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N A Lebedeva

Publications and source records attributed to N A Lebedeva.

At least 19 recordsLinked to original sources
PubMed2005

Analysis of interactions of DNA polymerase beta and reverse transcriptases of human immunodeficiency and mouse leukemia viruses with dNTP analogs containing a modified sugar residue.

Substrate properties of various morpholinonucleoside triphosphates in the reaction of DNA elongation catalyzed by DNA polymerase beta, reverse transcriptase of human immunodeficiency virus (HIV-1 RT), and reverse transcriptase of Moloney murine leukemia virus (M-MuLV RT) were compared. Morpholinonucleoside triphosphates were utilized by DNA polymerase beta and HIV-1 reverse transcriptase as substrates, which terminated further synthesis of DNA, but were virtually not utilized by M-MuLV reverse transcriptase. The kinetic parameters of morpholinoderivatives of cytosine (MorC) and uridine (MorU) were determined in the reaction of primer elongation catalyzed by DNA polymerase beta and HIV-1 reverse transcriptase. MorC was a more effective substrate of HIV-1 reverse transcriptase and significantly less effective substrate of DNA polymerase beta than MorU. The possible use of morpholinonucleoside triphosphates as selective inhibitors of HIV-1 reverse transcriptase is discussed.

Catalysis↗
PubMed2004

Interaction of replication protein A with photoreactive DNA structures.

A new photoreactive oligonucleotide derivative was synthesized with a perfluoroarylazido group attached to the 2'-position of the ribose fragment of the 5'-terminal nucleotide. Using this conjugate, photoreactive DNA duplexes were produced which contained single-stranded regions of different length, single-stranded breaks (nicks), and also ds duplex with a photoreactive group inside one of the chains. These structures imitate DNA intermediates generated at different stages of DNA replication and repair. The interaction of replication protein A (RPA) with the resulting DNA structures was studied using photoaffinity modification and gel retardation assay. Independently of the DNA structure, only the large subunit of RPA (p70) was crosslinked to photoreactive DNAs, and the intensity of its labeling increased with decrease in the size of the single-stranded region and was maximal in the case of the nick-containing DNA structure. By gel retardation, the most effective binding of RPA to this structure was shown, whereas the complexing of RPA with DNA containing the unmodified nick and also with the full duplex containing the photoreactive group inside the chain was significantly less effective. The data suggest that RPA should be sensitive to such damages in the double-stranded DNA structure.

Azides↗
PubMed2003

A new binary system for photosensitized labeling of DNA polymerases in nuclear extract.

A binary system of reagents was used for photosensitized labeling of proteins of bovine testis nuclear extract. A dUTP analog containing 4-azido-2,5-difluoro-3-chloropyridyl group (FAP-dUTP) was used for the first time as a component of the binary system, and a dUTP analog containing the pyrenyl group (Pyr-dUTP) was used as a photosensitizer. Photoaffinity labeling of proteins of nuclear extract was performed using the radioactively labeled DNA duplex with the photoreactive FAP group at the 3;-end of elongating DNA strand and analog of the deoxyribose phosphate residue (3-hydroxy-2-hydroxymethyltetrahydrofuran (F) 5;-phosphate) at the 5;-end of the nick. Such structure is formed by the action of nuclear extract enzymes from the initial DNA duplex containing a synthetic apurine/apyrimidine site and is a photoreactive analog of a long-patch base excision repair intermediate. UV-irradiation modified a limited number of proteins of the nuclear extract. As shown using specific antibodies, the new binary system of photoreagents increases the efficiency of DNA polymerase beta labeling.

Animals↗
PubMed2003

Thermostable DNA polymerase from Thermus thermophilus B35: preparation and study of a modified form of the enzyme with high affinity to ddNTP.

The hybrid protein consisting of Tte DNA polymerase fragment and mutant Taq DNA polymerase (F667Y) fragment in the ratio 20 : 1 was constructed. Affinity of the modified enzyme (substitutions F669Y, V667I, and S692Q) to ddNTP was two orders higher than that of the wild type enzyme. The modified enzyme was used for sequencing DNA fragment with total deoxyguanosine and deoxycytidine content of 68%. In the polymerase chain reaction, the modified enzyme exhibits properties typical of the wild type Tte DNA polymerase.

Autoradiography↗
PubMed2003

[Results of international study of quality of life of patients with stable angina treated with nitrates (IQOLAN)].

Effect of transition from multiple administration of short acting nitrates to once daily use of isosorbide-5-mononitrate on angina class and quality of life was studied in 280 patients with stable angina pectoris. Transition to isosorbide-5-mononitrate was associated with increases of (in units) exercise tolerance (Delta=-6.6+/-0.35, p<0.001), satisfaction from treatment (Delta=-2.1+/-0.12, p<0.001), degree of psychological discomfort (Delta=-2.8+/-0.21, p<0.001), decreases of numbers of attacks of angina (Delta=-4.0+/-0.22, p<0.001) and side effects (Delta=-4.1+/-0.29, p<0.001), increase of average distance of walking without chest pain or dyspnea (from 372.1+/-415.1 to 586+/-663.5 m), shortening of duration of episodes of angina (from 4.7+/-4.0 to 4.1+/-7.2 min), significant lowering of angina class (p<0.0001). Thus in patients with ischemic heart disease long acting formulation of isosorbide-5-mononitrate provided rapid onset and stability of therapeutic action what eventually resulted in effective prevention of attacks of angina and improvement of quality of life.

