Table 2.

Pre-steady-state kinetic analysis of wild-type and mutant T4 DNA polymerases with double-stranded 2AP-DNA substrate

Enzyme	Rate-limiting step before hydrolysis*
	1		2
	k1, s−1	a1	k2, s−1	a2
Wild type	3.6  ±  0.2	0.8	20.1  ±  2.7	0.2
G255S	0.34  ±  0.01†	0.6	14.9  ±  1.8†	0.4
D131N	0.5  ±  0.1	0.6	21.6  ±  2.9	0.4
D131G			0.02  ±  0.01	1.0

^*Rate constants k₁ and k₂ were determined from double-exponential curve fits of time courses measuring the increase in fluorescence intensity in proofreading reactions with the wild-type, G255S-, or D131N-DNA polymerase and the G+C-rich 2AP-DNA substrate. The relative amplitudes for the two rates are designated a₁ and a₂, respectively. The time course for the D131G-DNA polymerase was best described by a single-exponential equation, and the corresponding rate constant is given for comparison in the k₂ column. 

^†Data are from Marquez and Reha-Krantz (4).