FIGURE 8.

Similarly to the SSB proteins, DdrB enhances EcRecA and inhibits DrRecA ssDNA binding activities. RecA ssDNA-dependent ATPase activity, proportional to the amount of RecA bound to DNA, was monitored at 380_nm to follow the loss of NADH used in the coupled ATP regeneration system. The reactions were carried out at 37 °C. 3 μm RecA protein was added first to M13mp18 circular ssDNA (5 μm nucleotides), and the reactions were initiated by the addition of 3 mm ATP. The SSB or DdrB proteins (0.5 μm each) were added 10 min after (arrow). Curves from typical experiments are represented. A, effects on E. coli RecA. The k_cat of EcRecA on ssDNA alone is 11.42 ± 0.98 min⁻¹. Addition of EcSSB and DrSSB enhanced EcRecA ATPase activity to k_cat values of 30.76 ± 0.48 min⁻¹ and 30.26 ± 1.10 min⁻¹, respectively. Addition of DdrB had a similar but lesser effect on EcRecA yielding a k_cat of 24.80 ± 0.47 min⁻¹ (thick line). B, effects on D. radiodurans RecA. The k_cat value for DrRecA alone is 6.89 ± 0.15 min⁻¹. Addition of EcSSB or DrSSB produced an initial enhancement to k_cat values of, respectively, 10.72 ± 0.36 min⁻¹ and 9.45 ± 0.59 min⁻¹ followed by a slow inhibition to k_cat values of 0.77 ± 0.52 min⁻¹ and 1.13 ± 0.84 min⁻¹ with EcSSB and DrSSB, respectively. Addition of DdrB immediately decreased DrRecA ATPase activity to a k_cat value of 0.59 ± 0.09 min⁻¹ (thick line).