Relationship between human immunodeficiency virus type 1 Gag multimerization and membrane binding

The human immunodeficiency virus type I (HIV-I) Gag precursor, Pr55(Gag), i s necessary and sufficient far the assembly and release of viruslike partic les. Binding of Gag to membrane and Gag multimerization are both essential steps in virus assembly, get the domains responsible for these events have not been fully defined. In addition, the relationship between membrane bind ing and Gag-Gag interaction remains to be elucidated. To investigate these issues, we analyzed, in vivo, the membrane-binding and assembly properties of a series of C-terminally truncated Gag mutants. Pr-55Gag was truncated a t the C terminus of matrix (MAstop), between the N- and C-terminal domains of capsid (CA146stop), at the C terminus of capsid (p41stop), at the C term inus of p2 (p43stop), and after the N-terminal 35 amino acids of nucleocaps id (NC35stop). The ability of these truncated Gag molecules to assemble and release viruslike particles and their capacity to copackage into particles when coexpressed with full-length Gag were determined. We demonstrate that the amount of truncated Gag incorporated into particles is incrementally i ncreased by extension from CA416 to NC35, suggesting that multiple sites in this region are involved in Gag multimerization, Using membrane Botation c entrifugation, we observe that MA shows significantly reduced membrane bind ing relative to full-length Gag but that CA146 displays steady-state membra ne-binding properties comparable to those of Pr55(Gag). The finding that th e CA146 mutant, which contains only matrix and the N-terminal domain of cap sid, exhibits levels of steady-state membrane binding equivalent to those o f full-length Gag indicates that strong Gag-Gag interaction domains are not required for the efficient binding of HIV-1 Gag to membrane.