Oligomeric initiator protein-mediated DNA looping negatively regulates plasmid replication in vitro by preventing origin melting.
Although DNA looping between the initiator binding sites (iterons) of the replication origin (ori) of a plasmid and the iterons located in a cis-acting control sequence called inc has been postulated to promote negative control of plasmid DNA replication, not only was definitive evidence for such looping lacking, but also the detailed molecular mechanism of this control had not been elucidated. Here, we present direct evidence showing that both the monomeric and the dimeric forms of the RepE initiator protein of F factor together promote pairing of incC-oriF sites by DNA looping. By using a reconstituted replication system consisting of 26 purified proteins, we show further that the DNA loop formation negatively regulates plasmid replication by inhibiting the formation of an open complex at the replication origin, thus elucidating a key step of replication control.
Zzaman
Shamsu
S
Department of Biochemistry and Molecular Biology, Medical University of South Carolina, Charleston, 29425, USA.
Bastia
Deepak
D
D004247:DNA / Q000737:chemistry / Q000378:metabolism; D004261:DNA Replication; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D019281:Dimerization; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism; D046911:Macromolecular Substances; D009690:Nucleic Acid Conformation; D009691:Nucleic Acid Denaturation; D010957:Plasmids / Q000235:genetics / Q000378:metabolism*; D011487:Protein Conformation; D018741:Replication Origin; D012097:Repressor Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D018840:HSP70 Heat-Shock Proteins; D046911:Macromolecular Substances; C074164:RepE protein, E coli; D012097:Repressor Proteins; D004247:DNA; C063842:dnaK protein, E coli