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'-deox ythymidine (AZT) resistance by a mechanism that has not been previously rep roduced in vitro. We show that mutant RT has increased ability to remove AZ TMP from blocked primers through a nucleotide-dependent reaction, producing dinucleoside polyphosphate and extendible primer. In the presence of physi ological concentrations of ATP, mutant RT extended 12% to 15% of primers pa st multiple AZTMP termination sites versus less than 0.5% for wild type. Al though mutant RT also unblocked ddAMP-terminated primers more efficiently t han wild-type RT, the removal of ddAMP was effectively inhibited by the nex t complementary dNTP (ICS50 approximate to 12 mu M). In contrast, the remov al of AZTMP was not inhibited by dNTPs except at nonphysiological concentra tions (ICS50 > 200 mu M).