title: Resolving Holliday junctions with Escherichia coli UvrD helicase.
abstract: The Escherichia coli UvrD helicase is known to function in the mismatch repair and nucleotide excision repair pathways and has also been suggested to have roles in recombination and replication restart. The primary intermediate DNA structure in these two processes is the Holliday junction. UvrD has been shown to unwind a variety of substrates including partial duplex DNA, nicked DNA, forked DNA structures, blunt duplex DNA and RNA-DNA hybrids. Here, we demonstrate that UvrD also catalyzes the robust unwinding of Holliday junction substrates. To characterize this unwinding reaction we have employed steady-state helicase assays, pre-steady-state rapid quench helicase assays, DNaseI footprinting, and electron microscopy. We conclude that UvrD binds initially to the junction compared with binding one of the blunt ends of the four-way junction to initiate unwinding and resolves the synthetic substrate into two double-stranded fork structures. We suggest that UvrD, along with its mismatch repair partners, MutS and MutL, may utilize its ability to unwind Holliday junctions directly in the prevention of homeologous recombination. UvrD may also be involved in the resolution of stalled replication forks by unwinding the Holliday junction intermediate to allow bypass of the blockage.
authors.affiliation: Department of Biology, University of North Carolina, Chapel Hill, North Carolina 27599, USA.
authors.forename: Annamarie S
authors.initials: AS
authors.lastname: Carter
authors.forename: Kambiz
authors.initials: K
authors.lastname: Tahmaseb
authors.forename: Sarah A
authors.initials: SA
authors.lastname: Compton
authors.forename: Steven W
authors.initials: SW
authors.lastname: Matson
mesh_terms: D000251:Adenosine Triphosphatases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D018983:DNA Footprinting / Q000379:methods; D004265:DNA Helicases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D045566:DNA, Cruciform / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D003850:Deoxyribonuclease I / Q000737:chemistry; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000070956:MutL Proteins; D051719:MutS DNA Mismatch-Binding Protein / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D009692:Nucleic Acid Heteroduplexes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D004269:DNA, Bacterial; D045566:DNA, Cruciform; D029968:Escherichia coli Proteins; C074770:MutL protein, E coli; D009692:Nucleic Acid Heteroduplexes; D003850:Deoxyribonuclease I; D000251:Adenosine Triphosphatases; C506746:UvrD protein, E coli; D000070956:MutL Proteins; D051719:MutS DNA Mismatch-Binding Protein; C497175:MutS protein, E coli; D004265:DNA Helicases