology , Carcinoma/ultrastructure , Transcription Factors/physiology , ATPases Associated with Diverse Cellular Activities , Actin Cytoskeleton/metabolism , Actins/metabolism , Adaptor Proteins, Signal Transducing/antagonists & inhibitors , Adaptor Proteins, Signal Transducing/genetics , Carcinoma/genetics , Carcinoma/metabolism , Cell Line, Tumor , Cell Movement , Cytoskeletal Proteins/metabolism , Down-Regulation , Humans , LIM Domain Proteins , Proteasome Endopeptidase Complex , RNA Interference , Transcription Factors/antagonists & inhibitors , Transcription Factors/genetics
12.
Anesteziol Reanimatol ; (2): 62-6, 2005.
Article in Russian | MEDLINE | ID: mdl-15938101

ABSTRACT

This study was undertaken to evaluate the efficiency of complex intensive therapy for multiple organ dysfunction syndrome (MODS) after cardiosurgical interventions at the resuscitative and intensive care unit of the A. N. Bakulev Research Center of Cardiovascular Surgery, Russian Academy of Medical Sciences. In 2003-2004, MODS developed in 70 (37%) of the neonatal infants operated on the heart and vessels. The babies' age ranged from 6 hours of life to 1 month (8.3 +/- 2.1 days of life, their body weight was from 1.7 to 4.1 kg (3.0 +/- 0.49 kg). All the patients were found to have significant renal and respiratory failures. There were more than 4 (4.1 +/- 0.5) failing vital viscera. The use of phosphodiesterase (III) inhibitors in therapy for acute left ventricular insufficiency significantly improved the performance of the left heart whereas nitric oxide inhalation significantly lowered pulmonary pressure in babies with acute right ventricular insufficiency and improved oxygenation in patients with MODS. The efficiency of nitric oxide inhalation in MODS significantly increased when it was used in combination with endotracheal administration of a surfactant and high-frequency oscillatory ventilation. Peritoneal dialysis effectively replaced renal function when acute renal failure (ARF) developed. Nevertheless, the development of ARF in the pattern of MODS is a marker of high mortality (89% in ARF versus 46% in MODS without ARF).


Subject(s)
Cardiovascular Abnormalities/surgery , Intensive Care, Neonatal , Multiple Organ Failure/therapy , Acute Kidney Injury/therapy , Female , Health Status Indicators , Humans , Infant, Newborn , Male , Multiple Organ Failure/mortality , Peritoneal Dialysis , Postoperative Period , Syndrome
14.
Bioorg Khim ; 30(5): 499-506, 2004.
Article in Russian | MEDLINE | ID: mdl-15562971

ABSTRACT

A 147-bp cDNA fragment was isolated from human lymphocytes activated with concanavalin A using the method of direct selection. A complete copy of the selected gene having total homology with the mitochondrial ribosomal gene MRPL37 was obtained by the RACE (rapid amplification of cDNA ends) technique. The MRPL37 gene was localized on human chromosome 1 using a DNA panel composed of somatic cellular human-hamster hybrids. The Northern blotting and RT-PCR (reverse transcription-polymerase chain reaction) demonstrated that the RNA of the human MRPL37 gene is widely represented in the lymphoma populations of Raji B cells and MT4 T cells, as well as in pancreas, liver, and lung embryonic fibroblasts WI-38 and LEH. The highest expression level of the MRPL37 mouse homologue was found in the cells of skeletal muscles, the heart, and organs of reproductive system: the uterus, ovaries, and testicles. A comparative analysis of the MRPL37 amino acid sequence with those of proteins represented in the Fasta33 and GenBank databases showed a homologous region in MRPL37 and PDCD9 (programmed cell death 9, MPRS30) proteins. The chicken homologue of PDCD9 is interesting because its overexpression causes apoptosis of the mouse fibroblasts C3H10T1/2. The existence of a common domain indicates possible similar functional peculiarities of the PDCD9 and MRPL37 genes and may imply the MRPL37 involvement in the process of apoptosis. The English version of the paper: Russian Journal of Bioorganic Chemistry, 2004, vol. 30, no. 5; see also http: // www.maik.ru.


