Table 3.

Summary of pre-steady-state burst kinetics results for hpol κ and mutants. Each reaction was performed under enzyme-limiting conditions by rapid mixing of 100 nM 24/36-mer DNA substrate and hpol κ with 1 mM dNTP in the presence of 50 mM Tris-HCl buffer (pH 7.5), 3 mM DTT, 50 μg BSA ml⁻¹, and 5% (v/v) glycerol at 23 °C. The final enzyme concentrations after mixing for each set of reaction were 25 nM for WT hpol κ, 25 nM for W214Y/W392H, 15 nM for Y50W, and 14 nM for T408W. Results were obtained by fitting data to Eq. 1. (see Fig. 2)

hpol κ	Template base:dNTP pair	kobs s−1	kss s−1	Aa nM	[pol-DNA]active/[pol]total%
WT	G:C	2.1 ± 0.5	0.027 ± 0.012	17 ± 2	68
	8-oxoG:C	3.0 ± 0.7	0.034 ± 0.004	6 ± 1	24
	8-oxoG:A	2.2 ± 0.3	0.028 ± 0.006	10 ± 1	40
W214Y/W392H	G:C	5.9 ± 1.4	0.019 ± 0.002	16 ± 1	64
	8-oxoG:C	3.2 ± 1.4	0.024 ± 0.002	8 ± 1	32
	8-oxoG:A	1.9 ± 0.7	0.014 ± 0.004	14 ± 1	56
Y50W	G:C	4.7 ± 1.0	0.033 ± 0.005	8.6 ± 0.6	57
	8-oxoG:C	3.1 ± 0.8	0.029 ± 0.004	5.4 ± 0.6	36
	8-oxoG:A	3.8 ± 1.0	0.027 ± 0.005	6.4 ± 0.6	42
T408W	G:C	3.1 ± 0.7	0.048 ± 0.004	5.7± 0.5	41
	8-oxoG:C	0.82 ± 0.14	0.014 ± 0.001	4.8 ± 0.4	34
	8-oxoG:A	2.3 ± 0.7	0.019 ± 0.005	4.8 ± 0.6	34

^aA is the product amplitude.