title: RecG and UvsW catalyse robust DNA rewinding critical for stalled DNA replication fork rescue.
abstract: Helicases that both unwind and rewind DNA have central roles in DNA repair and genetic recombination. In contrast to unwinding, DNA rewinding by helicases has proved difficult to characterize biochemically because of its thermodynamically downhill nature. Here we use single-molecule assays to mechanically destabilize a DNA molecule and follow, in real time, unwinding and rewinding by two DNA repair helicases, bacteriophage T4 UvsW and Escherichia coli RecG. We find that both enzymes are robust rewinding enzymes, which can work against opposing forces as large as 35 pN, revealing their active character. The generation of work during the rewinding reaction allows them to couple rewinding to DNA unwinding and/or protein displacement reactions central to the rescue of stalled DNA replication forks. The overall results support a general mechanism for monomeric rewinding enzymes.
authors.affiliation: 1] Departament de Física Fonamental, Facultat de Física, Universitat de Barcelona, Diagonal 647, 08028 Barcelona, Spain [2] CIBER-BBN de Bioingenieria, Biomateriales y Nanomedicina, Instituto de Sanidad Carlos III, Madrid, Spain.
authors.forename: Maria
authors.initials: M
authors.lastname: Manosas
authors.forename: Senthil K
authors.initials: SK
authors.lastname: Perumal
authors.forename: Piero R
authors.initials: PR
authors.lastname: Bianco
authors.forename: Piero
authors.initials: P
authors.lastname: Bianco
authors.forename: Felix
authors.initials: F
authors.lastname: Ritort
authors.forename: Stephen J
authors.initials: SJ
authors.lastname: Benkovic
authors.forename: Vincent
authors.initials: V
authors.lastname: Croquette
mesh_terms: D017122:Bacteriophage T4 / Q000201:enzymology; D055162:Biocatalysis / Q000187:drug effects; D001696:Biomechanical Phenomena / Q000187:drug effects; D004247:DNA / Q000737:chemistry* / Q000378:metabolism*; D004265:DNA Helicases / Q000378:metabolism*; D004261:DNA Replication / Q000187:drug effects; D045566:DNA, Cruciform / Q000187:drug effects / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology; D029968:Escherichia coli Proteins / Q000378:metabolism*; D008274:Magnesium / Q000494:pharmacology; D060328:Magnetic Phenomena; D052898:Optical Tweezers; D009994:Osmolar Concentration; D013379:Substrate Specificity / Q000187:drug effects; D014764:Viral Proteins / Q000378:metabolism*
chemical_list: D045566:DNA, Cruciform; D029968:Escherichia coli Proteins; D014764:Viral Proteins; C071446:RecG protein, E coli; D004247:DNA; D004265:DNA Helicases; C107414:UvsW protein, Bacteriophage T4; D008274:Magnesium