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Biomedical subjects

S A Kuznetsova

Publications and source records attributed to S A Kuznetsova.

At least 19 recordsLinked to original sources
PubMed2002

Backward electron transport in photosystem 2 reaction center and temperature dependence of delayed luminescence characteristics.

The temperature dependence of parameters of light-induced changes in millisecond delayed luminescence (half-width of the maximum, maximal and steady-state luminescence intensity) is studied within the temperature range from -23 to 45 degrees C in leaf segments of Chinese rose (Hibiscus rosa sinensis). Delayed luminescence (DL) is induced and registered by a homemade setup based on a Lewis-Kasha-type phosphoroscope. The temperature dependence of steady-state luminescence intensity is shown to have two maxima, at -10 and 35 degrees C. At room temperatures, the steady-state value of luminescence intensity is minimal, and its value correlates with the temperature tolerance of the plant. Depending on cooling and heating regimes, the DL steady-state value vs. temperature curves is found to be different. We suppose this effect to be caused by temperature-induced destructive changes in the structure of photosystem 2 reaction centre and probably by salting out.

Electrons↗
PubMed2002

On the involvement of the water-polaron mechanism in energy trapping by reaction centers of purple bacteria.

A locus for binding a mobile water molecule was searched for in the immediate vicinity of the special pair in the reaction center. Using the PROTEUS PC-program (a part of the GRASP package) atomic structures of the reaction centers were analyzed in purple bacteria Rhodopseudomonas viridis and Rhodobacter sphaeroides. In both structures the loci for binding mobile water molecules were found at the distance of about 4.5 A from the middle of the special pair in the reaction center. The reorientation of a hydrogen atom of this water molecule in the electric field of the excited special pair required energy of no less than 40 MeV that corresponded to predictions of the water-polarization model of trapping of electron excitation which was developed by M. V. Fok and one of the authors of this article.

Electric Conductivity↗
PubMed1999

Efficient synthesis of DNA dumbbells using template-induced chemical ligation in double-stranded polynucleotides closed by minihairpin fragments.

The chemical ligation of 17 50-54-membered nicked DNA dumbbells with different closing fragments, nick positions, and nucleotides facing the nick were investigated. T4, T5, GTA4C, GCGA2GC, and GCGA3GC sequences were chosen as the closing fragments. The nicks were placed in the center of the duplex stem or were adjacent to the closing fragments. N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide and cyanogen bromide were used as the condensing agents. We showed that the ligation efficiency is 10%-90% depending on the sequence of the closing fragments, nick position, and nucleotides facing the nick. Coupling yields of 80%-90% were observed when the nick was situated in the middle of the molecule between two T residues or was adjacent to GCGA2GC or GCGA3GC minihairpins. In the last case, the reacting 3'-phosphate and 5'-hydroxy groups were brought close together by only two base pair minihairpins. The coupling yields did not depend on the nature of the condensing agent. On the basis of the results obtained, we believe a rational design of nicked DNA dumbbells has been developed for efficient chemical synthesis of closed dumbbells.

Base Pairing↗
PubMed1998

Design and synthesis of double-stranded oligonucleotides containing reactive acylphosphate internucleotide groups.

DNA duplex and dumbbells containing chemically active acylphosphate internucleotide groups were synthesized. To obtain these compounds the chemical ligation method was used. The acylphosphate group was inserted into a DNA duplex and dumbbells as a result of template-directed condensation of 5'-phosphate and especially introduced 3'-carboxy groups of oligonucleotides. 1-Ethyl-3-(3'-dimethylaminopropyl)carbodiimide (EDC) was used as a condensing agent. Oligonucleotides containing a carboxy group were obtained by the interaction of their 3'-phosphate with glycine methyl ester under the action of EDC, followed by ester hydrolysis. The yields of acylphosphate-containing double-stranded oligonucleotides achieved 15-25% depending on the structure of their precursors. It was shown that these compounds are acylating agents and are efficiently cleaved in near-physiological conditions under the action of ethylenediamine or N-methylimidazole. These results indicate that double-stranded oligonucleotides carrying acylphosphate internucleotide groups could constitute new crosslinking reagents for affinity modification of DNA recognizing proteins.

