title: Structural basis for the inhibition of RecBCD by Gam and its synergistic antibacterial effect with quinolones.
abstract: Our previous paper (Wilkinsonet al, 2016) used high-resolution cryo-electron microscopy to solve the structure of theEscherichia coliRecBCD complex, which acts in both the repair of double-stranded DNA breaks and the degradation of bacteriophage DNA. To counteract the latter activity, bacteriophage λ encodes a small protein inhibitor called Gam that binds to RecBCD and inactivates the complex. Here, we show that Gam inhibits RecBCD by competing at the DNA-binding site. The interaction surface is extensive and involves molecular mimicry of the DNA substrate. We also show that expression of Gam inE. coliorKlebsiella pneumoniaeincreases sensitivity to fluoroquinolones; antibacterials that kill cells by inhibiting topoisomerases and inducing double-stranded DNA breaks. Furthermore, fluoroquinolone-resistance inK. pneumoniaeclinical isolates is reversed by expression of Gam. Together, our data explain the synthetic lethality observed between topoisomerase-induced DNA breaks and the RecBCD gene products, suggesting a new co-antibacterial strategy.
authors.affiliation: Department of Medicine, Section of Structural Biology, Imperial College London, London, United Kingdom.
authors.forename: Martin
authors.initials: M
authors.lastname: Wilkinson
authors.affiliation: School of Biochemistry, University of Bristol, Bristol, United Kingdom.
authors.forename: Luca
authors.initials: L
authors.lastname: Troman
authors.affiliation: School of Cellular and Molecular Medicine, University of Bristol, Bristol, United Kingdom.
authors.forename: Wan Ak
authors.initials: WA
authors.lastname: Wan Nur Ismah
authors.affiliation: Department of Medicine, Section of Structural Biology, Imperial College London, London, United Kingdom.
authors.forename: Yuriy
authors.initials: Y
authors.lastname: Chaban
authors.affiliation: School of Cellular and Molecular Medicine, University of Bristol, Bristol, United Kingdom.
authors.forename: Matthew B
authors.initials: MB
authors.lastname: Avison
authors.affiliation: School of Biochemistry, University of Bristol, Bristol, United Kingdom.
authors.forename: Mark S
authors.identifier: 0000-0002-4612-7141
authors.initials: MS
authors.lastname: Dillingham
authors.affiliation: Department of Medicine, Section of Structural Biology, Imperial College London, London, United Kingdom.
authors.forename: Dale B
authors.identifier: 0000-0002-0786-6726
authors.initials: DB
authors.lastname: Wigley
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D010582:Bacteriophage lambda / Q000201:enzymology; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004357:Drug Synergism; D004926:Escherichia coli / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics; D043211:Exodeoxyribonuclease V / Q000037:antagonists & inhibitors*; D007711:Klebsiella pneumoniae / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics; D015363:Quinolones / Q000494:pharmacology*; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D000900:Anti-Bacterial Agents; D004268:DNA-Binding Proteins; D015363:Quinolones; D014764:Viral Proteins; C065410:gam protein, Coliphage; D043211:Exodeoxyribonuclease V
keywords: DNA repair; E. coli; RecBCD; antibacterials; biophysics; infectious disease; microbiology; structural biology