THE K65R MUTANT REVERSE-TRANSCRIPTASE OF HIV-1 CROSS-RESISTANT TO 2',3'-DIDEOXYCYTIDINE, 2',3'-DIDEOXY-3'-THIACYTIDINE, AND 2',3'-DIDEOXYINOSINE SHOWS REDUCED SENSITIVITY TO SPECIFIC DIDEOXYNUCLEOSIDE TRIPHOSPHATE INHIBITORS IN-VITRO

The K65R mutation in human immunodeficiency virus type 1 (HIV-1) rever se transcriptase (RT) encodes cross-resistance to 2',3'-dideoxycytidin e (ddC), 2',3'-dideoxy-3'-thiacytidine (3TC), and 2',3'-dideoxyinosine (ddI). We characterized the in vitro sensitivities of recombinant wil d type (wt) and K65R mutant RT to dideoxynucleoside triphosphate (ddNT P) inhibitors, using a variety of primer-templates, With poly(rA)-olig o(dT), the K65R mutant showed slight increases in K-i for ddTTP and 3' -azido, 3'-deoxythymidine triphosphate (AZTTP) compared to wt RT, but neither wt nor K65R RT was inhibited by ddCTP or ddATP. With poly(rI)- oligo(dC), the K65R mutant showed a 2-fold increase in Ii;, for dCTP a nd a 20-fold increase in K-i for ddCTP compared to wt, whereas ddATP, ddTTP, and AZTTP failed to inhibit either enzyme. With a heteropolymer ic primer-template, the K65R mutant showed 10-fold reduced sensitiviti es to ddCTP, 3TCTP, and ddATP, and 4-fold reduced sensitivity to AZTTP , compared to wt. In contrast, both enzymes were equally inhibited by ddTTP and ddGTP. HIV-1 cross-resistance to ddC/3TC/ddI resulting from the K65R mutation may therefore involve selective alterations in subst rate/inhibitor recognition. Additionally, competitive inhibition by dd NTPs noncomplementary to the template base appears to be unimportant i n the mechanism of inhibition of HIV-1 RT by dideoxynucleoside analogs .