ANTIGENIC STRUCTURE OF HEPATITIS A VIRUS

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Antigenic Structure of Hepatitis A Virus. Hepatitis A virus (HAV) is a member of the picornavirus family but is distantly related to other viruses in that family and will be classified into its own genus. While the virus has been cloned and sequenced in several laboratories, useful peptide or polypeptide antigens have not been produced. We have used 2 techniques to study the antigenic structure of HAV. A cDNA expression library consisting of random fragments of HAV capsid coding region was produced in lambda gt11. This library has been screened with over 20 neutralizing antibodies for HAV but no clones were recognized by these antibodies. On the other hand, when this library was screened with antibodies that were directed against synthetic oligopeptides representing regions of each of the 4 capsid proteins, clones were easily detected. This indicates that the antigens recognized by the monoclones were non-linear or conformation dependant. Sequence analysis (done in other laboratories) of neutralization escape mutants of HAV have revealed 2 potential antigenic sites. The first is around the 70th amino acid of VP3 (370) and the second around VP1 105. Using site directed mutagenesis we have produced a total of 30 mutants in and around predicted antigenic sites. Ten of these mutants to date are viable and are being analyzed. Thus far we have confirmed the 370 site and produced a mutant at that position that, unlike the antibody selected mutants, is stable to reversion. We have also extended the VPl site to amino acid 114 (1114). We have initiated a project to study T-cell immunity to HAV in the chimpanzee model. T-cell colones are being developed which will eventually aid in the analysis of the T cell epitopes.