Angina Pectoris↗
PubMed2002

Highly efficient labeling of DNA polymerases by a binary system of photoaffinity reagents.

A binary system of photoaffinity reagents was proposed earlier for highly efficient labeling of DNA polymerases by 5;-[32P]DNA primers. In the present study we demonstrate the feasibility of this approach to increase the efficiency of DNA polymerase labeling. A photoactive 2,3,5,6-tetrafluoro-4-azidobenzoyl (FAB) group was incorporated at the 3;-end of 5;-[32P]DNA primers synthesized by DNA polymerase beta or Tte in the presence of one of the dTTP analogs--FAB-4-dUTP, FAB-9-dUTP, or FAB-4-ddUTP. The reaction mixture was irradiated by light with wavelength of 334-365 nm (direct labeling) or 365-450 nm in the presence of photosensitizer, one of dTTP analogs containing a pyrene moiety, Pyr-6-dUTP or Pyr-8-dUTP. In the case of the binary system of photoaffinity reagents, a FAB group is activated by energy transfer from sensitizer localized in the dNTP-binding site of DNA polymerase in the triple complex, comprised by reagent, DNA polymerase, and Pyr-6(8)-dUTP. Direct activation of the FAB group under these conditions is negligible. The most efficient photolabeling of DNA polymerases was observed with a primer containing a FAB-4-dUMP group at the 3;-end, and Pyr-6-dUTP as a photosensitizer. Using 10-fold molar excess of photoreagent to DNA polymerase beta, the labeling efficiency was shown to achieve 60%, which is 2-fold higher than the efficiency of the direct DNA polymerase labeling under harsher conditions (334-365 nm).

DNA Polymerase beta↗
PubMed2001

A binary system of photoreagents for high-efficiency labeling of DNA polymerases.

To increase the efficiency of photoaffinity labeling of DNA polymerases, a binary system of photoaffinity reagents was applied. Photoreactive radioactive primers were synthesized by DNA polymerases beta (pol beta) or DNA polymerase from Thermus thermophilus (pol Tte) using a template-primer duplex in the presence of a dTTP analogue containing 4-azidotetrafluorobenzoyl group linked via spacers of varying length to 5-position of uridine ring- 5-[N-(2,3,5,6-tetrafluoro-4-azidobenzoyl)-amino-trans-propenyl-1]-2'-deoxyuridine-5'-triphosphate (FAB-4-dUTP) or 5-[N-[[(2,3,5,6-tetrafluoro-4-azidobenzoyl)-butanoyl]-amino]-trans-3-aminopropenyl-1]-2'-deoxyuridine-5'-triphosphate (FAB-9-dUTP). The reaction mixtures were UV irradiated (lambda = 365-450 nm) in the absence or presence of a dTTP analog, containing a pyrene moiety-5-[N-(4-(1-pyrenyl)-butylcarbonyl)-amino-trans-propenyl-1]-2'-deoxyuridine-5'-triphosphate (Pyr- 8-dUTP) or 5-[N-(4-(1-pyrenyl)-ethylcarbonyl)-amino-trans-propenyl-1]-2'-deoxyuridine-5'-triphosphate (Pyr-6-dUTP). The most efficient crosslinking of both DNA polymerases was observed in the case of photoreactive DNA primer, carrying the FAB-4-dUMP moiety at the 3'-end, and Pyr-6-dUTP as a sensitizer. The binary system of photoaffinity reagents allows increasing photoaffinity labeling of the both DNA polymerases in comparison to the primer crosslinking without photosensitizer.

DNA Primers↗
PubMed2000

Highly selective affinity labeling of DNA-polymerase from Thermus thermophilus B35 by a binary system of photoreactive agents.

The thermostable DNA-polymerase from Thermus thermophilus B35 (Tte-polymerase) was affinity labeled by a binary system of photoreagents comprising base-substituted TTP analogs. The 5;-[32P]-labeled primer was elongated by Tte-polymerase in the presence of a TTP analog containing the photoreactive 2,3,5, 6-tetrafluoro-4-azidobenzoyl group (FAB-4-dUTP). Then the reaction mixture was UV-irradiated (365-450 nm) in the presence or the absence of a photosensitizer (TTP analog containing a pyrene moiety, Pyr-dUTP). The initial rate of the Pyr-dUTP-sensitized photomodification was almost 10-fold higher than the rate of direct photomodification (in the absence of Pyr-dUTP); in the case of the sensitized modification, the product of covalent cross-linking of the photoreactive primer with Tte-polymerase was apparently homogenous according to the data of electrophoresis. The enzyme was protected from the photosensitized modification by dNTP. To confirm the selectivity of the photosensitized modification of Tte-polymerase, another DNA-binding protein (human replication factor A, RPA) was added to the reaction mixture. In the presence of the photosensitizer (Pyr-dUTP), RPA was not labeled and only Tte-polymerase was modified, whereas in the case of direct modification, Tte-polymerase and the p32 and p70 subunits of RPA were labeled. The suggested method enables highly selective affinity modification of DNA-polymerases.

