CryoEM structures of human CMG–ATPγS–DNA and CMG–AND-1 complexes
Open Access
- 26 May 2020
- journal article
- research article
- Published by Oxford University Press (OUP) in Nucleic Acids Research
- Vol. 48 (12), 6980-6995
- https://doi.org/10.1093/nar/gkaa429
Abstract
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.Keywords
Funding Information
- Wellcome Trust (104641/Z/14/Z, 202905/Z/16/Z, 206171/Z/17/Z)
- Engineering and Physical Sciences Research Council (EP/P020259/1)
- Science and Technology Facilities Council
- University of Cambridge OA funds
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