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Updated: Jun 4, 2025

Author Spotlight: Investigating the Motion Dynamics of the Eukaryotic Replisome Components at the Single-Molecule Level
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MCM2-7 ring closure involves the Mcm5 C-terminus and triggers Mcm4 ATP hydrolysis

Sarah V Faull1, Marta Barbon1,2, Audrey Mossler1

  • 1DNA Replication Group, Institute of Clinical Science, Imperial College London, London, UK.

Nature Communications
|January 2, 2025
PubMed

Insights

The MCM2-7 helicase loading requires ring closure and ATP hydrolysis. Mcm4 ATPase activity is key for regulating pre-replicative complex assembly, with a stable Mcm2/Mcm5 interface being essential.

Area of Science:

  • Molecular Biology
  • Cellular Biology
  • Biochemistry

Background:

  • The eukaryotic MCM2-7 helicase is loaded onto replication origins by ORC, Cdc6, and Cdt1, forming a double-hexamer.
  • DNA insertion into MCM2-7 causes partial ring closure, and ATP hydrolysis is critical for pre-replicative complex (pre-RC) assembly.
  • Mechanisms controlling MCM2-7 ring closure and ATP hydrolysis remain unclear.

Purpose of the Study:

  • To elucidate the control mechanisms of MCM2-7 ring closure and ATP hydrolysis during pre-RC assembly.
  • To identify the key ATPase regulating helicase loading and complex stability.

Main Methods:

  • Cryo-electron microscopy (cryo-EM) to determine the structure of an ORC-Cdc6-Cdt1-MCM2-7 intermediate.
  • Analysis of mutations at the Mcm2/Mcm5 interface affecting helicase loading and stability.
  • Investigation of Mcm4 ATP hydrolysis during helicase loading.

Main Results:

  • A cryo-EM structure revealed a remodelled, fully-closed Mcm2/Mcm5 interface, with the Mcm5 C-terminus contacting Orc3.
  • Normal helicase loading triggers Mcm4 ATP hydrolysis, leading to MCM2-7 complex reorganization and Cdt1 release.
  • Defective Mcm2/Mcm5 interface closure causes MCM2-7 ring splitting and complex disassembly.

Conclusions:

  • Mcm4 is identified as the key ATPase regulating pre-RC formation.
  • A stable Mcm2/Mcm5 interface is essential for ATP hydrolysis-dependent MCM2-7 remodelling and productive helicase loading.
  • Understanding these mechanisms provides insights into DNA replication initiation.

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