title: pCGR2 copy number depends on the par locus that forms a ParC-ParB-DNA partition complex in Corynebacterium glutamicum.
abstract: OBJECTIVE
  To characterize the par system of Corynebacterium glutamicum pCGR2 and to manipulate the par components to effectively manipulate plasmid copy number.
  
  
  RESULTS
  ParB binds sequence specifically to centromere-binding sites around the parAB operon and serves as an autorepressor. A small ORF (orf4, later named parC) downstream of parAB encodes a protein with 23.7% sequence identity with ParB. ParB is also implicated in the repression of parC transcription. Nonetheless, this ParC protein does not bind to centromere-binding sites and is not essential for plasmid stability. Introduction of a frameshift mutation within ParC implicated the protein in regulation of both parAB and parC. Electrophoretic Mobility Shift Assay confirmed a previously unreported ParC-ParB-parS partition complex. ParC also interacts directly with ParB without the mediation of the centromere sites. Deletion of the par components resulted in different plasmid copy numbers.
  
  
  CONCLUSIONS
  A previously unreported ParC-ParB-parS partition complex is formed in pCGR2, where interaction of ParC with ParB-parS may affect the level of repression by ParB. Modifying the par components and antisense RNA enables manipulation of plasmid copy number to varying degrees.
  
  
  CONCLUSIONS
  Genetically manipulating the par components, in combination with deactivation of antisense RNA, is a novel approach to artificially elevate plasmid copy number. This approach can be applied for development of new genetic engineering tools.
authors.affiliation: Research Institute of Innovative Technology for the Earth, Kizugawa, Kyoto, Japan.
authors.forename: Naoko
authors.initials: N
authors.lastname: Okibe
authors.forename: Nobuaki
authors.initials: N
authors.lastname: Suzuki
authors.forename: Masayuki
authors.initials: M
authors.lastname: Inui
authors.forename: Hideaki
authors.initials: H
authors.lastname: Yukawa
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D002503:Centromere / Q000378:metabolism; D048230:Corynebacterium glutamicum / Q000235:genetics* / Q000378:metabolism; D004269:DNA, Bacterial / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism; D008969:Molecular Sequence Data; D009876:Operon; D010957:Plasmids / Q000235:genetics*; D012097:Repressor Proteins / Q000737:chemistry / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D012097:Repressor Proteins
keywords: Corynebacterium glutamicum; antisense RNA; pCGR2; partition; plasmid copy number