TABLE 1.

Kinetic measurement of templateda nucleotide incorporation by pol X polymerases

polNTPKm (μM)kcat (s−1)kcat/Km (mM−1 s−1)Sugar selectivityb
pol μdATP15.6 ± 0.70.12 ± 0.017.9 ± 1.211
rATP114 ± 150.08 ± 0.020.70 ± 0.19
dCTP40.2 ± 9.20.12 ± 0.033.13 ± 0.491.4
rCTP78 ± 250.16 ± 0.022.17 ± 0.45
dGTP11.6 ± 1.50.54 ± 0.1246.5 ± 5.84.0
rGTP24.1 ± 6.40.28 ± 0.0511.7 ± 1.0
dTTP31.3 ± 5.70.21 ± 0.017.0 ± 1.97.2
rUTP189 ± 240.18 ± 0.020.97 ± 0.04
TdTdATP113 ± 170.74 ± 0.186.6 ± 1.68.9
rATP108 ± 100.08 ± 0.010.74 ± 0.15
dCTP254 ± 661.11 ± 0.054.51 ± 0.872.6
rCTP300 ± 8.40.53 ± 0.011.76 ± 0.05
pol βdATP2.40 ± 0.350.71 ± 0.13299 ± 846,000
rATP1,040 ± 4300.05 ± 0.030.05 ± 0.02
dCTP0.32 ± 0.010.81 ± 0.012,540 ± 952,000
rCTP62.1 ± 1.60.08 ± 0.021.25 ± 0.29
rGTP606 ± 2800.07 ± 0.030.11 ± 0.03NDc
rUTP1,320 ± 300.08 ± 0.010.06 ± 0.01ND
  • a Using standard 1-nt-gapped substrates (Fig. 1), with template base varied for each tested NTP as appropriate; detectable pol μ and pol β activity was template dependent, while TdT activity was independent of the template base, as expected. Data are means (± standard deviations) from replicate experiments.

  • b Sugar selectivity = [(kcat/Km)dNTP]/[(kcat/Km)rNTP].

  • c ND, not determined.