Figure 3

Rescue DNA polymerization initiated from AZTMP-, d4TMP-, and tenofovir-terminated primers annealed to a DNA template. Reactions were carried out with D38/25PGA or D38T/25PGA complexes (sequences given above). The 25-nucleotide primer (lane P) is first blocked with the nucleotide analogue (lane B). The excision of the inhibitor, followed by extension of the primer is achieved after addition of a mixture containing 3.2 mM ATP and the four dNTPs. A fully extended 38-nucleotide product is formed. The gel on the right shows a representative time course experiment of a primer rescue reaction. Lanes 1 to 9 correspond to aliquots removed 2, 4, 6, 8, 10, 12, 15, 20, and 30 minutes after the addition of 3.2 mM ATP. Graphs of time course experiments of primer rescue reactions initiated from inhibitor-terminated primers are given below. All dNTPs in the assays were supplied at 100 μM, except for dATP or dTTP (depending on the reaction) whose concentration was 1 μM. Template-primer and active RT concentrations in these assays were 30 and 24 nM, respectively. The values (averaged ± standard deviations [error bars]) were obtained from three independent experiments.