MECHANISM OF RESISTANCE OF HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 TO 2',3'-DIDEOXYINOSINE

A molecular done containing the wild-type reverse transcriptase (RT) c oding region of human immunodeficiency virus type 1 (HIV-1) was constr ucted, and site-directed mutagenesis was used to introduce mutations-L eu74 --> Val (L74V), T215Y, and the combination L74V/T215Y-into the RT coding region. The proteins were purified by immunoaffinity chromatog raphy. Assays were performed with mutant and wild-type RT to determine substrate and inhibitor specificity. All three mutant enzymes catalyz ed the incorporation of substrate 2'-deoxynucleoside 5'-triphosphates (dNTPs) as efficiently as wild-type HIV-1 RT. Small changes were obser ved in the K(m) values for dNTPs with all three mutant enzymes, while more significant changes were noted in sensitivity to nucleoside 5'-tr iphosphate analogues that inhibit the enzyme activity. Results suggest that altered substrate recognition by the HIV-1 RT is involved in the mechanism of resistance.