[TiO2-DNA nanocomposites capable of penetrating into cells].

Methods of noncovalent immobilization of DNA fragments onto titanium dioxide nanoparticles (TiO2) were developed, which led to TiO2-DNA nanocomposites capable of penetrating through cell membranes. TiO2 nanoparticles of different forms (amorphous, anatase, brookit) with enhanced agglomeration stability were synthesized. The particles were characterized by X-ray diffraction, small angle X-ray scattering, infrared spectroscopy and atomic force microscopy.

[Cloning of Escherichia coli K12 xylose isomerase (glucose isomerase) and studying the enzymatic properties of its expression product].

The coding region of Escherichia coli K12 xylose (glucose) isomerase gene was inserted into the pRAC expression vector and cloned in E. coli BL21 (DE3) cells. After induction of expression of the cloned gene, the proportion of recombinant xylose isomerase accounted for 40% of the total protein content.

An examination of the ability of titanium dioxide nanoparticles and its conjugates with oligonucleotides to penetrate into eucariotis cells.

In this study we examine the possibility that TiO nanoparticles and their conjugates can penetrate into cultivated cells without any special transfection procedures. Oligonucleotides and their derivates were conjugated with the TiO nanoparticles, which were obtained as colloidal solutions at a concentration of TiO 0.3M by TiCl hydrolysis.

[Genetic polymorphism of Commander Islands polar foxes Alopex lagopus semenovi Ognev, 1931 and Alopex lagopus beringiensis Merriam, 1902].

The polymorphism of the mitochondrial DNA (mtDNA) control region sequence was examined in 30 polar foxes from Bering Island and 30 polar foxes from Mednyi Island. Seven haplotypes were revealed in polar foxes from Bering Island, and one, in polar foxes from Mednyi Island. The age of divergence of these populations (12 000 +/- 600 years) was calculated based on a fragment of the D-loop.

[Consumption of hydrocarbons by psychrotolerant degrader strains].

Oil-oxidizing microorganisms have been sampled in various regions of Siberia and used in strain associations, which degrade n-alkanes of oil from various fields by 64-92% after 6 days of growth in a wide temperature range. These strains are salt-tolerant and psychrotolerant. They are compatible with aboriginal soil microflora.

[Isolation of total RNA from baker's yeast].

A simple method of production of total RNA from baker's yeast was developed. Total RNA was isolated from yeast (Saccharomyces cerevisiae) biomass using lysis with sodium dodecyl sulfate at 100 degrees C for 40-60 min and subsequent precipitation of the target product with 3 M NaCl. The preparation obtained was characterized in detail: yield of total RNA from 1 kg of pressed yeast, 9.

[Influence of the salt concentration in culture on the copy number of plasmid pSH1 in cells of Micrococcus sp. 9].

The study of heterotrophic bacteria isolated from the brackish waters of Lake Shira has shown that some of them contain plasmid pSH1 of approximately 2.7 kb in size. The number of plasmid copies in plasmid-containing strains cultivated at a minimal concentration of sodium chloride is found to be low, whereas the subculturing of these strains at high salt concentrations increases the plasmid number.

[Construction of an expression plasmid vector for accomplishing in vivo delivery of recombinant biologically active proteins. 2. Synthesis of HBcAG in a vaccine strain of Salmonella choleraesuis].

The capacity of a previously described plasmid vector pAZ to deliver bioactive proteins to targets in vivo has been studied. This vector molecule has a strong constitutive promoter, is extremely stable in cells of vaccinal S. choleraesuis strain, and encodes the synthesis of marker protein beta-galactosidase which helps monitor the vector's fate in the host.

[Antigenic determinants of proteins. The humoral immune response].

There exist two opposing points of view on the organization of antigenic determinants of proteins: 1) the determinants are stringently predetermined regions of the protein molecule; 2) all the surface of the protein globule is potentially antigenic, and the immune response is generated occasionally in every individual, although some sites of the protein surface could be preferable for eliciting the immune response. In this work, taking into account the results of recent investigations, we propose some reconciliation: a correlation between the rigidity of the protein architecture and the antigenicity of the particular sites of the antigen molecule. Some general principles of the antigenic determinants of proteins are formulated.

[The principles of creating standard serum panels with a low concentration of HIV-1 antibodies for the quality control of commercial immunoenzyme test systems].

The main approaches are defined to the production of sera with the required concentration of antibodies to HIV by the dilution method and to formation on their basis of serum panels according to theoretical distribution of optic density (OD) values. It was shown to be principally possible to prepare panels of positive sera with low concentration of HIV-specific antibodies in a dry stable form, and their practical importance in the evaluation of the sensitivity of enzyme immunoassays was demonstrated. The evaluation of the quality of commercial test systems is based on the position and shape of the histogram of OD values distribution for panel sera for the controlled test systems.

[In vitro processing of the artificial analog of beta-lipotropin].

The processing of the recombinant analogue of beta-lipotropin (beta-LHP) having 11 additional N-terminal amino acid residues and separated from the hormone by the processing signal, was investigated using rat adrenal secretory granule lysate as a test system of processing "in vitro". It was found that incubation of the beta-LPH analogue with secretory granule enzymes leads to its limited specific degradation with a release of native beta-endorphin. It is concluded that the additional N-terminal amino acids induced no qualitative changes in beta-LPH processing.

[Solid-phase immunoenzyme analysis of apoprotein A-1].

