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Structure paper

TitleThe structural mechanism of dimeric DONSON in replicative helicase activation.
Journal, issue, pagesMol Cell, Vol. 83, Issue 22, Page 4017-44031.e9, Year 2023
Publish dateNov 16, 2023
AuthorsMilos A Cvetkovic / Paolo Passaretti / Agata Butryn / Alicja Reynolds-Winczura / Georgia Kingsley / Aggeliki Skagia / Cyntia Fernandez-Cuesta / Divyasree Poovathumkadavil / Roger George / Anoop S Chauhan / Satpal S Jhujh / Grant S Stewart / Agnieszka Gambus / Alessandro Costa /
PubMed AbstractThe MCM motor of the replicative helicase is loaded onto origin DNA as an inactive double hexamer before replication initiation. Recruitment of activators GINS and Cdc45 upon S-phase transition ...The MCM motor of the replicative helicase is loaded onto origin DNA as an inactive double hexamer before replication initiation. Recruitment of activators GINS and Cdc45 upon S-phase transition promotes the assembly of two active CMG helicases. Although work with yeast established the mechanism for origin activation, how CMG is formed in higher eukaryotes is poorly understood. Metazoan Downstream neighbor of Son (DONSON) has recently been shown to deliver GINS to MCM during CMG assembly. What impact this has on the MCM double hexamer is unknown. Here, we used cryoelectron microscopy (cryo-EM) on proteins isolated from replicating Xenopus egg extracts to identify a double CMG complex bridged by a DONSON dimer. We find that tethering elements mediating complex formation are essential for replication. DONSON reconfigures the MCM motors in the double CMG, and primordial dwarfism patients' mutations disrupting DONSON dimerization affect GINS and MCM engagement in human cells and DNA synthesis in Xenopus egg extracts.
External linksMol Cell / PubMed:37820732
MethodsEM (single particle)
Resolution3.03 - 3.53 Å
Structure data

18191

8q6o

EMDB-18191, PDB-8q6o:
X. laevis CMG dimer bound to dimeric DONSON - without ATPase
Method: EM (single particle) / Resolution: 3.14 Å

18192

8q6p

EMDB-18192, PDB-8q6p:
X. laevis CMG dimer bound to dimeric DONSON - MCM ATPase
Method: EM (single particle) / Resolution: 3.53 Å

EMDB-18195: Single CMG purified from replicating Xenopus egg extracts
Method: EM (single particle) / Resolution: 3.03 Å

Chemicals

ChemComp-ZN:
Unknown entry

ChemComp-ATP:
ADENOSINE-5'-TRIPHOSPHATE / ATP, energy-carrying molecule*YM

Source
  • xenopus laevis (African clawed frog)
KeywordsREPLICATION / DNA replication initiation / Xenopus egg extract / primordial dwarfism / replicative helicase / genome stability

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