Figure 4.

Elemental effect on correct and incorrect nucleotide incorporation. A, a preincubated solution of hPolε exo− (100 nm, UV concentration) and 5′-radiolabeled D-1 DNA substrate (20 nm) was rapidly mixed with 5 μm dTTP (●) or S_p-dTTPαS (○) and Mg²⁺ for varying incubation times before quenching with the addition of EDTA. Product concentration was plotted against time and the data were fit to Equation 5 to yield fast rate constants of 127 ± 15 s⁻¹ and 118 ± 21 s⁻¹ for dTTP and S_p-dTTPαS, respectively, resulting in an elemental effect of 1.1. B, the elemental effect on incorrect dATP incorporation was tested under identical conditions, except the reaction was initiated with the addition of 500 μm dATP (●) or S_p-dATPαS (○) and Mg²⁺. Product concentration was plotted against time and the data for dATP incorporation were fit to Equation 6, whereas the data for S_p-dATPαS were fit to a line. The incorporation rate constants were 0.073 ± 0.007 s⁻¹ for dATP and 0.0016 s⁻¹ for S_p-dATPαS, giving an elemental effect of 46.