CHARACTERIZATION OF HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 PR55(GAG) MEMBRANE ASSOCIATION IN A CELL-FREE SYSTEM - REQUIREMENT FOR A C-TERMINALDOMAIN

Association of the human immunodeficiency virus type 1 (HIV-1) gag pol yprotein precursor with cellular membranes is necessary for assembly o f virions. We used in vitro synthesized HIV-1 gag to study its associa tion with isolated cellular membranes. Rabbit reticulocyte lysates pro grammed with HIV-1 gag mRNA incorporated [S-35]methionine and [H-3]myr istate into two predominant species of 55 kDa and 40 kDa. Radioimmunop recipitation with HIV-1-specific antibodies suggested that the 55-kDa protein represented the polyprotein precursor (Pr55(gag)), while the 4 0-kDa protein was a mixture of N- or C-terminal truncations of the gag precursor. The Pr55(gag) protein bound to cellular membranes, while t he 40-kDa mixed protein species did not. Membrane binding studies with C terminus-truncated and point mutants revealed that the seven-amino acid sequence located between the two Cys-His arrays in the nucleocaps id region was necessary for stable association to occur. Therefore, we propose that signals in addition to myristate are required for the me mbrane association of HIV-1 gag proteins and that these signals includ e a domain in the nucleocapsid protein.