title: Chi hotspots trigger a conformational change in the helicase-like domain of AddAB to activate homologous recombination.
abstract: In bacteria, the repair of double-stranded DNA breaks is modulated by Chi sequences. These are recognised by helicase-nuclease complexes that process DNA ends for homologous recombination. Chi activates recombination by changing the biochemical properties of the helicase-nuclease, transforming it from a destructive exonuclease into a recombination-promoting repair enzyme. This transition is thought to be controlled by the Chi-dependent opening of a molecular latch, which enables part of the DNA substrate to evade degradation beyond Chi. Here, we show that disruption of the latch improves Chi recognition efficiency and stabilizes the interaction of AddAB with Chi, even in mutants that are impaired for Chi binding. Chi recognition elicits a structural change in AddAB that maps to a region of AddB which resembles a helicase domain, and which harbours both the Chi recognition locus and the latch. Mutation of the latch potentiates the change and moderately reduces the duration of a translocation pause at Chi. However, this mutant displays properties of Chi-modified AddAB even in the complete absence of bona fide hotspot sequences. The results are used to develop a model for AddAB regulation in which allosteric communication between Chi binding and latch opening ensures quality control during recombination hotspot recognition.
authors.affiliation: School of Biochemistry, University of Bristol, Bristol BS8 1TD, UK.
authors.forename: Neville S
authors.initials: NS
authors.lastname: Gilhooly
authors.affiliation: Department of Macromolecular Structures, Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Cientificas, 28049 Cantoblanco, Madrid, Spain.
authors.forename: Carolina
authors.initials: C
authors.lastname: Carrasco
authors.affiliation: Department of Macromolecular Structures, Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Cientificas, 28049 Cantoblanco, Madrid, Spain.
authors.forename: Benjamin
authors.identifier: http://orcid.org/0000-0001-6476-4614
authors.initials: B
authors.lastname: Gollnick
authors.affiliation: Institute of Cancer Research, Chester Beatty Laboratories, 237 Fulham Road, London SW3 6JB, UK Section of Structural Biology, Department of Medicine, Imperial College London, South Kensington Campus, London SW7 2AZ, UK.
authors.forename: Martin
authors.initials: M
authors.lastname: Wilkinson
authors.affiliation: Institute of Cancer Research, Chester Beatty Laboratories, 237 Fulham Road, London SW3 6JB, UK Section of Structural Biology, Department of Medicine, Imperial College London, South Kensington Campus, London SW7 2AZ, UK.
authors.forename: Dale B
authors.initials: DB
authors.lastname: Wigley
authors.affiliation: Department of Macromolecular Structures, Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Cientificas, 28049 Cantoblanco, Madrid, Spain.
authors.forename: Fernando
authors.initials: F
authors.lastname: Moreno-Herrero
authors.affiliation: School of Biochemistry, University of Bristol, Bristol BS8 1TD, UK mark.dillingham@bristol.ac.uk.
authors.forename: Mark S
authors.initials: MS
authors.lastname: Dillingham
mesh_terms: D001412:Bacillus subtilis / Q000201:enzymology* / Q000235:genetics; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D053903:DNA Breaks, Double-Stranded; D004265:DNA Helicases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004269:DNA, Bacterial / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D005090:Exodeoxyribonucleases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D008958:Models, Molecular; D009154:Mutation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D059767:Recombinational DNA Repair
chemical_list: D001426:Bacterial Proteins; D004269:DNA, Bacterial; D005090:Exodeoxyribonucleases; C407424:AddAB enzyme; D004265:DNA Helicases