title: RusA Holliday junction resolvase: DNA complex structure--insights into selectivity and specificity.
abstract: We have determined the structure of a catalytically inactive D70N variant of the Escherichia coli RusA resolvase bound to a duplex DNA substrate that reveals critical protein-DNA interactions and permits a much clearer understanding of the interaction of the enzyme with a Holliday junction (HJ). The RusA enzyme cleaves HJs, the fourway DNA branchpoints formed by homologous recombination, by introducing symmetrical cuts in the phosphodiester backbone in a Mg2+ dependent reaction. Although, RusA shows a high level of selectivity for DNA junctions, preferring to bind fourway junctions over other substrates in vitro, it has also been shown to have appreciable affinity for duplex DNA. However, RusA does not show DNA cleavage activity with duplex substrates. Our structure suggests the possible basis for structural selectivity as well as sources of the sequence specificity observed for DNA cleavage by RusA.
authors.affiliation: Department of Molecular Biology and Biotechnology, University of Sheffield, Firth Court, Western Bank, Sheffield S10 2TN, UK.
authors.forename: Rachel
authors.initials: R
authors.lastname: Macmaster
authors.forename: Svetlana
authors.initials: S
authors.lastname: Sedelnikova
authors.forename: Patrick J
authors.initials: PJ
authors.lastname: Baker
authors.forename: Edward L
authors.initials: EL
authors.lastname: Bolt
authors.forename: Robert G
authors.initials: RG
authors.lastname: Lloyd
authors.forename: John B
authors.initials: JB
authors.lastname: Rafferty
mesh_terms: D019943:Amino Acid Substitution; D001665:Binding Sites; D004247:DNA / Q000737:chemistry; D045566:DNA, Cruciform / Q000737:chemistry* / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D045565:Holliday Junction Resolvases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D013379:Substrate Specificity
chemical_list: D045566:DNA, Cruciform; D029968:Escherichia coli Proteins; D004247:DNA; D045565:Holliday Junction Resolvases; C112057:RusA protein, E coli