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A conserved MCM single-stranded DNA binding element is essential for replication initiation.

Clifford A Froelich1, Sukhyun Kang, Leslie B Epling

  • 1Department of Structural Biology, St Jude Children's Research Hospital, Memphis, United States.

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|April 3, 2014
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Summary
This summary is machine-generated.

The MCM helicase

Keywords:
DNA replicationarchaeacrystallographygeneticshelicase

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Area of Science:

  • Molecular biology
  • Biochemistry
  • Structural biology

Background:

  • The MCM helicase complex is crucial for DNA replication, loading as an inactive double-hexamer onto duplex DNA at origins.
  • Helicase activation transforms the double-hexamer into two active single hexamers that encircle single-stranded DNA (ssDNA).
  • The precise molecular interactions between MCM and DNA during these critical events remain largely uncharacterized.

Purpose of the Study:

  • To elucidate the molecular mechanisms of MCM-DNA interactions during helicase loading and activation.
  • To define the structural basis of MCM's interaction with single-stranded DNA (ssDNA).
  • To investigate the functional significance of identified MCM-ssDNA interactions in eukaryotic DNA replication.

Main Methods:

  • Determined the crystal structure of the Pyrococcus furiosus MCM N-terminal domain hexamer bound to ssDNA.
  • Defined a conserved MCM-ssDNA binding motif (MSSB) through structural analysis.
  • Investigated the function of the MSSB in eukaryotic MCM complexes using mutant analysis in S. cerevisiae.

Main Results:

  • ssDNA binds to the interior of the MCM ring, perpendicular to the central channel, with a defined polarity.
  • A conserved MCM-ssDNA binding motif (MSSB) was identified and structurally characterized.
  • Mutations in the MSSB in S. cerevisiae Mcm2-7 complexes result in non-viable cells, with defects in helicase loading and activation at replication origins.

Conclusions:

  • The study identifies a critical interaction between MCM and ssDNA mediated by the MSSB.
  • This interaction is essential for MCM helicase activation and likely plays a role in selecting the DNA strand for translocation.
  • The findings provide fundamental insights into the mechanism of DNA replication initiation and regulation.