Binding properties of the human immunodeficiency virus type 1 nucleocapsidprotein p7 to a model RNA: Elucidation of the structural determinants for function

HIV-1 nucleocapsid protein (NCp7) is a double zinc-fingered protein that ha s been traditionally implicated in viral RNA recognition and packaging, in addition to its tight association with genomic RNA and tRNA primer within t he virion nucleocapsid. The availability of large quantities of viral or re combinant wild-type NCp7 and mutant p7 has made possible the assignment of the different roles that structural motifs within the protein play during R NA binding. At low ionic strength binding to the homopolymeric fluorescent RNA, poly(epsilon A), is electrostatically driven and four sodium ions are displaced. Arg7 in the flanking N-terminal region, Lys20 and Lys26 in the f irst zinc finger and one positively charged residue (attributed to Lys41) i n the second zinc finger are involved in electrostatic contacts with RNA. T he p7 zinc fingers do not function independently but concomitantly. The fir st zinc finger (both isolated or in the context of the full-length protein) has a more prominent electrostatic interaction than the second one. The se cond zinc finger dominates the non-electrostatic stabilization of the bindi ng to RNA due to stacking of its Trp residue with nucleic acid bases. Mutat ions in the highly conserved retroviral Zn-coordinating residues (CCHC) to steroid hormone receptor (CCCC) or transcription factor (CCHH) metal cluste r types do not affect RNA binding. Ln spite of the limited impact in RNA bi nding affinity in vitro or RNA packaging in vivo that such mutations or str uctural alterations impart, they impair or abolish virus infectivity. It is likely that such an effect stems from the involvement of NCp7 in crucial s teps of the virus life cycle other than RNA binding. (C) 1999 Academic Pres s.