TABLE 2.

dTTP hydrolysis activity of wild-type gp4 and the gp4 variants

The rate of dTTP hydrolysis by wild-type gp4 and gp4 variants in the presence or absence of ssDNA is compared. dTTP hydrolysis reactions were performed as described under “Experimental Procedures.” Wild-type gp4 hydrolyzes dTTP approximately 75-fold faster in the presence of 5 nm M13 ssDNA as compared to the absence of DNA. The relative dTTP hydrolysis by the gp4 variants were compared with the wild-type activity. The K_DNA was calculated from the Fig. 2B.

gp4-helicase	No M13 ssDNA				5 nm M13 ssDNA				KDNA
	kcat	Km	kcat/Km	Relative kcat	kcat	Km	kcat/Km	Relative kcat
	s−1	mm	s−1 mm−1		s−1	mm	s−1mm−1		nm
Wild-type	0.24 ± 0.02	0.07 ± 0.02	3.4	100	17.6 ± 1	2.3 ± 0.3	7.6	100	0.14
F523A	0.07 ± 0.01	0.2 ± 0.01	0.35	29	3.9 ± 0.7	0.5 ± 0.3	7.8	22	0.54
F523V	0.12 ± 0.01	0.2 ± 0.03	0.6	50	3.7 ± 0.8	0.5 ± 0.3	7.4	21	0.91
F523I	0.15 ± 0.02	0.3 ± 0.02	0.5	62	12.4 ± 1.2	1.6 ± 0.4	7.7	70	0.29
F523L	0.35 ± 0.03	0.2 ± 0.03	1.75	145	10.8 ± 0.9	1.3 ± 0.3	8.3	61	0.38
F523H	0.12 ± 0.01	0.1 ± 0.01	1.2	50	5.4 ± 0.4	0.5 ± 0.1	10.8	31	0.13
F523Y	0.63 ± 0.04	0.2 ± 0.04	3.15	262	9.9 ± 0.8	1.1 ± 0.3	9	56	0.37