title: A mutation in helicase motif III of E. coli RecG protein abolishes branch migration of Holliday junctions.
abstract: The RecG protein of Escherichia coli catalyses branch migration of Holliday junctions made by RecA and dissociates synthetic X junctions into duplex products in reactions that require hydrolysis of ATP. To investigate the mode of action of this enzyme a chromosomal mutation that inactivates recG (recG162) was cloned and sequenced. The recG162 mutation is a G:C to A:T transition, which produces an Ala428 to Val substitution in the protein. This change affects a motif (motif III) in the protein that is highly conserved in DNA and RNA helicases. RecG162 protein was purified and shown to retain the ability to bind synthetic X and Y junctions. However, it does not dissociate these junctions and fails to catalyse branch migration of Holliday junction intermediates purified from a RecA strand exchange reaction. RecG162 retains a DNA-dependent ATPase activity, but this is much reduced relative to the wild-type protein, especially with single-stranded DNA as a co-factor. These results suggest that branch migration by RecG is related to a junction-targeted DNA helicase activity.
authors.affiliation: Department of Genetics, University of Nottingham, Queens Medical Centre, UK.
authors.forename: G J
authors.initials: GJ
authors.lastname: Sharples
authors.forename: M C
authors.initials: MC
authors.lastname: Whitby
authors.forename: L
authors.initials: L
authors.lastname: Ryder
authors.forename: R G
authors.initials: RG
authors.lastname: Lloyd
mesh_terms: D000251:Adenosine Triphosphatases / Q000235:genetics / Q000378:metabolism; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D004265:DNA Helicases / Q000235:genetics / Q000378:metabolism*; D017931:DNA Primers / Q000737:chemistry; D004260:DNA Repair; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology* / Q000528:radiation effects; D029968:Escherichia coli Proteins; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D011995:Recombination, Genetic; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D014466:Ultraviolet Rays
chemical_list: D001426:Bacterial Proteins; D017931:DNA Primers; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C071446:RecG protein, E coli; D000251:Adenosine Triphosphatases; D004265:DNA Helicases