A mechanism of AZT resistance: An increase in nucleotide-dependent primer unblocking by mutant HIV-1 reverse transcriptase

Mutations in HIV-1 reverse transcriptase (RT) give rise to 3'-azido-3'- deoxythymidine (AZT) resistance by a mechanism that has not been previously reproduced in vitro. We show that mutant RT has increased ability to remove AZTMP from blocked primers through a nucleotide-dependent reaction, producing dinucleoside polyphosphate and extendible primer. In the presence of physiological concentrations of ATP, mutant RT extended 12% to 15% of primers past multiple AZTMP termination sites versus less than 0.5% for wild type. Although mutant RT also unblocked ddAMP-terminated primers more efficiently than wild-type RT, the removal of ddAMP was effectively inhibited by the next complementary dNTP (IC₅₀ ≃ 12 μM). In contrast, the removal of AZTMP was not inhibited by dNTPs except at nonphysiological concentrations (IC₅₀ > 200 μM).