5'-Single-stranded/duplex DNA junctions are loading sites for E. coli UvrD translocase

Escherichia coli UvrD is a 3'-5' superfamily 1A helicase/ translocase involved in a variety of DNA metabolic processes. UvrD can function either as a helicase or only as an singlestranded DNA (ssDNA) translocase. The switch between these activities is controlled in vitro by the UvrD oligomeric state; a monomer has ssDNA translocase activity, whereas at least a dimer is needed for helicase activity. Although a 30- ssDNA partial duplex provides a high-affinity site for a UvrD monomer, here we show that a monomer also binds with specificity to DNA junctions possessing a 50-ssDNA flanking region and can initiate translocation from this site. Thus, a 50-ss-duplex DNA junction can serve as a high-affinity loading site for the monomeric UvrD translocase, whereas a 30-ss-duplex DNA junction inhibits both translocase and helicase activity of the UvrD monomer. Furthermore, the 2B subdomain of UvrD is important for this junction speci- ficity. This highlights a separation of helicase and translocase function for UvrD and suggests that a monomeric UvrD translocase can be loaded at a 50-ssDNA junction when translocation activity alone is needed.