STRUCTURE OF HEPATITIS B ANTIGENS

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The objective is to obtain detailed information concerning the structure of hepatitis B viral proteins, and to determine the relationship between their protein structure and their antigenic function. These studies will include research on the HBsAg, including the pre-S gene products, HBeAg, HBcAg and the "X" protein. Studies of the HBsAg will include studies of the intact particle and its protein and lipid components. We will utilize our panel of monoclonal anti-HBs antibodies (which contain anti-group specific and anti-subtypes specific antibodies) in conjunction with amino acid sequence analysis, chemical modification studies, synthetic peptides, immunoblotting techniques, olignucleotide directed site, specific mutagenesis, and eukaryotic expression vectors for the production of HBsAg from the cloned viral gene to attempt to determine the physical structure of HBsAg and the location and structure of the various antigenic determinants. We will also use physical techniques such as freeze ETCH and freeze fracture electron microscopy low angle x-ray scattering, fourier transform infrared spectorphotometry, and circular dichroism to gain information on the number, arrangement, and secondary structure of the protein subunits with HBsAg and how the lipid components are related to the maintenance of this structure. The role of the pre-S proteins in the particle structure and antigenic activity will also be investigated. The chemistry of the conversion of HBcAg to HBeAg will be studied and this chemical conversion correlated with the antigenic alteration. The location of the relevant antigenic sites will be sought by the same methods as used for HBsAg. We will also use anti-synthetic peptide antibodies in an attempt to characterize the product of the viral "X" gene. Mammalian expression vectors will be used in an effort to synthesize the protein for further characterization. The antibody produced in humans in response to HBV infection or to immunization by the current vaccine (Heptavax) as well as that produced in response to the experimental yeast HBsAg vaccines will be examined by competition studies with our monoclonal antibodies to determine the relative amounts and specificities of these human antibodies. It is expected that this will help define the major immunogenic determination recognized by humans.