FIG 2.

DDDP, RDDP, and RNase H activities of HIV-1 RT mutants. The purified recombinant WT and mutant HIV-1 RT versions were tested. In the gels shown in panels A, B, and C, the order of all employed RT mutants is the same. (A) DDDP. (B) RDDP. Both assays were done by primer extensions on either DNA (DDDP) or RNA (RDDP) templates (see panel D). The mutant designations are indicated. Lanes C, control with no enzyme present. In panel A, “S” indicates the 21-nt substrate, and “P” indicates the 54-nt product of the primer extension. In panel B, “S” indicates the 20-nt substrate, and “P” indicates the 30-nt product of the primer extension. (C) The HIV-1 RT versions were assayed for their RNase H function, as described in Materials and Methods and in panel D. S, 33-nt labeled substrate. The reaction products are labeled as follows: C1, the 27-nt primary RNA product (that corresponds to the 17- to 18-nt cleavage according to the DNA 3′ end); C2, the secondary 18-nt product (corresponding to 8 to 9 nt from the DNA 3′ end). (D) Schematic descriptions of the performed assays. Upper panel, primer extensions (both RDDP and DDDP); lower panel, RNase H assay.