Base Sequence↗
PubMed1996

Crosslinking of double-stranded oligonucleotides containing O-methyl-substituted pyrophosphate groups to the HNF1 transcription factor in nuclear cell extract.

Probing of the HNF1 (hepatocyte nuclear factor I) DNA-binding region using a set of DNA duplexes containing pyrophosphate or O-methyl-substituted pyrophosphate internucleotide groups at different positions of the HNF1 recognition sequence was performed. The histidine-tagged HNF1/1-281 DNA binding domain and nuclear extract from rat liver were used. We showed that HNF1 from these species specifically binds to modified DNA duplexes. A correlation in binding affinity of both types of duplexes was detected. Crosslinking of the HNF1 DNA-binding domain and HNF1 in nuclear liver extract to DNA duplexes carrying O-methyl-substituted pyrophosphate groups was observed. The crosslinking efficiency of HNF1 in liver extract to substituted pyrophosphate-modified DNA duplex, containing a reactive internucleotide group between nucleotides G and T of the GT dinucleotide immediately 5' to the TAAT recognition sequence, amounts to 40% of the efficiency of non-covalent association. Nonspecific crosslinking of the reactive DNA duplexes to other components of nuclear extract was not observed. These results indicate that DNA duplexes carrying substituted pyrophosphate internucleotide groups can specifically bind and crosslink with DNA-binding proteins, especially transcription factors in crude preparations and could constitute a potential tool to control the expression of disease-causing genes.

Animals↗
PubMed1996

Cross-linking of SsoII restriction endonuclease to cognate and non-cognate DNAs.

Specific and non-specific interactions of SsoII restriction endonuclease (R.SsoII) were probed by the method of covalent attachment to modified DNA containing an active monosubstituted pyrophosphate internucleotide bond instead of a phosphodiester one. R.SsoII with six N-terminal His residues was shown to be cross-linked to duplexes with this type of modification, either containing or not the recognition sequence. Competition experiments with covalent attachment of R.SsoII to activated DNAs demonstrated the similar affinity of the enzyme to cognate and non-cognate DNAs in the absence of cofactor, Mg2+ ions.

Base Sequence↗
PubMed1996

[Synthesis and properties of covalently-closed DNA duplexes containing pyrophosphate and and substituted pyrophosphate internucleotide groups].

A new method for the efficient synthesis of covalently closed DNA duplexes (DNA dumbbells) and the introduction of pyrophosphate and substituted pyrophosphate internucleotide groups into their structure is proposed. The method is based on chemical ligation in DNA duplexes that are formed by a polynucleotide the ends of which are brought together due to the introduction of the minihairpin structure [sequence: see text]. DNA dumbbells containing a pyrophosphate (substituted pyrophosphate) group result from the interaction as being between the 3'-terminal phosphate (methylphosphate) group of the polynucleotide and the 5'-terminal phosphate group of deoxyguanosine of the minihairpin sequence, which flanks the polynucleotide from the 5' end. 1-Ethyl-3-(3'-dimethylaminopropyl) carbodiimide was used as a condensing agent. The yield of covalently closed 42-mer DNA duplexes containing a pyrophosphate group was 98%, that of duplexes with a substituted pyrophosphate group was 25%. The reactivity of the substituted pyrophosphate group incorporated into DNA dumbbells was studied. It is shown that the group efficiently interacts with nucleophiles in an aqueous medium at pH 8.0.

DNA↗
PubMed1995

Modified substrates as probes for studying uracil-DNA glycosylase.

