S-DNA and RecA/RAD51-Mediated Strand Exchange in Vitro

S-DNA (stretched DNA) is an elongated base-paired DNA conformation under high tension. Because the RecA/Rad51 family DNA recombinases form helical filaments on DNA and mediate the formation of DNA triplex (D-loop) in which the DNA is stretched, and because the extension of these nucleoprotein filaments are similar to the extension of S-DNA, S-DNA has long been hypothesized as a possible intrinsic state of DNA that participant in RecA/Rad51-mediated DNA strand-exchange in homologous recombination. Such a hypothesis, however, is still lacking directly experimental studies. In this work, we have studied the polymerization of and strand-exchange on S-DNA mediated by Escherichia coli RecA, human Rad51 and Saccharomyces cerevisiae Rad51 by single-molecule magnetic-tweezers. We report that RecA/Rad51 polymerizes faster on S-DNA than on B-DNA at the same buffer conditions. Furthermore, RecA/Rad51 mediated DNA triplexes forms faster from S-DNA than from B-DNA together with the homologous single-stranded DNA. These...