Table 4.

Pre-steady-state kinetic analysis of single-turnover DNA polymerase assays under enzyme excess conditions. Data were obtained by rapid mixing of 1.5 μM Y50W, 50 nM 24/36-mer DNA substrate, 3 mM DTT, 50 μg BSA ml⁻¹, and 5% (v/v) glycerol with dNTP concentrations from 0.1 to 200 μM. Actual results are shown in Fig. 7C. Data were fit to Eq. 2 to extract the maximal rate constant for single nucleotide incorporation (k_pol) and the apparent constant of nucleotide dissociation from kinetically active ternary complex (K_d,dNTP,app).

hpol κ	Template base:dNTP pair	kpol s−1	Kd,dNTP,app μM	kpol/ Kd,dNTP,app μM−1 s−1
WTa	G:C	10 ± 1	36 ± 14	0.28
	8-oxoG:C	0.40 ± 0.04	90 ± 35	0.0044
	8-oxoG:A	8.2 ± 0.2	13 ± 3	0.63
Y50W	G:C	1.4 ± 0.1	1.0 ± 0.1	1.5
	8-oxoG:C	1.3 ± 0.2	17 ± 7	0.076
	8-oxoG:A	1.6 ± 0.2	0.81 ± 0.39	2.0

^aData were from ref. [19].