A single DNA binding site of DprA dimer is required to facilitate RecA filament nucleation

The DprA (a.k.a. Smf) protein has emerged as a RecA mediator during natural chromosomal transformation, but its ubiquity suggests a possible role in DNA metabolism beyond natural transformation. We show that Bacillus subtilis dprA increases the frequency of Escherichia coli Hfr conjugation. RecA{middle dot}ATP binds and cooperatively polymerises in a 50-nucleotide (nt) poly deoxy T (dT)50 ssDNA to form a dynamic filament with SSB competing for binding, but B. subtilis DprA (DprABsu) counters the inhibitory effects of SSB on RecA{middle dot}ATP filaments. RecA bound to (dT)21 is poorly active as dATPase, with DprABsu significantly improving RecA dATP hydrolysis. RecABsu{middle dot}dATP-(dT)20 complexes were readily formed, while DprABsu exerts an allosteric effect on RecABsu-(dT)15 complexes competent for dATP hydrolysis. Combining experimental data with a full-atomic model of the RecA-DprA-ssDNA complexs spatial structure, we proposed a molecular mechanism for DprA-mediated loading of RecA onto short ssDNA stretches. Our results suggest that steric constraints allow for the participation of only one DNA binding site of the DprA dimer in RecA-mediated dATP hydrolysis.