CHARACTERIZATION OF THE P68 P58 HETERODIMER OF HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-2 REVERSE-TRANSCRIPTASE/

Recently we demonstrated that the p58 subunit of p68/p58 HIV-2 reverse transcriptase (RT) heterodimer, produced by processing of p68/p68 hom odimer with recombinant HIV-2 protease, terminates at Met(484) [Fan, N ., et al. (1995) J. Biol. Chem. 270, 13573-13579]. Here we describe pu rification and characterization of the p68/p58 heterodimer of recombin ant HIV-2 RT. It exhibited both RT and RNase H activities, obeyed Mich aelis-Menten kinetics, and was competitively inhibited by the DNA chai n terminator ddTTP (K-i[app] = 305 +/- 20 nM). The HIV-2 RT-associated RNase H exhibited a marked preference for RNA hydrolysis from a HIV-1 gag-based heteropolymeric RNA/DNA hybrid in the presence of either Mg 2+ or Mn2+, compared to the [H-3]poly(rA). poly(dT) or [H-3]poly(rG). poly(dC) homopolymeric substrates. Relative to HIV-1 RT, the RNase H a ctivity of HIV-2 RT was only 5% toward the [H-3]poly(rA) poly(dT) in t he presence of Mg2+. The size distribution of products generated from [H-3]poly(rA). poly(dT) by HIV-2 RT-associated RNase H was markedly di stinct from that of HIV-1 RT in the presence of Mg2+ Or Mn2+. The p68/ p58 HIV-2 RT heterodimer, produced by specific cleavage using HIV-2 pr otease, should be useful for inhibition and biophysical studies aimed at discovering and designing drugs directed toward HIV-2.