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TitleCryoEM structures of human CMG-ATPγS-DNA and CMG-AND-1 complexes.
Journal, issue, pagesNucleic Acids Res, Vol. 48, Issue 12, Page 6980-6995, Year 2020
Publish dateJul 9, 2020
AuthorsNeil J Rzechorzek / Steven W Hardwick / Vincentius A Jatikusumo / Dimitri Y Chirgadze / Luca Pellegrini /
PubMed AbstractDNA unwinding in eukaryotic replication is performed by the Cdc45-MCM-GINS (CMG) helicase. Although the CMG architecture has been elucidated, its mechanism of DNA unwinding and replisome interactions ...DNA unwinding in eukaryotic replication is performed by the Cdc45-MCM-GINS (CMG) helicase. Although the CMG architecture has been elucidated, its mechanism of DNA unwinding and replisome interactions remain poorly understood. Here we report the cryoEM structure at 3.3 Å of human CMG bound to fork DNA and the ATP-analogue ATPγS. Eleven nucleotides of single-stranded (ss) DNA are bound within the C-tier of MCM2-7 AAA+ ATPase domains. All MCM subunits contact DNA, from MCM2 at the 5'-end to MCM5 at the 3'-end of the DNA spiral, but only MCM6, 4, 7 and 3 make a full set of interactions. DNA binding correlates with nucleotide occupancy: five MCM subunits are bound to either ATPγS or ADP, whereas the apo MCM2-5 interface remains open. We further report the cryoEM structure of human CMG bound to the replisome hub AND-1 (CMGA). The AND-1 trimer uses one β-propeller domain of its trimerisation region to dock onto the side of the helicase assembly formed by Cdc45 and GINS. In the resulting CMGA architecture, the AND-1 trimer is closely positioned to the fork DNA while its CIP (Ctf4-interacting peptide)-binding helical domains remain available to recruit partner proteins.
External linksNucleic Acids Res / PubMed:32453425 / PubMed Central
MethodsEM (single particle)
Resolution3.29 - 6.77 Å
Structure data

EMDB-10619, PDB-6xtx:
CryoEM structure of human CMG bound to ATPgammaS and DNA
Method: EM (single particle) / Resolution: 3.29 Å

EMDB-10620:
CryoEM structure of human CMG bound to ATPgammaS and DNA - MCM2-7 C-tier
Method: EM (single particle) / Resolution: 3.41 Å

EMDB-10621, PDB-6xty:
CryoEM structure of human CMG bound to AND-1 (CMGA)
Method: EM (single particle) / Resolution: 6.77 Å

Chemicals

ChemComp-ZN:
Unknown entry

ChemComp-ADP:
ADENOSINE-5'-DIPHOSPHATE / ADP, energy-carrying molecule*YM / Adenosine diphosphate

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

ChemComp-MG:
Unknown entry

Source
  • homo sapiens (human)
  • synthetic construct (others)
KeywordsREPLICATION / CMG / Helicase / ATPase / Replisome

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