MYRISTYLATION OF THE HEPATITIS-B VIRUS LARGE SURFACE PROTEIN IS ESSENTIAL FOR VIRAL INFECTIVITY

The hepatitis B Virus (HBV) envelope contains equimolar amounts of thr ee viral proteins: the major (S), middle, and large (L) polypeptides. Their roles in the adsorption and penetration of the virus have not ye t been elucidated. We have used a highly efficient in vitro model that permits reproducible HBV infection to investigate whether N-myristyla tion, a posttranslational modification of the L protein, was essential for viral infectivity. A point mutation abolishing myristylation was introduced into the HBV genome. Mutant virions were produced by transf ecting viral DNA into hepatoma cells and their infectivity was evaluat ed on primary human hepatocyte cultures. No difference between mutant and wild-type viral RNA production could be observed. Furthermore, int ermediate DNA replicative forms were observed in transfected cells dem onstrating replication competence of mutant viral genomes. In addition , complete Virions were produced in the cell supernatant However, we f ound that mutant viral particles contained viral DNA with a reduced me an size, probably corresponding to a larger single-stranded region in the relaxed circular DNA form. We have evidenced the presence of pre-S i, pre-SP, and S epitopes at the outer surface of these virions by usi ng immunoprecipitation with specific monoclonal antibodies. This resul t confirmed that mutant viruses were normally assembled. By contrast, myristylation-defective mutants completely lost their infectivity for human hepatocytes in primary cultures as shown by the absence of HBs a ntigen production and viral intermediate replicative forms in hepatocy tes. In conclusion, the myristylation of the L protein is not required for the production of Dane-like particles but it is absolutely necess ary for HBV infectivity. (C) 1995 Academic Press, Inc.