Polymerization activity and AZT resistance of SFVmac PR-RT variants. (A). SFVmac PR-RT variants used in this study harboring one to four of the relevant resistance substitutions are highlighted by red rectangles. (B) Polymerization initiation. Initiation of polymerization was checked with 20 nM 5′ 32P-P30/T50 substrate, 250 μM dNTPs and 10 nM enzyme at 37°C for 10 min. The PR-RTs used are indicated on top of the gel. WT, wild type; Control, sample without enzyme. (C) AZTMP removal in the presence of ATP. 5 nM of an AZTMP-terminated and 5′ 32P-labeled substrate P30-AZTMP/T50, 0.02 U of pyrophosphatase and 5 mM ATP were preincubated for 5 min. Reactions were started by the addition of 320 nM enzyme as indicated on top and stopped after 20 min. Reaction products were separated on 10% sequencing gels and visualized by phosphoimaging. P30-AZT, AZTMP terminated primer; P30, unblocked primer.