THE FIDELITY OF MISINSERTION AND MISPAIR EXTENSION THROUGHOUT DNA-SYNTHESIS EXHIBITED BY MUTANTS OF THE REVERSE-TRANSCRIPTASE OF HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-2 RESISTANT TO NUCLEOSIDE ANALOGS

The AIDS-causing retroviruses, human immunodeficiency virus types 1 an d type 2 (HIV-1 and HIV-2, respectively) undergo extensive genetic var iations, which effect their pathogenesis and resistance to drug therap y. It was postulated that this genetic hypervariability results From h igh rates of viral replication in conjugation with a relatively low fi delity of DNA synthesis [typical to the reverse transcriptases (RT) of these retroviruses]. As part of studying structure/function relations hip in HN RT. mutational analyses were conducted to identify amino aci d residues which are involved in affecting the fidelity of DNA synthes is. The formation of 3'-mispaired DNA due to nucleotide misinsertions, and the subsequent elongation of this mismatched DNA were shown to be major determinants in affecting those substitutions during DNA synthe sis (exhibited in vitro by HIV RT). It was interesting to find a corre lation between sensitivity to nucleoside analogs (due to the ability t o incorporate or reject an incoming analog) and the fidelity of DNA sy nthesis (which depends on the capacity to incorporate and extend a wro ng nucleotide). Such a connection has already been found for several d rug-resistant mutants of HIV-1 RT, with an increased fidelity of DNA s ynthesis relative to the wild-type RT. In the present study we have ex amined the fidelity of DNA synthesis using the same parameters of misi nsertion and mispair extension for five novel drug-resistant mutants o f HIV-2 RT, i.e. the single mutants [Val74]RT, [Gly89]RT and [Tyr215]R T and the double mutants [Val74,Tyr215]RT and [Gly89,Tyr215]RT. This c omparative study suggests that unlike the Val74 mutant of HIV-1 RT, wh ich was shown earlier to display a substantially enhanced fidelity, th e comparable mutant of HIV-2 RT has fidelity similar to that of the wi ld-type RT. Depending on the assay employed and the DNA sequences exte nded, must other mutants of HIV-2 RT display moderate effects on the e nzyme, leading to mild increases in fidelity of DNA synthesis. This im plies a more complex and less distinctive correlation between drug-res istance, misinsertion and mispair extension in HIV-2 RT in contrast to HIV-1 RT, providing evidence for potential biochemical differences be tween these two related RT.