Figure 2.

Mfd WA550 exhibits elevated ATPase and DNA translocase activities. (A) Rates of ATP hydrolysis by Mfd derivatives. ATP hydrolysis rates were measured at 37°C using an ATP/NADH-coupled assay. Reactions were carried out in repair buffer and contained 50 nM protein and 1 mM ATP. 150 µM herring sperm DNA was added where indicated. Data are the average of three independent experiments and are shown with standard deviation. (B) Analysis of DNA translocation by Mfd derivatives. TFO displacement was analysed on 5 nM supercoiled pSRTB1 containing a triplex, which was incubated with 250 nM Mfd or its derivatives. Reactions were initiated with 2 mM ATP or ATPγS and samples were removed at time intervals, quenched and analysed by gel electrophoresis. The data shown are the percentage of TFO displaced normalized to the amount of free TFO at t = 0. The data are the average of three independent experiments shown with standard deviation. The TFO displacement data for WT Mfd and Mfd D7AAA have been published previously (5) and are shown for comparison.