title: Analysis of the pSK1 replicon, a prototype from the staphylococcal multiresistance plasmid family.
abstract: Multidrug-resistant staphylococci often harbour plasmids that carry genes conferring resistance to several antimicrobial compounds. Many of these multiresistance plasmids appear to utilize a related theta-type replication system for which multiresistance plasmid pSK1 serves as a prototype. Essential pSK1 replication elements were identified by cloning segments of the replication region and testing the resulting plasmids for replication proficiency. An iterated region within rep and a DNA segment of up to 68 bp upstream of the rep promoter were both found to be essential for origin activity. The Rep protein was overexpressed as a 6xHis-tagged C-terminal fusion protein and was shown to bind in vitro to four Rep boxes located within the rep coding region. Inactivation of a divergently oriented promoter upstream of rep, designated P(rnaI), resulted in an elevated plasmid copy number. Comparative analyses suggest that the replication systems of many staphylococcal multiresistance plasmids share a similar genetic organization and utilize an antisense-RNA-mediated regulatory mechanism for copy number control.
authors.affiliation: School of Biological Sciences, University of Sydney, New South Wales 2006, Australia.
authors.forename: Stephen M
authors.initials: SM
authors.lastname: Kwong
authors.affiliation: School of Biological Sciences, University of Sydney, New South Wales 2006, Australia.
authors.forename: Ricky
authors.initials: R
authors.lastname: Lim
authors.affiliation: School of Biological Sciences, University of Sydney, New South Wales 2006, Australia.
authors.forename: Rebecca J
authors.initials: RJ
authors.lastname: LeBard
authors.affiliation: School of Biological Sciences, University of Sydney, New South Wales 2006, Australia.
authors.forename: Ronald A
authors.initials: RA
authors.lastname: Skurray
authors.affiliation: School of Biological Sciences, University of Sydney, New South Wales 2006, Australia.
authors.forename: Neville
authors.initials: N
authors.lastname: Firth
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics; D001483:Base Sequence; D004261:DNA Replication; D004269:DNA, Bacterial / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics; D024901:Drug Resistance, Multiple, Bacterial / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics; D018628:Gene Dosage; D015964:Gene Expression Regulation, Bacterial; D008969:Molecular Sequence Data; D009154:Mutation; D010957:Plasmids / Q000235:genetics*; D011401:Promoter Regions, Genetic; D016372:RNA, Antisense / Q000235:genetics; D012329:RNA, Bacterial / Q000235:genetics; D012093:Replicon; D013211:Staphylococcus aureus / Q000235:genetics*; D024363:Transcription Initiation Site
chemical_list: D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D016372:RNA, Antisense; D012329:RNA, Bacterial