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Cryo-EM structures and dynamics of substrate-engaged human 26S proteasome

Nature (2018) | Download Citation


The proteasome is an ATP-dependent, 2.5-megadalton machine responsible for selective protein degradation in eukaryotic cells. Here we present cryo-EM structures of the substrate-engaged human proteasome in seven conformational states at 2.8-3.6 Å resolution, captured during breakdown of a polyubiquitylated protein. These structures visualize a continuum of dynamic substrate-proteasome interactions from ubiquitin recognition to substrate translocation, during which ATP hydrolysis sequentially navigates through all six ATPases. Three principal modes of coordinated hydrolysis are observed, featuring hydrolytic events in two oppositely positioned ATPases, in two adjacent ATPases, and in one ATPase at a time. These hydrolytic modes regulate deubiquitylation, translocation initiation and processive unfolding of substrates, respectively. ATP hydrolysis powers a hinge-like motion in each ATPase that regulates its substrate interaction. Synchronization of ATP binding, ADP release and ATP hydrolysis in three adjacent ATPases drives rigid-body rotations of substrate-bound ATPases that are propagated unidirectionally in the ATPase ring and unfold the substrate.

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Author notes

  1. These authors contributed equally: Yuanchen Dong, Shuwen Zhang.


  1. State Key Laboratory for Artificial Microstructures and Mesoscopic Physics, School of Physics, Peking University, Beijing, 100871, China

    • Yuanchen Dong
    • , Shuwen Zhang
    • , Zhaolong Wu
    • , Wei Li Wang
    • , Yanan Zhu
    •  & Youdong Mao
  2. Center for Quantitative Biology, Peking University, Beijing, 100871, China

    • Shuwen Zhang
    • , Yanan Zhu
    •  & Youdong Mao
  3. Intel Parallel Computing Center for Structural Biology, Dana-Farber Cancer Institute, Boston, MA, 02215, USA

    • Yuanchen Dong
    • , Wei Li Wang
    •  & Youdong Mao
  4. Department of Cancer Immunology and Virology, Dana-Farber Cancer Institute, Department of Microbiology and Immunobiology, Harvard Medical School, Boston, MA, 02115, USA

    • Yuanchen Dong
    • , Wei Li Wang
    •  & Youdong Mao
  5. Electron Microscopy Laboratory, School of Physics, Peking University, Beijing, 100871, China

    • Xuemei Li
    •  & Youdong Mao
  6. Center for Nanoscale Systems, Harvard University, Cambridge, MA, 02138, USA

    • Svetla Stoilova-McPhie
  7. Department of Systems Biology, Harvard Medical School, Boston, MA, 02115, USA

    • Ying Lu
  8. Department of Cell Biology, Harvard Medical School, Boston, MA, 02115, USA

    • Daniel Finley


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Corresponding author

Correspondence to Youdong Mao.

Supplementary information

  1. Supplementary Information

    This file contains Supplementary Discussions regarding “Power stroke”, “Long-range quaternary allosteric regulation”, and “Functional asymmetry” along with Supplementary References

  2. Reporting Summary

  3. Video 1: Substrate processing by the human 26S proteasome.

    The complete dynamic process of the substrate engagement, deubiquitylation and translocation within the human 26S holoenzyme. The video is directly interpolated from the seven atomic structures of the substrate-bound human proteasome. The video shows smooth motions of the entire 26S holoenzyme, intuitively suggesting that the seven conformational states are on the pathway of substrate processing by the proteasome

  4. Video 2: Substrate interactions with the axial channel of the proteasomal ATPase ring.

    The closeup view of the dynamic interactions between the substrate and the ATPase ring during substrate processing, showing how the differential rotations are driven by ATP hydrolysis and how such conformational changes mechanically translate the substrate toward the pore particle (below the ATPase but not shown for clarity). The video is directly interpolated from the seven atomic structures of the substrate-bound human proteasome

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