Figure 5. RuvA_Mge inhibits HJ resolution by RecU_Mge.

(A, B) HJ resolution assays [19] were performed in volumes of 10 µl and contained 12.3 nM HJ substrate HJ 1.1 (6-FAM-labeled), RecU_Mge (0.2 µM) and various concentrations of RuvA_Mge, as indicated above the lanes. Reactions were preincubated for 2 min with either RecU_Mge (A) or RuvA_Mge (B), followed by addition of the other protein. After incubation for 30 min at 37°C, the reaction products were separated on 12% polyacrylamide gels, and analyzed by fluorometry. The locations of the HJ substrate and resolution products are indicated schematically at the right-hand side of the gels. (C) Quantification of the influence of RuvA_Mge on RecU_Mge activity. The relative RecU_Mge (resolution) activity was measured from the gels shown in (A) and (B) and expressed as percentage of the protein's activity in the absence of RuvA_Mge. The data from (A) and (B) are represented by the closed squares (▪) and the open squares (□), respectively.