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·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 (DprA Bsu ) counters the inhibitory effects of SSB on RecA·ATP filaments. RecA bound to (dT) 21 is poorly active as dATPase, with DprA Bsu significantly improving RecA dATP hydrolysis. RecA Bsu ·dATP-(dT) 20 complexes were readily formed, while DprA Bsu exerts an allosteric effect on RecA Bsu -(dT) 15 complexes competent for dATP hydrolysis. Combining experimental data with a full-atomic model of the RecA-DprA-ssDNA complex’s 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.