Figure 3. Biochemical activities of WT and mutant G2L4 and GII RTs.

(A-D) Primer extension assays with 50-nt 3’-blocked DNA or RNA templates annealed to primers of various lengths. Reactions were initiated by adding 1 mM labeled dNTPs (1 mM each of dATP, dCTP, dGTP, and dTTP plus trace [α–³²P]-dTTP) and incubated for 240 min at 37°C.

(E and F) Terminal transferase assays with 5’-labeled 50-nt DNA or RNA templates (see above) without a 3’-blocking group, and 1 mM of the indicated dNTP incubated for 20 min at 37°C.

(G and H) Translesion DNA synthesis time courses with 50-nt 3’-blocked DNA templates containing an AP site or 8-oxoguanine 23 nt from the 3’ end. The tables show rate constants (k_obs) and amplitudes (Ampl.) for production of the labeled 50-nt DNA product obtained by fitting the data to a first-order rate equation.

The numbers to the left of the gels indicate the positions of size markers in a parallel lane.