Affinity Labels↗
PubMed1999

Sensitized photomodification of mammalian DNA polymerase beta. A new approach for highly selective affinity labeling of polymerases.

To enhance the specificity of polymerase photoaffinity labeling, a novel approach based on sensitized photomodification has been developed. A base-substituted analog of TTP containing a pyrene group (PyrdUTP) was synthesized and used as an active site-bound photosensitizer for photoaffinity modification of DNA polymerase beta (pol beta). 5'-[32P]-labeled primer was elongated in situ by pol beta with a photoreactive analog of TTP (FAB-4-dUTP). The pyrene sensitizer (PyrdUTP), excited by light (365-450 nm), can activate the photoreagent, cross-linking it to pol beta as a result of fluorescence resonance energy transfer. The initial rate of pol beta photomodification was shown to increase by a factor of ten. The selectivity of pol beta photosensitized modification was proved by adding human replication protein A.

Animals↗
PubMed1999

[Sub-atrophic and atrophic pharyngitis as a manifestation of dystrophic pharyngeal mucosa in presenile, senile and long living persons].

The authors consider diagnostic, clinical and symptom characteristics of atrophic pharyngitis (AP) in presenile, senile and long-living persons and conclude that AP is age-specific dystrophic lesion of the pharynx. Diagnosis formulation by three degrees are suggested: pharyngeal dystrophy (PD) of degree I (only local symptoms cured after timely treatment), PD of degree II (local symptoms + neurotic condition which manifests with cancerophobia, persistent sensation of a foreign body), PD of degree III (persistent cancerophobia demanding psychotherapeutic treatment or psychiatric care).

Aged↗
PubMed1999

Thermostable DNA polymerase from Thermus thermophilus B35: cloning, sequence analysis, and gene expression.

The nucleotide sequences of three thermostable DNA polymerase (Taq, Tth, and Tfl) genes were analyzed and high conserved regions typical for this polymerase family were identified. Using primers for one of the conserved regions, the genomic DNA fragment of T. thermophilus B35 strain was amplified. The resulting fragment was cloned into a plasmid and used as a hybridization probe with digests of T. thermophilus B35 DNA cleaved by different restriction endonucleases. A restriction DNA fragment carrying the full-length Tte polymerase gene was found, cloned, and sequenced. The primary structures of the Tte and Tth DNA polymerase genes were analyzed. The Tte-pol gene was recloned into an expression vector and recombinant protein was purified to homogeneity. The properties of Tte-pol in the polymerase chain reaction were investigated.

Amino Acid Sequence↗
PubMed1998

Thermostable DNA-polymerase from Thermus thermophilus B35: isolation and characterization of some properties.

Thermostable Tte DNA-polymerase was isolated from the strain Thermus thermophilus B35 which was found in hot spring water. The enzyme with molecular mass 87 kD was isolated using sequential chromatography on DEAE-Sepharose, hydroxylapatite, hexyl-agarose, and heparin-Sepharose. Biochemical properties of Tte DNA-polymerase are similar to those of Tth DNA-polymerase isolated from Thermus thermophilus HB8; however, practical application of Tte-Pol seems to be more favorable due to higher temperature optimum of this enzyme and lack of restriction endonucleases in the initial strain.

Cations, Divalent↗
PubMed1991

[The effect of proton therapy on the protein-amino acid metabolism in Itsenko-Cushing disease].

The composition and blood concentration of free aminoacids and other aminocompounds were investigated using automatic aminoacid analysis by a physiological program in 12 patients with diagnosed corticotropin secreting hypophyseal disease (Itsenko-Cushing disease) before and 1-3 years after proton "hypophysectomy". Imbalance of components of the glucose-alanine cycle, expressed in hypoalaninemia and raised concentration of aminoacids with a bifurcated lateral chain, was detected against a background of marked hypercorticotropinemia and hypercorticolism. Single irradiation of the pituitary gland with a proton beam at absorbed doses of 100-120 Gy in the maximum of a deep field resulted in remission and normalization of aminograms of blood plasma (suggesting pathogenicity of proton-beam therapy) for 6-12 mos. The authors discussed probable mechanisms of the interrelationship of protein-amino-acid and carbohydrate metabolic derangements and their correlation with a type of changes in the hormonal balance.

Adult↗
PubMed1989

[Osteopathy in malabsorption syndromes].

Electrolytic imbalance is a frequent finding in malabsorption syndrome. Derangement of calcium metabolism present clinically in different variants is most serious. Some patients develop hypocalcemia manifesting clinically with specific myasthenia, paresthesias, convulsions, hemorrhages, etc. In other variants severe skeletal lesions are seen which may become dominating in the clinical picture though hypocalcemia was absent. Investigation of calcium metabolism, hormonal profile (parathormone, in particular) in malabsorption syndrome can prognosticate and prevent the onset of osteomalacia.

Adolescent↗