FIG. 3.

Kinetics of assembly of WT FMDV 3Dpol-s/s-U complexes. (A) Sequence of s/s-U. (B) AMP incorporation into s/s-U by WT FMDV 3Dpol. Products were resolved by electrophoresis on a denaturing 23% polyacrylamide gel. (C) Concentration dependence of WT FMDV 3Dpol. Reaction mixtures contained 1 μM s/s-U (0.5 μM duplex), 500 μM ATP, 5 mM MgCl₂, and either 0.25 (○), 0.5 (•), 1 (□), or 5 (▪) μM 3Dpol. Reactions were initiated by the addition of 3Dpol, incubated at 30°C, and quenched at the indicated times by the addition of EDTA (50 mM). The solid lines represent the fit of the data to a single exponential. The observed rate of assembly was 0.003 s⁻¹ for all concentrations of 3Dpol. (D) Concentration dependence of s/s-U. Reactions were performed as described above, but the mixtures contained 1 μM 3Dpol and either 1 (•), 2 (○), 4 (▪), 10 (□), or 20 (▴) μM s/s-U. The solid lines represent the fit of the data to a single exponential. The observed rates of assembly were 0.004, 0.004, 0.002, 0.002, and 0.004 s⁻¹ for 20, 10, 4, 2, and 1 μM s/s-U, respectively (average, 0.003 s⁻¹).