In order to study the mechanism of action of uracil-DNA glycosylase (UDG) from human placenta, single-stranded (ss) and double-stranded (ds) oligodeoxyribonucleotides (oligos), containing deoxyuridine (dU) and a wide variety of their analogs were used. It was shown that UDG has a twofold preference for ss oligos over ds oligos and a twofold preference for intermolecular duplexes over similar hairpin-like duplexes. The replacement of dU with 1-(beta-D-2'-deoxy-threo-pentofuranosil)uracil (xU) or 1-(beta-D-3'-deoxy-threo-pentofuranosil)uracil (tU), which results in a change in sugar hydroxyl configuration, has no influence on UDG binding to such substrates, but inhibits uracil removal. A oligo containing 2'-deoxy-2'-fluorouridine (flU), with a 3'-endo conformation of modified sugar is recognized by UDG 100-200-fold less efficiently than the natural ones. F or Br atoms or a methyl group were introduced at position 5 of a dU residue in an oligo. It was shown that the nature of a substituent at this position is essential for UDG function.

Base Sequence↗
PubMed1994

[Affinity modification of EcoRII restriction endonuclease by a DNA-duplex containing a monosubstituted pyrophosphate internucleotide bond].

Oligonucleotide duplex with an active monosubstituted pyrophosphate bond within the recognition site of the EcoRII restriction endonuclease was cross-linked to this enzyme with a yield of 10-15%. The cross-linking specificity was proved by the absence of the cross-linking to a DNA duplex with the same modification but without the EcoRII recognition site as well as by unmodified EcoRII substrate's inhibition of the cross-linking.

Affinity Labels↗
PubMed1991

[Chemical reactions in double helical nucleic acids. XIII. Directed introduction of acylphosphate internucleotide bonds into the DNA-duplex structure].

DNA duplex, containing an acylphosphate internucleotide bond in a predetermined position of the sugar-phosphate backbone, was synthesized. The synthesis was carried out by condensing on the complementary matrix two heptanucleotides, one of which possessed at the 3'-end a glycine residue, connected with the oligonucleotide by the phosphoramide bond, whereas the 5'-end phosphate group of the other was activated with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDAC). The yield of the oligonucleotide with an acylphosphate bond was 24%. The stability and chemical properties of the synthesized compound were studied in comparison with analogous oligonucleotide containing a substituted pyrophosphate internucleotide bond. The former was shown to be an effective acylating agent in the aqueous medium in contrast to the latter which is a phosphorylating agent.

Acylation↗
PubMed1990

[Chemical reactions in double-stranded nucleic acids. IX. Directed introduction of substituted pyrophosphate bonds into DNA structure].

An effective synthesis of oligodeoxyribonucleotides containing a substituted pyrophosphate bond in the definite position of the sugar-phosphate backbone has been developed by template-directed condensation of two heptanucleotides. One of them containing 5'-phosphate group to be activated and 3'-phosphate group of the other being substituted with ethoxy-, buthylamino-, morpholino- or ethyl glycinate residues. Water-soluble carbodiimide (EDAC) proved to be more efficient in the phosphate group activation than N-hydroxybenzotriazole ester (yields of substituted pyrophosphates 35-80 and 10-15% respectively). The substituted pyrophosphate bong is quite stable in neutral aqueous solution. Mild conditions of selective cleavage of this bond yielding the initial oligonucleotides were found.

Base Sequence↗
PubMed1989

[Significance of immunocytochemical indices of body reactivity of workers engaged in the production of carbonic plastics on the base of formaldehyde-containing and epoxide resins].

Findings of the complex study carried out by means of hematologic, cytochemical and immunologic methods on 130 workers engaged for the 5-year period in carbonic plastic production on the basis of methanol-containing and epoxy resins are presented. Dependence of the trends and intensity of reactivity changes on the character of leading unfavourable occupational factors has been established. Immunotoxicity of carbonic plastic dust and prevalence of sensitization reaction under the impact of industrial chemical allergens (methanol, epichlorohydrine) are mainly demonstrated. The study has indicated immunocytochemical signs of forming skin allergic pathology under cytotoxic effect. Groups of workers have been registered for medical supervision on the basis of an individual analysis of functionally interdependent immunologic and cytochemical indices.

