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Structure paper

TitleStructure of DNA-CMG-Pol epsilon elucidates the roles of the non-catalytic polymerase modules in the eukaryotic replisome.
Journal, issue, pagesNat Commun, Vol. 9, Issue 1, Page 5061, Year 2018
Publish dateNov 29, 2018
AuthorsPanchali Goswami / Ferdos Abid Ali / Max E Douglas / Julia Locke / Andrew Purkiss / Agnieszka Janska / Patrik Eickhoff / Anne Early / Andrea Nans / Alan M C Cheung / John F X Diffley / Alessandro Costa /
PubMed AbstractEukaryotic origin firing depends on assembly of the Cdc45-MCM-GINS (CMG) helicase. A key step is the recruitment of GINS that requires the leading-strand polymerase Pol epsilon, composed of Pol2, ...Eukaryotic origin firing depends on assembly of the Cdc45-MCM-GINS (CMG) helicase. A key step is the recruitment of GINS that requires the leading-strand polymerase Pol epsilon, composed of Pol2, Dpb2, Dpb3, Dpb4. While a truncation of the catalytic N-terminal Pol2 supports cell division, Dpb2 and C-terminal Pol2 (C-Pol2) are essential for viability. Dpb2 and C-Pol2 are non-catalytic modules, shown or predicted to be related to an exonuclease and DNA polymerase, respectively. Here, we present the cryo-EM structure of the isolated C-Pol2/Dpb2 heterodimer, revealing that C-Pol2 contains a DNA polymerase fold. We also present the structure of CMG/C-Pol2/Dpb2 on a DNA fork, and find that polymerase binding changes both the helicase structure and fork-junction engagement. Inter-subunit contacts that keep the helicase-polymerase complex together explain several cellular phenotypes. At least some of these contacts are preserved during Pol epsilon-dependent CMG assembly on path to origin firing, as observed with DNA replication reconstituted in vitro.
External linksNat Commun / PubMed:30498216 / PubMed Central
MethodsEM (single particle)
Resolution4.4 - 4.98 Å
Structure data

0287

6hv8

EMDB-0287, PDB-6hv8:
Cryo-EM structure of S. cerevisiae Polymerase epsilon deltacat mutant
Method: EM (single particle) / Resolution: 4.4 Å

0288

6hv9

EMDB-0288, PDB-6hv9:
S. cerevisiae CMG-Pol epsilon-DNA
Method: EM (single particle) / Resolution: 4.98 Å

Chemicals

ChemComp-ZN:
Unknown entry

ChemComp-AGS:
PHOSPHOTHIOPHOSPHORIC ACID-ADENYLATE ESTER / ATP-gamma-S, energy-carrying molecule analogue*YM

Source
  • saccharomyces cerevisiae (brewer's yeast)
KeywordsDNA BINDING PROTEIN / Polymerase epsilon / DNA replication / enzyme / DNA polymerase / Helicase / Polymerase / AAA+ protein

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