Evidence of interactions between the nucleocapsid protein NCp7 and the reverse transcriptase of HIV-1

The human immunodeficiency virus (HIV-1) nucleocapsid protein NCp7 containi ng two CX2CX4HX4C-type zinc fingers was proposed to be involved in reverse transcriptase (RT)-catalyzed proviral DNA synthesis through promotion of tR NA(3)(LYS) annealing to the RNA primer binding site, improvement of DNA str and transfers, and enhancement of RT processivity. The NCp7 structural char acteristics are crucial because mutations altering the finger domain confor mation led to noninfectious viruses characterized by defects in provirus in tegration. These findings prompted us to study a putative RT/NCp7 protein-p rotein interaction. Binding as says using far Western analysis or RT immobi lized on beads clearly showed the formation of a complex between NCp7 and R T. The affinity of NCp7 for p66/p51RT was 0.60 mu M with a 1:1 stoichiometr y. This interaction was confirmed by chemical cross-linking and co-immunopr ecipitation of the two proteins in a viral environment. Competition experim ents using different NCp7 mutants showed that alteration of the finger stru cture disrupted RT recognition giving insights into the loss of infectivity of corresponding HIV-1 mutants. Together with structural data on RT, these results suggest that the role of NCp7 could be to enhance RT processivity through stabilization of a p51-induced active form of the p66 subunit and o pen the way for designing new antiviral agents.