Figure 5. Site-directed mutagenesis study of the HIV-1 RT p66 subunit with NRTI resistance.

Single substitutions of amino acids around the predicted ATP-binding site in Figure 3 were introduced into the p66 chain of ERT-mt6. The overall DNA polymerization activity (Figure S4B), IC₅₀ of AZTTP (Figure S4C), and K_m, k_cat, K_i^ATP, and K_i′ ^ATP values were measured using the [α-³²P]dTTP and poly (rA)·p(dT)_12-18 system, and fold increases in the K_m (A), k_cat (B), K_i^ATP (C), and K_i′ ^ATP values (D) compared to those for the ERT-mt6 were calculated. Results for the RT mutants, D113A, D113N, K219Q, and K219A, which retained sufficient polymerization activity for a kinetic study, are shown.