ENZYMATIC-PROPERTIES OF 2 MUTANTS OF REVERSE-TRANSCRIPTASE OF HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 (TYROSINE-181 -] ISOLEUCINE AND TYROSINE-188 -] LEUCINE), RESISTANT TO NONNUCLEOSIDE INHIBITORS

A number of structurally diverse compounds have been shown to be poten t inhibitors of the DNA polymerase activity of human immunodeficiency virus (HIV-1) reverse transcriptase (RT). The compounds can be grouped into two broad classes: nucleoside analogs and nonnucleoside inhibito rs. The nonnucleoside inhibitors are quite specific for the polymerase activity of HIV-1 RT; they do not affect the polymerase activity of H IV-2 RT or the ribonuclease H (RNase H) activity of either HIV-1 RT or HIV-2 RT. Structural, biochemical, and genetic analyses showed that t his group of inhibitors binds in a hydrophobic pocket near the polymer ase active site. Mutations in amino acids that line this hydrophobic p ocket, for example at tyrosine 181, tyrosine 188, or lysine 103, lead to enzymes that are resistant to the nonnucleoside inhibitors. We have investigated the enzymatic properties of two mutants of HIV-1 RT in w hich residues 181 and 188 were replaced by the corresponding amino aci ds in HIV-2 RT (tyrosine 181 --> isoleucine and tyrosine 188 --> leuci ne). The two tyrosine mutants closely resemble the wild-type HIV-1 RT in almost all the catalytic functions tested, including the heat stabi lity, sensitivity of the DNA polymerase activity to inhibition by deox ynucleoside analogs, inhibition by the zinc chelator o-phenanthroline, and the K-m values calculated for the DNA polymerase activity. There is, however, a slight difference in the effect of orthophenanthroline on the RNase H activity. In addition, there is a subtle disparity in t he fidelity of DNA synthesis (analyzed by a mispair extension assay), thus indicating that these mutant RTs are not likely to confer any sel ective advantages or disadvantages to the variant virions over wildtyp e virus.