Roles of Bacillus subtilis DprA and SsbA in RecA-mediated Genetic Recombination

Background: ATP-bound RecA cannot catalyze DNA recombination without crucial accessory factors. Results: DprA facilitates RecA assembly onto SsbA- or SsbB-coated single-stranded DNA, but only DprA and SsbA enable RecAATP to mediate strand exchange. Conclusion: RecAATP transition to an active state requires DprA and SsbA. Significance: DprA and SsbA may serve as a two-component mediator during chromosomal transformation. Bacillus subtilis competence-induced RecA, SsbA, SsbB, and DprA are required to internalize and to recombine single-stranded (ss) DNA with homologous resident duplex. RecA, in the ATPMg(2+)-bound form (RecAATP), can nucleate and form filament onto ssDNA but is inactive to catalyze DNA recombination. We report that SsbA or SsbB bound to ssDNA blocks the RecA filament formation and fails to activate recombination. DprA facilitates RecA filamentation; however, the filaments cannot engage in DNA recombination. When ssDNA was preincubated with SsbA, but not SsbB, DprA was able to activate DNA strand exchange dependent on RecAATP. This work demonstrates that RecAATP, in concert with SsbA and DprA, catalyzes DNA strand exchange, and SsbB is an accessory factor in the reaction. In contrast, RecAdATP efficiently catalyzes strand exchange even in the absence of single-stranded binding proteins or DprA, and addition of the accessory factors marginally improved it. We proposed that the RecA-bound nucleotide (ATP and to a lesser extent dATP) might dictate the requirement for accessory factors.