Table 1.

V_max, K_m, and k_cat Values for Incorporations of TTP and AZTTP, dCTP and ddCP, or dATP and EFdATP by HIV-1 RT at 6 and 0.25 mM Mg²⁺

conditiona	[RT] (nM)	Vmax (nM/min)	kcatb (min−1)	Km (μM)	kcat/Kmc (x-fold decrease)	p valued
6 mM Mg2+ dTTP	0.8	1.1 ± 0.1	1.3 ± 0.09	2.3 ± 0.2	0.57	–
0.25 mM Mg2+ dTTP	0.8	0.65 ± 0.07	0.81 ± 0.09	5.7 ± 0.7	0.14 (4.1)	<0.001
6 mM Mg2+ AZTTP	0.8	0.95 ± 0.08	1.2 ± 0.09	2.3 ± 0.4	0.52	–
0.25 mM Mg2+ AZTTP	0.8	0.26 ± 0.01	0.32 ± 0.01	12 ± 1	0.027 (19)	<0.001
6 mM Mg2+ dCTP	2	2.1 ± 0.1	1.0 ± 0.1	1.3 ± 0.4	0.77	–
0.25 mM Mg2+ dCTP	2	1.4 ± 0.1	0.68 ± 0.03	3.9 ± 1.7	0.17 (4.5)	0.05
6 mM Mg2+ ddCTP	0.5	1.4 ± 0.2	0.72 ± 0.08	5.0 ± 0.3	0.14	–
0.25 mM Mg2+ ddCTP	0.5	0.23 ± 0.05	0.12 ± 0.03	12.5 ± 3.5	0.01 (18)	<0.001
6 mM Mg2+ dATP	0.8	2.1 ± 0.2	2.6 ± 0.3	2.2 ± 0.8	1.2	–
0.25 mM Mg2+ dATP	0.8	0.72 ± 0.08	0.90 ± 0.09	2.5 ± 1.1	0.36 (3.3)	0.1
6 mM Mg2+ EFdATP	0.8	0.72 ± 0.02	0.90 ± 0.01	1.1 ± 0.1	0.82	–
0.25 mM Mg2+ EFdATP	0.8	0.53 ± 0.27	0.66 ± 0.35	3.5 ± 2.2	0.19 (4.3)	0.1

^aAssays were conducted with a 20-nucleotide template and a 19-nucleotide 5′ end-labeled primer as described previously.³⁷ The single template-directed nucleotide was a T opposite an A, a C opposite a G, or an A opposite a T for AZT, ddCTP, or EFdATP, respectively. All assays used 0.8 nM RT. Results are averages of three experiments ± SD.

^bk_cat was calculated by dividing V_max by the enzyme concentration (0.8 nM).

^cThe x-fold decrease in enzyme efficiency (as judged by k_cat/K_m) compared to the 6 mM result (number above) with the same nucleotide or inhibitor.

^dp values were calculated using a standard Student’s t test for k_cat/K_m values between 0.25 and 6 mM Mg²⁺.