[Characterization of a panel of monoclonal antibodies to hepatitis C NS3 recombinant protein ].

Recombinant protein rNS3 imitating helicase region (1356-1459 amino acid residues) of hepatitis C virus (HCV) was expressed in E. coli cells and used for BALB/c mice immunization. Seven hybrydoma clones producing monoclonal antibodies (MAbs) to rHS3 were obtained.

[Preparation and purification of a polypeptide containing antigenic determinants of hepatitis C core protein].

A method has been developed for preparing and purifying recombinant polypeptide--hepatitis C virus core protein (HCcoreAg) expressed in E. coli from pFC105-302 plasmid coding for 150 N-terminal amino acids of HCcoreAg in the hybrid from the C-terminal with beta-galactosidase. HCcoreAg was purified by ion-exchange chromatography on P-11 phosphocellulose.

[Immunochemical properties and specificity of monoclonal antibodies against N- and C-terminal sites of the recombinant protein of the nucleocapsid of hepatitis C virus (HCcAg)].

Highly affine murine monoclonal antibodies (MAB) to recombinant nucleocapsid (core) protein of hepatitis C virus (rHCcAg) expressed in E. coli were obtained. The MABs were analyzed by solid-phase enzyme immunoassay (EIA), immunodot, immunoblotting, and competitive immunochemical analysis.

[Heterologous epitopes in the central part of the hepatitis B virus core protein].

A set of plasmids was constructed determining synthesis of hybrid proteins with insertions of antigenic determinants of preS1 and preS2 regions of HBV in the middle part of HBcAg. The polypeptides containing the 31-36 or monomeric 94-105 preS1 epitopes were water-soluble and formed particles similar to the 27-nm ones of native HBcAg, possessing antigenic properties of both HBcAg and the inserted epitope, while the hybrids containing 133-143 of preS2 or a trimeric form of the 94-105 preS1 epitope became membrane-bound. Another hybrid encoded by plasmid pPS2M31 contained two regions of HBcAg modification: insertion of amino acids 133-143 a.

[Introduction of heterologous epitopes at the N-terminal part of the hepatitis B core protein].

For investigation of proteins possessing assigned immunological properties, plasmids pPS31-42, pPS1-5, pPS2-17, and pPS1P-30 were constructed encoding the hepatitis B core protein (HBcAg) with N-terminally inserted immunodominant epitopes of preS regions (amino acids 31-36 or 94-105 of preS1, or 133-143 of preS2). Analysis of the hybrid proteins with the use of ELISA and immunoelectron microscopy showed that the insertions did not prevent specific aggregation of the protein molecules, the inserted sequences being exposed on the surface of the particles obtained, and both HBcAg and the corresponding preS determinants were antigenically active.

[Localization of the immunodominant site in the hepatitis B virus core antigen using synthetic peptides].

Based on theoretical analysis of secondary structure and hydrophilicity data, the region (residues 73-89) of the HBV core-antigen presumably containing the antigenic determinant has been revealed. The epitope mapping of this region with the use of synthetic peptides, obtained by the pin technology and solid phase method, was carried out. Peptides were synthesized in two variants with different amino acids residues in the position 80 (Ala, Ile).

[Synthesis and antigenic activity of peptides from the C-terminal part of the NS4-protein of the hepatitis A virus].

A set of four peptides from the HCV NS4-protein C-terminal region (aa 1921-1940) were obtained by solid-phase synthesis using activated esters and symmetrical anhydrides of Boc-amino acids. Peptide 1921-1940 has demonstrated a positive reaction in ELISA with individual anti-HCV-positive sera from patients with acute and chronic hepatitis C (80% and 56%, respectively). We analysed the antigenic properties of the peptide 1921-1940 and its fragments and suggested at least two antibody recognizing sites to be contained in this region.

[Synthesis and antigenic activity of peptides of the nucleocapsid protein of the hepatitis delta virus].

