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Open-ringed structure of the Cdt1–Mcm2–7 complex as a precursor of the MCM double hexamer

Nature Structural & Molecular Biology volume 24, pages 300308 (2017) | Download Citation

Abstract

The minichromosome maintenance complex (MCM) hexameric complex (Mcm2–7) forms the core of the eukaryotic replicative helicase. During G1 phase, two Cdt1–Mcm2–7 heptamers are loaded onto each replication origin by the origin-recognition complex (ORC) and Cdc6 to form an inactive MCM double hexamer (DH), but the detailed loading mechanism remains unclear. Here we examine the structures of the yeast MCM hexamer and Cdt1–MCM heptamer from Saccharomyces cerevisiae. Both complexes form left-handed coil structures with a 10–15-Å gap between Mcm5 and Mcm2, and a central channel that is occluded by the C-terminal domain winged-helix motif of Mcm5. Cdt1 wraps around the N-terminal regions of Mcm2, Mcm6 and Mcm4 to stabilize the whole complex. The intrinsic coiled structures of the precursors provide insights into the DH formation, and suggest a spring-action model for the MCM during the initial origin melting and the subsequent DNA unwinding.

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Acknowledgements

We thank the Tsinghua University Branch of the China National Center for Protein Sciences (Beijing, China) for technical support with cryo-EM data collection and for computation resources. Part of the computation was done on the Computing Platform of the Center for Life Science, Peking University. We also thank J. Diffley and S. Gasser (Francis Crick Institute, London, UK) for yeast and E. coli strains. This work was supported by the Ministry of Science and Technology of China (grants 2013CB910404 and 2016YFA0500700 to N.G.), the National Natural Science Foundation of China (grants 31422016, 31470722 and 31630087 to N.G.), the Research Grants Council of Hong Kong (grants GRF16138716 to B.-K.T.; GRF664013, HKUST12/CRF/13G, GRF16104115 and GRF16143016 to Y.Z. and B.-K.T.; and IGN15SC02 to Y.Z.). N.L. is supported by a postdoctoral fellowship from the Peking-Tsinghua Center for Life Sciences.

Author information

Author notes

    • Yuanliang Zhai
    •  & Erchao Cheng

    These authors contributed equally to this work.

Affiliations

  1. Division of Life Science, Hong Kong University of Science and Technology, Clear Water Bay, Hong Kong, China.

    • Yuanliang Zhai
    • , Philip Yuk Kwong Yung
    •  & Bik-Kwoon Tye
  2. Institute for Advanced Study, Hong Kong University of Science and Technology, Clear Water Bay, Hong Kong, China.

    • Yuanliang Zhai
  3. Ministry of Education Key Laboratory of Protein Sciences, Beijing Advanced Innovation Center for Structural Biology, School of Life Sciences, Tsinghua University, Beijing, China.

    • Erchao Cheng
    • , Hao Wu
    •  & Ning Gao
  4. Peking-Tsinghua Center for Life Sciences, School of Life Sciences, Peking University, Beijing, China.

    • Ningning Li
  5. Department of Molecular Biology and Genetics, College of Agriculture and Life Sciences, Cornell University, Ithaca, New York, USA.

    • Bik-Kwoon Tye

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Contributions

Y.Z. and P.Y.K.Y. purified proteins; E.C., H.W. and N.L. collected and processed data; P.Y.K.Y., Y.Z. and E.C. generated animations; and Y.Z., E.C., N.G. and B.-K.T. prepared the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding authors

Correspondence to Ning Gao or Bik-Kwoon Tye.

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Supplementary information

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  1. 1.

    Supplementary Text and Figures

    Supplementary Figures 1–7

Videos

  1. 1.

    Supplementary Video 1

    Structure of the Cdt1-Mcm2-7 complex and its transition to the MCM double hexamer. The cryo-EM map of the Cdt1-MCM heptamer (AMPPNP saturated state) is shown in transparent surface representation, superimposed with color-coded subunits and Cdt1. After a continuous rotation around the channel axis, the open-ring conformation of the heptamer morphs into the closed ring conformation of the double hexamer.

  2. 2.

    Supplementary Video 2

    Proposed spring-action model for the translocation of the MCM complex on DNA. The MCM core of the CMG helicase, which translocates on DNA via a spring-action mechanism, is illustrated by animation. The repeated spring action of transitioning between the open- and closed-ring conformations translocates the helicase in an inchworm-like motion along one strand of the vertically displayed duplex DNA from bottom to top. The gap-forming Mcm5 and Mcm2 are labeled.

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DOI

https://doi.org/10.1038/nsmb.3374