Deoxynucleotide polymerization by HIV-1 reverse transcriptase is terminated by site-specific styrene oxide adducts after translesion synthesis

In an effort to integrate an understanding of human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) structure with its function, the action of HIV-1 RT was examined in vitro on DNA templates modified with a model bulky DNA adduct. Styrene oxide was site-specifically and stereospecifically coupled to the N⁶ position of adenine to form six different adducted templates. Primer extension assays were conducted under conditions defining both single and multiple encounters between the polymerase and the damaged template-primer. The extent of polymerization observed for each adduct was found to depend on both the chirality of the damage and the lesion sequence context. When HIV-1 RT polymerization was limited by single encounters with damaged DNA, the SO lesions were readily bypassed as evidenced by minimal pausing at the adducted base. However, RT replication of all SO-modified templates was significantly terminated 3-5 nucleotides after translesion synthesis but before reaching the end of the template. These truncated products could be readily extended when additional encounters between enzyme and template-primer were permitted. A model is presented to explain these results in the context of HIV-1 RT structure during DNA replication.