THE FUNCTION OF THE SECONDARY DNA-BINDING SITE OF RECA PROTEIN DURINGDNA STRAND EXCHANGE

RecA protein features two distinct DNA-binding sites. During DNA stran d exchange, the primary site binds to single-stranded DNA (ssDNA), for ming the helical RecA nucleoprotein filament. The weaker secondary sit e binds double-stranded DNA (dsDNA) during the homolog search process. Here we demonstrate that this site has a second important function. I t binds the ssDNA strand that is displaced from homologous duplex DNA during DNA strand exchange, stabilizing the initial heteroduplex DNA p roduct. Although the high affinity of the secondary site for ssDNA is essential for DNA strand exchange, it renders DNA strand exchange sens itive to an excess of ssDNA which competes with dsDNA for binding. We further demonstrate that single-stranded DNA-binding protein can seque ster ssDNA, preventing its binding to the secondary site and thereby a ssisting at two levels: it averts the inhibition caused by an excess o f ssDNA and prevents the reversal of DNA strand exchange by removing t he displaced strand from the secondary site.