Subject(s)
Gene Expression Regulation , Ribosomal Proteins/genetics , Amino Acid Sequence , Animals , Apoptosis/genetics , Base Sequence , Cell Cycle Proteins/genetics , Cell Cycle Proteins/metabolism , Cells, Cultured , Chickens/genetics , Chromosome Mapping , Chromosomes, Human, Pair 1 , Cloning, Molecular , DNA, Complementary , Female , Fibroblasts/pathology , Fibroblasts/physiology , Humans , Lymphoma, B-Cell/genetics , Lymphoma, B-Cell/pathology , Male , Mice , Mitochondrial Proteins , Molecular Sequence Data , Ribosomal Proteins/metabolism , Sequence Homology, Amino Acid , T-Lymphocytes/pathology , T-Lymphocytes/physiology , Tumor Suppressor Protein p53/genetics
15.
Gene ; 210(2): 239-45, 1998 Apr 14.
Article in English | MEDLINE | ID: mdl-9573374

ABSTRACT

The ril gene encoding a LIM domain protein of an unknown function was previously identified by differential expression cloning as a candidate tumor suppressor gene in rat fibroblasts (Kiess, M., Scharm, B., Aguzzi, A., Hajnal, A., Klemenz, R., Schwarte-Waldhoff, I., Schafer, R., 1995. Expression of ril, a novel LIM domain gene, is down-regulated in HRAS-transformed cells and restored in phenotypic revertants. Oncogene 10, 61-68). Searching for novel genes on human chromosome 5q31.1 by the cDNA selection technique, we isolated a cDNA clone identical with the cDNA of the human RIL gene (GenBank Accession No. X93510). The human 5q31.1 region is of interest because it contains the cytokine gene cluster and is frequently deleted in the malignant cells of patients with myelodysplasia and myeloid leukemia. Using Southern blot analysis and restriction mapping of genomic YAC (yeast artificial chromosome) and cosmid clones, we located the human RIL gene 240-260 kb telomeric to the IRF1 gene and characterized its genomic structure. PCR analysis indicated the presence of two alternative RIL transcripts in human fetal brain mRNA. The major transcript is identical with the RIL cDNA previously deposited in GenBank and contains seven exons distributed over 14.5 kb of genomic DNA with the two last 3'-exons coding a LIM domain. The minor transcript lacks the sixth exon compared with the major transcript, which leads to the loss of the LIM domain. We also identified two putative transcription start points (tsp) and sequenced the 5'-flanking region of RIL to reveal potential binding sites for transcriptional factors.


Subject(s)
Alternative Splicing , Chromosomes, Human, Pair 5 , DNA-Binding Proteins/genetics , Base Sequence , Cell Line , Chromosome Mapping , DNA, Complementary , Humans , LIM Domain Proteins , Molecular Sequence Data , Sequence Analysis, DNA
16.
Gene ; 155(2): 277-81, 1995 Apr 03.
Article in English | MEDLINE | ID: mdl-7721105

ABSTRACT

The genes encoding human interleukin-4 (IL-4) and interleukin-13 (IL-13) are located on segment q23-31 of chromosome 5 and encode two multifunctional lymphokines with some common functions. We have cloned 72 kb of human genomic DNA that contain IL-4 and IL-13 and their flanking sequences, and constructed a restriction map of this region. Using Southern analysis, we have shown that IL-13 is located 12 kb 5' to IL-4 and linked in a 'tail-to-head' fashion. We have also determined the complete nucleotide sequence of the DNA fragment (about 4.8 kb) containing IL-13 and its 5' flanking regulatory region (2.1 kb) with a 'CpG island'. We identified potential binding sites for a different transcription factors in the 5' flanking region and in the first intron of IL-13. Comparison of IL-13 and IL-4 revealed considerable similarity in the structural organization of these genes and also many potential binding sites for transcription factors common to both genes: AP1, AP2, AP3, PEA3, HRE, TCF-1, GATA-3 and the interferon-inducible and enhancer elements. These results, along with the similarity in functional activity of IL-4 and IL-13 suggests that their expression may be coregulated.


Subject(s)
Chromosomes, Human, Pair 5/genetics , Interleukin-13/genetics , Interleukin-4/genetics , Amino Acid Sequence , Base Sequence , Chromosome Mapping , Cloning, Molecular , Gene Expression Regulation , Humans , Molecular Sequence Data , Sequence Homology, Nucleic Acid , Transcription, Genetic
17.
Mol Biol (Mosk) ; 28(2): 307-12, 1994.
Article in Russian | MEDLINE | ID: mdl-8183262

ABSTRACT

Inhibition of the influenza virus protein NP mRNA with derivatives of an antisense oligonucleotide complementary to the 5' terminus of the mRNA was investigated. The derivatives were prepared by conjugation of aromatic 2-chloroethylamine, cholesterol, porphyrin, and phenazine groups to the 5'-terminal phosphate of the oligonucleotide. The most efficient inhibitors were found to be the conjugates bearing the alkylating, cholesterol and phenaznium groups.