The authors have optimized indirect solid-phase enzyme immunoassay for measuring human blood serum apoA-1. Immunochemical identification of the obtained apoA-1 and specific antibodies has been carried out, working regimens for the components and conjugates employed have been selected. Relationship between the method of preliminary treatment of the serum and the completeness of detected apoA-1 antigenic determinants is demonstrated.

[Effect of poly(dA).poly(dT) sequence inserted into EcoR1 site of pBr322 plasmid on the expression of tet and bla genes].

The 99 base pair poly(dA).poly(dT) stretch was inserted into EcoRI site of pBR plasmid. The effect of this insertion on tet-gene expression from P2 promoter, and bla-gene expression from P1 and P3 promoters was studied.

[Behavior of the large fragment of DNA polymerase I (the Klenow fragment) during fractionation of a cell-free extract of E. coli MRE-600].

Distribution of the DNA polymerase I large fragment (Klenow fragment) was studied during fractionation of the E. coli MRE-600 cell-free extract with polyethylenimine. On the basis of the results obtained a simple procedure is proposed that enables the Klenow fragment to be obtained as a coproduct of DNA polymerase I, RNA polymerase, polynucleotide phosphorylase, nucleotide kinases with acetokinase and nucleoside deoxy-ribosyltransferase in the framework of a combined technological scheme.

[Effectiveness of the promoter Ptac' in vitro and in mini-cells].

The efficiency of hybrid promotor Ptac', comprising a synthetic trp-promoter and lacUV5 "-10" sequence, was studied. By means of electrophoresis and hybridization of RNA-products obtained in vitro and in minicells, with promotor-containing plasmids; the hybrid promotor was found to be 6 times and 3-4 times as efficient as trp and lacUV5 promoters, respectively.

[Isolation and properties of immobilized alkaline phosphatase from E. coli].

Preparations of alkaline phosphatase from E. coli, immobilized on Sepharose, with a specific activity of 40-60 U/g wet weight were obtained. The immobilized enzyme was stable up to 50 degrees C; at higher temperatures it was inactivated.

[Mechanism of reaction catalyzed by RNA-ligase from bacteriophage T4].

The dissociation constants of the complexes of RNA-ligase with acceptors, donors and the adenylylated donor A(5')ppAp have been determined on the basis of the inhibition of ATP-pyrophosphate exchange reaction. The dissociation constants of the complexes of the enzyme with "poor" acceptors (oligouridilates) have been shown to be slightly different from those with "good" acceptors (oligoadenylates). The dependence of the reaction velocity of the formation of ligation products on the concentration of acceptors (pA)4, (pU)4 and the adenylylated donor A(5)ppAp has been studied.

[Distribution of enzymes of nucleic acid biosynthesis and their precursors during fractionation of Escherichia coli MRE-600 extract].

Distribution of the enzymes of template-dependent and template-independent polynucleotide syntheses (DNA-polymerase I, RNA-polymerase, polynucleotide phosphorylase) as well as those of the biosynthesis of nucleic acids precursors (nucleotide kinases, acetokinase and nucleoside deoxyribosyltransferases) during fractionation of Escherichia coli MRE-600 cell extract was studied. On the basis of the results obtained a technological scheme was developed that enabled to combine routine procedures of purification of the above mentioned enzymes.

[Study of ATP-independent stages of reaction catalyzed by phage T4 RNA-ligase].

The isotope exchange between [5'-32P]pAP and A(5')ppAp catalyzed by enzyme was shown not to take place in the absence of the acceptor; i. e. the necessity of the acceptor presence during the second step of the process was demonstrated.

[Expression of the gene for hybrid beta-lactamase pBR322 with C-terminal fragment containing tuftsin (Thr-Lys-Pro-Arg)].

A hybrid beta-lactamase gene with a synthetic tuftsin-coding DNA fragment inserted at the Pst I-site of pBR322 plasmid has been obtained and its expression has been studied. Radioactive amino acids have been used to show that in E. coli chi 925 minicells up to 30% of newly synthesized chimeric protein is secreted into periplasm providing the tuftsin transport.

[Circular transcription map of the plasmid pBR322].

The plasmid pBR322 transcription in the isolated E. coli DNA-dependent RNA-polymerase system was studied. Transcription regions as well as transcripts orientation were defined using both the technique of "criss-cross" hybridization and the annealing of RNA-products with L- and H-strands of plasmid.

[Termination of DNA transcription by a poly(dA).poly(dT) fragment inserted into pBR325 plasmid].

A recombinant DNA was constructed by inserting polynucleotide (dA).(dT) of 80-100 base pairs long into EcoRI site of the pBR325 plasmid DNA. Transcription of this DNA was studied in E.

[Kinetic characteristics of the ATP-pyrophosphate isotope exchange catalyzed by RNA ligase from T4 bacteriophage].

The dependence of initial rate v0 of ATP--PPi exchange reaction catalyzed by RNA-ligase of bacteriophage T4 on the concentration of ATP(s), pyrophosphate (z) and Mgcl2 has been determined. The dependence of v0 on s and z described by the equation v0 = k-1k2E0/(k-1 + K2) (1 + K1/s + k2/z) has been obtained for the reaction of E + S in equilibrium ES in equilibrium E1 + Z, where E--enzyme, E1--adenylylenzyme, S--ATP, Z--pyrophosphate, K1 and K2--constants of equilibrium, k-1, k2--velocity constants of transition of ES to E + S and E1 + Z, E0--complete concentration of enzyme. The low inhibition of the ATP--PPi exchange by the acceptor A(pA)2 and donors pAp, p(Ap)3, pCp has been shown.