B-Lymphocytes↗
PubMed1988

[Synthesis and properties of DNA duplexes containing hydrocarbon bridges instead of a nucleoside residue].

DNA duplexes 14 bp long containing an EcoRII and MvaI restriction site in which a nucleoside is substituted by 1,3-diaminopropane or 1,3-propanediol residue have been chemically synthesized. Diaminopropane bridge was introduced by the chemical ligation, whereas the oligonucleotide containing propanediol was prepared by automatic solid phase phosphoroamidite method on "Victoria-4M" synthesizer. As CD and UV spectra show, the modification destabilises the duplex by 18-20 degrees C without essential distortion of the double helix, except for increase of the conformational mobility in the modified site.

Base Sequence↗
PubMed1988

[Chemical reactions in double-stranded nucleic acids. V. Directed introduction into the DNA sugar-phosphate backbone of aliphatic diamine or glycol residues].

The chemical ligation method was used for directed introduction of aliphatic diamines or glycols into sugar-phosphate backbone of DNA. Via condensation of heptanucleotide derivatives at the terminal phosphate (aminoalkylamides or hydroxyalkyl esters) with the adjacent heptanucleotide on the corresponding template, 14 bp DNA duplexes containing residues of ethylene-, propylene- or hexamethylenediamine, as well as residues of ethylene- or propyleneglycol in one of its strands, were synthesised. In a similar way duplexes were obtained in which residues of the above diamines or glycols are substituted for a mononucleotide in one of the complementary strands. Examplified by synthesis of DNA duplexes containing ethylenediamine or ethyleneglycol residues, three methods of the phosphate group activation using carbodiimide, imidazolide and N-hydroxybenzotriazole ester were tested; the last method gave the highest yields and purity of the products. Yields of duplexes without nucleotide omissions were 50, 36 and 7% for aminoethyl, aminopropyl and aminohexylamides, and 20 and 17% for hydroxyethyl and hydroxypropyl esters, respectively, whereas duplexes with nucleotide omissions were synthesised with lower yields. Each of the modified DNA duplexes thus obtained contains a recognition site of EcoRII, SsoII or MvaI restriction nucleases, thus being a potential substrate of these enzymes.

Base Sequence↗
PubMed1987

[Interaction of EcoRII restriction and modification enzymes with synthetic DNA fragments. IX. Cleavage of substrates with point modifications in the recognition site and flanking sequences].

Ability of the EcoRII restriction endonuclease to cleave 14-base-pair DNA duplexes with nucleotide substitutions in the recognition site CCA/TGG and in the adjacent base pair has been studied. Modifications leading to a local change in the substrate conformation (rU residue in and outside the recognition site, A.A- or A.C-pairs in the flanking sequence) reduce the rate of hydrolysis, the effect being maximal when the modified base pair is outside the recognition site. No digestion occurs when the internal dC-residue of the recognition site is 5-methylated in one or both strands. Replacement of dT residue in the EcoRII recognition site by dfl5U residue results in a dramatic inhibition of hydrolysis. Km and kcat for the cleavage of 14-base-pair DNA duplex have been determined. The cleavage rate of the dT-containing strand of the recognition site in 1.5 fold higher comparing with the dA-containing strand. The cleavage of both strands of the substrate by EcoRII endonuclease is confirmed to proceed in one enzyme-substrate complex.

Base Sequence↗
PubMed1987

[Chemical reactions in double-stranded nucleic acids. The nature of the bond formed during chemical ligation using a cis-diol group].

Chemical and enzymatic ligation between the 5'-terminal phosphate of one oligonucleotide and the 3'-terminal 2',3'-cis-diol group of the other oligonucleotide on a complementary template was studied. Carbodiimide, imidazolide and N-hydroxybenzotriazole ester methods were used for chemical activation of the phosphate group, and T4 DNA ligase for enzymatic ligation. All the chemical activation methods produced 3',5'- and 2',5'-phosphodiester bonds (40-45 and 55-60%, resp.), whereas enzymatic ligation gave the product only with 3',5'-phosphodiester bond.

Base Sequence↗