Hepatitis delta virus (HDV), a recently discovered infectious agent, participates in severe, often lethal forms of acute and chronic hepatitis and liver cirrhosis. Based on theoretical analysis of secondary structure, hydrophilicity and acrophilicity data, several regions of HDV antigen, presumably containing B-epitopes, have been revealed and the corresponding peptides have been synthesized by the solid phase method. All the peptides obtained reacted with the respective antipeptide rabbit sera.

[Synthesis of peptides of the preS1-region of the viral hepatitis B envelope protein and localization of antigenic determinants].

We synthesized the 24-41, 30-36, 31-36, 24-30 fragments of the preS1-region of the hepatitis B (subtype ayw) envelope. The peptides were prepared by the solid phase synthesis on perfluorpolyethylene polymer grafted with polystyrene. The peptide chains were elongated from C-terminus using activated esters and symmetrical anhydrides of Boc-amino acids, cleaved off the solid phase by HBr or TFMSA in TFA, purified by gel filtration, and, after conjugation with protein carriers, inoculated into test animals.

[Construction and properties of a hybrid operon coding the HBcAG and beta-galactosidase of the hepatitis B virus in Escherichia coli].

A set of plasmids was constructed, that carries the hybrid operons with an artificial region for translation initiation of the second cistron. The SD-sequence situated close to the termination signal of the previous cistron facilitates the reinitiation of translation. Both HBcAg and beta-galactosidase coding cistrons are functionally active.

[Synthesis and immunological properties of the peptide fragment of the hepatitis B virus envelope preS-protein].

We have synthesized the 24-41 fragment of the preS region of the hepatitis B (subtype ayw) envelope. The peptide was prepared by the solid phase synthesis on perfluoropoly-ethylene polymer grafted with polystyrene. The peptide chain was elongated from C-terminus using pentafluorophenyl- and p-nitrophenyl esters of Boc-amino acids.

[Plasmids containing 2 new variants of the E.coli lacZ gene].

New plasmids containing partially deleted lacZ genes were obtained. These genes determine high-level synthesis of polypeptides of molecular mass 43-45 and 49-51 kD under the control of the lambda phage PR-promoter; inspite of the deletion, E. coli cells carrying new plasmids were found to possess beta-galactosidase activity.

[Correlation between the effectiveness of translation initiation and secondary structure of mRNA in the hybrid gene cro-lacIZ].

A set of plasmids containing short DNA deletions in the N-part of cro-lacIZ coding zone as constructed using Bal31 nuclease. Development of models of mRNA secondary structure was carried out stepwise beginning from the 5'-end by taking into consideration the hairpins first formed during mRNA synthesis. Comparison of the results of mRNA secondary structure determination and protein production analysis demonstrated a correlation between the efficiency of translation initiation and the appearance of a single-stranded region upon disruption of the mRNA local secondary structure in the translation initiation zone generated by ribosomes.

[The Shine-Dalgarno sequence and the effectiveness of translation initiation].

On the basis of theoretical analysis of different mRNAs secondary structure it is suggested that the efficiency of procaryotic translation initiation depends to a great extent on the possibility to generate a single-stranded region around the initiation codon. The local disruption of the mRNA secondary structure is mostly determined by interaction according to Shine--Dalgarno of 16S rRNA with the complementary mRNA region. Other mechanisms of single-stranded region generation in the initiation zone of mRNA are discussed.

[Recognition site in the restriction endonuclease Eco CK].

5'-Terminal nucleotide was degenerated in the fragments produced by means of hydrolysis with vestrictase Eco CK. Analysis of the nucleotide sequence, carried out in four DNA fragments after the enzymatic hydrolysis' enabled to detect the only one common 13 residues nucleotide sequence, which is apparently involved in the recognizing site for vestrictase Eco CK: (A/T) (A/T) N (A/T) CGCNCNNNG. This sequence was not found in several DNA molecules with well-known primary structure, stable to the action of this enzyme.