title: DNA binding polarity, dimerization, and ATPase ring remodeling in the CMG helicase of the eukaryotic replisome.
abstract: The Cdc45/Mcm2-7/GINS (CMG) helicase separates DNA strands during replication in eukaryotes. How the CMG is assembled and engages DNA substrates remains unclear. Using electron microscopy, we have determined the structure of the CMG in the presence of ATPγS and a DNA duplex bearing a 3' single-stranded tail. The structure shows that the MCM subunits of the CMG bind preferentially to single-stranded DNA, establishes the polarity by which DNA enters into the Mcm2-7 pore, and explains how Cdc45 helps prevent DNA from dissociating from the helicase. The Mcm2-7 subcomplex forms a cracked-ring, right-handed spiral when DNA and nucleotide are bound, revealing unexpected congruencies between the CMG and both bacterial DnaB helicases and the AAA+ motor of the eukaryotic proteasome. The existence of a subpopulation of dimeric CMGs establishes the subunit register of Mcm2-7 double hexamers and together with the spiral form highlights how Mcm2-7 transitions through different conformational and assembly states as it matures into a functional helicase.
authors.affiliation: London Research Institute, Cancer Research UK, London, United Kingdom alessandro.costa@cancer.org.uk.
authors.forename: Alessandro
authors.initials: A
authors.lastname: Costa
authors.affiliation: London Research Institute, Cancer Research UK, London, United Kingdom Department of Imaging, National Institute for Biological Standards and Control, Potters Bar, United Kingdom.
authors.forename: Ludovic
authors.initials: L
authors.lastname: Renault
authors.affiliation: London Research Institute, Cancer Research UK, London, United Kingdom.
authors.forename: Paolo
authors.initials: P
authors.lastname: Swuec
authors.affiliation: Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States.
authors.forename: Tatjana
authors.initials: T
authors.lastname: Petojevic
authors.affiliation: Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States.
authors.forename: James J
authors.initials: JJ
authors.lastname: Pesavento
authors.affiliation: Institute of Technology, Faculty of Science and Technology, University of Tartu, Tartu, Estonia.
authors.forename: Ivar
authors.initials: I
authors.lastname: Ilves
authors.affiliation: Department of Imaging, National Institute for Biological Standards and Control, Potters Bar, United Kingdom.
authors.forename: Kirsty
authors.initials: K
authors.lastname: MacLellan-Gibson
authors.affiliation: Department of Imaging, National Institute for Biological Standards and Control, Potters Bar, United Kingdom.
authors.forename: Roland A
authors.initials: RA
authors.lastname: Fleck
authors.affiliation: Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States mbotchan@berkeley.edu.
authors.forename: Michael R
authors.initials: MR
authors.lastname: Botchan
authors.affiliation: Department of Biophysics and Biophysical Chemistry, Johns Hopkins University School of Medicine, Baltimore, United States jberge29@jhmi.edu.
authors.forename: James M
authors.initials: JM
authors.lastname: Berger
mesh_terms: D000251:Adenosine Triphosphatases / Q000737:chemistry / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism; D000818:Animals; D018797:Cell Cycle Proteins / Q000737:chemistry* / Q000378:metabolism; D002868:Chromosomal Proteins, Non-Histone / Q000737:chemistry* / Q000378:metabolism; D004247:DNA / Q000737:chemistry / Q000378:metabolism; D004261:DNA Replication; D004277:DNA, Single-Stranded / Q000737:chemistry / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism; D029721:Drosophila Proteins / Q000737:chemistry* / Q000378:metabolism; D004331:Drosophila melanogaster / Q000378:metabolism; D005057:Eukaryotic Cells / Q000378:metabolism; D008854:Microscopy, Electron; D064110:Minichromosome Maintenance Proteins / Q000737:chemistry* / Q000378:metabolism*; D008958:Models, Molecular; D046912:Multiprotein Complexes / Q000737:chemistry* / Q000378:metabolism / Q000648:ultrastructure; D011485:Protein Binding; D055503:Protein Multimerization; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism; D000072260:RNA Splicing Factors; D016601:RNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism; D012097:Repressor Proteins / Q000737:chemistry / Q000378:metabolism
chemical_list: C547399:Cdc45 protein, Drosophila; D018797:Cell Cycle Proteins; D002868:Chromosomal Proteins, Non-Histone; D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D029721:Drosophila Proteins; D046912:Multiprotein Complexes; D021122:Protein Subunits; D000072260:RNA Splicing Factors; D016601:RNA-Binding Proteins; C120397:RSF1 protein, Drosophila; D012097:Repressor Proteins; C022571:adenosine 5'-O-(3-thiotriphosphate); D000255:Adenosine Triphosphate; D004247:DNA; D000251:Adenosine Triphosphatases; D064110:Minichromosome Maintenance Proteins
keywords: AAA+ ATPase; DNA replication; Mcm2-7; helicase; motor proteins; replication fork