THE RECG BRANCH MIGRATION PROTEIN ESCHERICHIA-COLI DISSOCIATES R-LOOPS

The RuvAB and RecG proteins of Escherichia coli promote branch migrati on of Holliday junction intermediates in genetic recombination. Both a re structure-specific helicases that unwind and rewind DNA at the junc tion point. The helicase activities of these proteins were investigate d using RNA:DNA hybrid molecules. RuvAB catalyses the unwinding of RNA :DNA partial duplexes of at least 218 bp in a reaction that requires b oth RuvA and RuvB, ATP and Mg2+. RecG failed to unwind these substrate s even when the duplex region was reduced to 35 bp. In contrast, RecG rapidly removes a 218 nt RNA from an R-loop substrate, whereas RuvAB d oes not. RecG's ability to dissociate R-loops is correlated with an ab ility to reduce the copy number of pUC plasmids and other constructs b ased on the ColE1 replicon. Copy number is reduced severely when the p lasmid carries recG(+). RecG is assumed to reduce copy number by inter fering with RNA II's ability to form an R-loop at the plasmid origin o f replication and prime DNA synthesis. The dissociation of R-loops by RecG is discussed in terms of the functions needed to promote recombin ation and to prime DNA replication at D-loops formed during the early stages of RecA-mediated recombination. (C) 1996 Academic Press Limited