The genome of the AIDs retrovirus (HIV), the etiologic agent of AIDS, codes for several polypeptides in an extremely complex scheme of mRNA biogenesis. We have used kinetic analysis of viral proteins using defined antisera, purified and sequenced several viral proteins both from extracellular virus and cellular lysates. Where possible, we have tried to correlate in vivo protein synthesis with cell free translation of selected viral mRNAs. To study the functional significance of different viral gene products, we have mutagenized the proviral DNA and studied their effects on viral replication using DNA transfections of animal cells. Using a cell line (8E5) containing a single copy of defective proviral DNA with a premature termination codon in the p64 RT domain of the pol ORF, we successfully identified and sequenced the p64 and p51 RT subunits of pol ORF, as well as the p34 endonuclease mapping near the C-terminus of the pol ORF. We have extended this project and successfully identified and sequenced a p10 gag-pol protease responsible for the processing of the gag and gag-pol fusion proteins. The localization of the N- terminus of the gag protease has allowed us to dissect the steps involved in the frameshifting mechanism responsible for the synthesis of gag-pol fusion proteins. We have also developed a sensitive assay for the protease using defined oligopeptides, and this will enable us to screen several protease inhibitors as potential antiviral drugs. During the studies relating to the mechanisms of gag and gag-pol protein processing, we also identified two forms of a gag precursor protein p41 and are presently trying to determine the functional correlation of a p41 protein initiated internally 141 residues from the authentic AUG codon of the gag ORF.
