Mechanisms of RecA filament nucleation on ssDNA by the DprA protein

The DprA protein has been implicated in the protecton of incoming DNA. However, the presence of dprA gene family members, also known as smf, across diverse bacterial species suggests a broader functonality for their gene products. We eaamined the role of the Escherichia coli DprA/Smf homologue in conjugaton. Remarkably, Bacillus subtilis dprA/smf can complement an E. coli dprA mutant, indicatng interchangeability of dprA/smf genes between competent and non-competent species in conjugatonal processes. The DprA protein forms a complex with DNA, facilitatng the nucleaton of RecA protein flaments onto circular single-stranded DNA coated with SSB protein. To focus on RecA nucleaton, we employed short DNA oligonucleotdes that restrict RecA-DNA binding but allow for DprA-RecA-DNA binding. Analysis of dATPase actvity revealed that RecA-DNA complexes were readily formed only with olig50, while DprA-RecA-DNA complexes were also feasible with olig21. Combining eaperimental data with a full-atomic model of the RecA-DprA-ssDNA complex’s spatal structure, we proposed a molecular mechanism for DprA-mediated loading of RecA proteins onto ssDNA. Our fndings suggest that only one DprA-ssDNA interacton can occur sterically, occupying one strand of ssDNA in the complex.