CHARGED AMINO-ACID-RESIDUES OF HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 NUCLEOCAPSID P7 PROTEIN INVOLVED IN RNA PACKAGING AND INFECTIVITY

Interaction of the human immunodeficiency virus type 1 (HIV-1) Gag pre cursor polyprotein (Pr55(Gag)) with the viral genomic RNA is required for retroviral replication. Mutations that reduce RNA packaging effici ency have been localized to the highly basic nucleocapsid (NC) p7 doma in of Pr55(Gag), but the importance of the basic amino acid residues i n specific viral RNA encapsidation and infectivity has not been thorou ghly investigated in vivo. We have systematically substituted the posi tively charged residues of the NC domain of Pr55(Gag) in, an HIV-1 vir al clone by using alanine scanning mutagenesis and have assayed the ef fects of these mutations on virus replication, particle formation, and RNA packaging in vivo, Analysis of viral clones with single substitut ions revealed that certain charged amino acid residues are more critic al for RNA packaging efficiency and infectivity than others, Analysis of viral clones with multiple substitutions indicates that the presenc e of positive charge in each of three independent domains - the zinc-b inding domains, the basic region that links them, and the residues tha t flank the two zinc-binding domains - is necessary for efficient HIV- 1 RNA packaging. Finally, we note that some mutations affect virus rep lication more drastically than RNA incorporation, providing in vivo ev idence for the hypothesis that NC p7 may be involved in aspects of the HIV life cycle in addition to RNA packaging.