Subject(s)
Nucleoproteins , Oligonucleotides, Antisense/pharmacology , Orthomyxoviridae/metabolism , Protein Biosynthesis/drug effects , RNA, Messenger/genetics , Viral Core Proteins/genetics , Base Sequence , Cholesterol/metabolism , Ethylamines/metabolism , Molecular Sequence Data , Nucleocapsid Proteins , Phenazines/metabolism , Porphyrins/metabolism
18.
Mol Biol (Mosk) ; 27(4): 781-9, 1993.
Article in Russian | MEDLINE | ID: mdl-7689694

ABSTRACT

We investigated the interaction of AMV reverse transcriptase and Klenow fragment with oligonucleotide derivatives carrying different 3'- or 5'-terminal reactive groups. It was shown that the attachment of phenazinium, ethidium, and daunomycin residues to the 5'-terminal phosphate stabilized the enzyme template primer complexes, while cholesterol and hemin residues generally decreased their stability. The increased stability in solution correlated to a certain extent with the increase in affinity of the modified primers to the enzyme template complex. Coupling of bulky R residues to the primers had a weak effect on the maximal rate of primer conversion, which is likely to be a result of the lack of strong contacts between the substituents and the enzyme, and steric obstacles hindering translocation of the primer enzyme complex. We analyzed the inhibitory effect of 23 oligonucleotide derivatives (both complementary and noncomplementary to the template) with modified 3'- and 5'-ends, and revealed several analogs inhibiting polymerization catalyzed by AMV reverse transcriptase by 70-100% at 0.1-1 microM concentrations of the reagents.


Subject(s)
Avian Myeloblastosis Virus/enzymology , DNA, Single-Stranded/chemistry , RNA-Directed DNA Polymerase/metabolism , Catalysis , Cholesterol/chemistry , DNA Polymerase I/chemistry , Daunorubicin/chemistry , Ethidium/chemistry , Phenazines/chemistry , Polymers , Porphyrins/chemistry
19.
Mol Biol Rep ; 18(1): 43-7, 1993 Jun.
Article in English | MEDLINE | ID: mdl-7694077

ABSTRACT

Km and Vmax values for d(pT8) and its derivatives containing various 5'-end groups were estimated in the reaction of DNA polymerization alpha catalyzed by DNA polymerase alpha and HIV-RT. The effect of 5'-end modification of primer is more pronounced in the case of HIV-RT. Strong influence is observed for an intercalating (ethidium) group. The affinity of EtpT8 is 200-fold higher than that of d(pT8). Attachment of Phn-, Dnm- and Hem-groups results in the increase of affinity of modified primer from 10 up to 20 times. For DNA polymerase alpha the influence of modifiers on primer affinity is much weaker. The effect of 5'-end residues on the Vmax values is also more pronounced for HIV RT. The way to improve selective interaction of oligonucleotide derivatives with the primer site of HIV RT is suggested.


Subject(s)
DNA Polymerase II/metabolism , HIV-1/enzymology , Poly T/metabolism , RNA-Directed DNA Polymerase/metabolism , DNA/metabolism , HIV Reverse Transcriptase , Humans , Molecular Structure
20.
Bioorg Khim ; 19(4): 439-54, 1993 Apr.
Article in Russian | MEDLINE | ID: mdl-8494567

ABSTRACT

A method for coupling 2,4-di[alpha-(2-hydroxyethoxy)ethyl] deuteroporphyrin dimethyl ether (IX), DDPOH and its complexes with metals to the 5'- or 3'-end of oligonucleotides was elaborated. In the presence of an oxidizing agent (H2O2), Fe(III)DDP-derivatives of oligonucleotides modified single-stranded DNA. The reaction was strictly site-specific and occurred at two neighbouring guanosine residues. A few types of modification were observed: cross-linking, modification leading to DNA cleavage upon piperidine treatment, and direct chain scission. The total modification yield reached 90%. Covalent attachment of Fe(III)DDP-group to oligonucleotides increased the efficiency of their uptake and the melting temperature of their complementary complexes.


Subject(s)
DNA/chemistry , Deuteroporphyrins/chemistry , Metals/chemistry , Oligonucleotides/chemistry , Porphyrins/chemistry , Base Sequence , Cross-Linking Reagents , Molecular Sequence Data , Oxidation-Reduction
SELECTION OF CITATIONS
SEARCH DETAIL
...