title: Frequent exchange of the DNA polymerase during bacterial chromosome replication.
abstract: The replisome is a multiprotein machine that carries out DNA replication. InEscherichia coli, a single pair of replisomes is responsible for duplicating the entire 4.6 Mbp circular chromosome. In vitro studies of reconstitutedE. colireplisomes have attributed this remarkable processivity to the high stability of the replisome once assembled on DNA. By examining replisomes in liveE. coliwith fluorescence microscopy, we found that the Pol III* subassembly frequently disengages from the replisome during DNA synthesis and exchanges with free copies from solution. In contrast, the DnaB helicase associates stably with the replication fork, providing the molecular basis for how theE. colireplisome can maintain high processivity and yet possess the flexibility to bypass obstructions in template DNA. Our data challenges the widely-accepted semi-discontinuous model of chromosomal replication, instead supporting a fully discontinuous mechanism in which synthesis of both leading and lagging strands is frequently interrupted.
authors.affiliation: Department of Biology, McGill University, Montreal, Canada.
authors.forename: Thomas R
authors.initials: TR
authors.lastname: Beattie
authors.affiliation: Department of Biology, McGill University, Montreal, Canada.
authors.forename: Nitin
authors.identifier: 0000-0003-4514-7631
authors.initials: N
authors.lastname: Kapadia
authors.affiliation: Department of Biochemistry, University of Oxford, Oxford, United Kingdom.
authors.forename: Emilien
authors.initials: E
authors.lastname: Nicolas
authors.affiliation: Department of Biochemistry, University of Oxford, Oxford, United Kingdom.
authors.forename: Stephan
authors.initials: S
authors.lastname: Uphoff
authors.affiliation: Biological Physical Sciences Institute, Departments of Physics and Biology, University of York, Heslington, United Kingdom.
authors.forename: Adam Jm
authors.initials: AJ
authors.lastname: Wollman
authors.affiliation: Biological Physical Sciences Institute, Departments of Physics and Biology, University of York, Heslington, United Kingdom.
authors.forename: Mark C
authors.initials: MC
authors.lastname: Leake
authors.affiliation: Department of Biology, McGill University, Montreal, Canada.
authors.forename: Rodrigo
authors.identifier: 0000-0002-5330-3481
authors.initials: R
authors.lastname: Reyes-Lamothe
mesh_terms: D002876:Chromosomes, Bacterial / Q000378:metabolism*; D004261:DNA Replication; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics*; D008856:Microscopy, Fluorescence
chemical_list: D004259:DNA-Directed DNA Polymerase
keywords: DNA replication; E. coli; bacteria; biochemistry; chromosomes; genes; genome integrity; replisome; single-molecule microscopy