title: Molecular basis of the supercoil deficit induced by the mini-F plasmid partition complex.
abstract: Formation of a partition complex on plasmid F by binding of SopB protein to the sopC centromere is the first step in the partition process that ensures stability of F in dividing cells. Establishment of the complex enables nonspecific binding of SopB to neighboring DNA, which extends the partition complex and provokes reduction of negative supercoiling of the plasmid. This reduction is believed to reflect winding of DNA into positive supercoils about SopB to create a nucleoprotein structure of probable importance to partition. We have searched for evidence that SopB alters plasmid topology. Permutation analysis indicated only modest bending of linear DNA fragments, and in vivo DNase I footprinting revealed no enhanced cleavages indicating curvature. In vitro, SopB binding left no topological trace in relaxed-circular DNA treated with topoisomerase I or in nicked circles closed by ligase. In vivo, novobiocin-mediated inhibition of DNA gyrase relaxed a plasmid carrying the partition complex but left no residue of positive supercoils. Hence, SopB does not reduce plasmid supercoiling directly. We did observe that SopB partly prevented removal of negative supercoils from plasmid DNA by topoisomerase I and partly prevented ligation of nicked circles, indicating that it acts as a physical obstacle. The supercoil deficit is thus better explained as SopB recoating of just-replicated DNA, which shelters it from gyrase and from topological changes in SopB-free DNA. This topological simplicity distinguishes the Sop partition complex from other complexes described.
authors.affiliation: Laboratoire de Microbiologie et Gánátique Moláculaires, CNRS, F-31000 Toulouse, France and LMGM, Universitáde Toulouse, UPS, F-31000 Toulouse, France; Laboratoire de Microbiologie et Gánátique Moláculaires, CNRS, F-31000 Toulouse, France and LMGM, Universitáde Toulouse, UPS, F-31000 Toulouse, France. Electronic address: jean-yves.bouet@ibcg.biotoul.fr.
authors.forename: Jean-Yves
authors.initials: JY
authors.lastname: Bouet
authors.affiliation: Laboratoire de Microbiologie et Gánátique Moláculaires, CNRS, F-31000 Toulouse, France and LMGM, Universitáde Toulouse, UPS, F-31000 Toulouse, France.
authors.forename: David
authors.initials: D
authors.lastname: Lane
mesh_terms: D027081:DNA Gyrase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004264:DNA Topoisomerases, Type I / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D004278:DNA, Superhelical / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000378:metabolism; D010957:Plasmids / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding / Q000502:physiology
chemical_list: D004278:DNA, Superhelical; D029968:Escherichia coli Proteins; C489223:SopB protein, E coli; D004264:DNA Topoisomerases, Type I; D027081:DNA Gyrase