Stable synapsis of homologous DNA molecules mediated by the Escherichia coli RecA protein involves local exchange of DNA strands

Abstract

Escherichia coli RecA protein promotes stable synapsis between a single-stranded DNA and a homologous duplex DNA, resulting in the formation of a complex of RecA with three DNA strands. To gain insight into the molecular interactions responsible for DNA synapsis, the base-pairing status within the synaptic complex was analyzed by using dimethylsulfate and potassium permanganate as probes. The results indicate that the original base pairs in the parental duplex are disrupted; one strand is displaced and the other strand appears to be involved in Watson-Crick base-pairing with the incoming single-stranded DNA. The state of base-pairing thus resembles that of the end products of strand exchange and not a canonical DNA triple helix involving non-Watson-Crick base-pairing. The results also indicate that this local strand exchange can occur without homology at the ends of the DNA substrates (i.e., when axial rotation of the product heteroduplex with respect to the axis of the parental duplex is obstructed). Taken together, these results suggest that exchange of DNA strands mediated by RecA occur at or before the stage of stable DNA synapsis by a process that does not require DNA rotation.