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Single-particle cryo-EM at atomic resolution

Author

Listed:
  • Takanori Nakane

    (MRC Laboratory of Molecular Biology)

  • Abhay Kotecha

    (Thermo Fisher Scientific)

  • Andrija Sente

    (MRC Laboratory of Molecular Biology)

  • Greg McMullan

    (MRC Laboratory of Molecular Biology)

  • Simonas Masiulis

    (MRC Laboratory of Molecular Biology
    Thermo Fisher Scientific)

  • Patricia M. G. E. Brown

    (MRC Laboratory of Molecular Biology)

  • Ioana T. Grigoras

    (MRC Laboratory of Molecular Biology
    Imperial College London)

  • Lina Malinauskaite

    (MRC Laboratory of Molecular Biology)

  • Tomas Malinauskas

    (University of Oxford)

  • Jonas Miehling

    (MRC Laboratory of Molecular Biology)

  • Tomasz Uchański

    (Vrije Universiteit Brussel
    VIB)

  • Lingbo Yu

    (Thermo Fisher Scientific)

  • Dimple Karia

    (Thermo Fisher Scientific)

  • Evgeniya V. Pechnikova

    (Thermo Fisher Scientific)

  • Erwin Jong

    (Thermo Fisher Scientific)

  • Jeroen Keizer

    (Thermo Fisher Scientific)

  • Maarten Bischoff

    (Thermo Fisher Scientific)

  • Jamie McCormack

    (Thermo Fisher Scientific)

  • Peter Tiemeijer

    (Thermo Fisher Scientific)

  • Steven W. Hardwick

    (University of Cambridge)

  • Dimitri Y. Chirgadze

    (University of Cambridge)

  • Garib Murshudov

    (MRC Laboratory of Molecular Biology)

  • A. Radu Aricescu

    (MRC Laboratory of Molecular Biology)

  • Sjors H. W. Scheres

    (MRC Laboratory of Molecular Biology)

Abstract

The three-dimensional positions of atoms in protein molecules define their structure and their roles in biological processes. The more precisely atomic coordinates are determined, the more chemical information can be derived and the more mechanistic insights into protein function may be inferred. Electron cryo-microscopy (cryo-EM) single-particle analysis has yielded protein structures with increasing levels of detail in recent years1,2. However, it has proved difficult to obtain cryo-EM reconstructions with sufficient resolution to visualize individual atoms in proteins. Here we use a new electron source, energy filter and camera to obtain a 1.7 Å resolution cryo-EM reconstruction for a human membrane protein, the β3 GABAA receptor homopentamer3. Such maps allow a detailed understanding of small-molecule coordination, visualization of solvent molecules and alternative conformations for multiple amino acids, and unambiguous building of ordered acidic side chains and glycans. Applied to mouse apoferritin, our strategy led to a 1.22 Å resolution reconstruction that offers a genuine atomic-resolution view of a protein molecule using single-particle cryo-EM. Moreover, the scattering potential from many hydrogen atoms can be visualized in difference maps, allowing a direct analysis of hydrogen-bonding networks. Our technological advances, combined with further approaches to accelerate data acquisition and improve sample quality, provide a route towards routine application of cryo-EM in high-throughput screening of small molecule modulators and structure-based drug discovery.

Suggested Citation

  • Takanori Nakane & Abhay Kotecha & Andrija Sente & Greg McMullan & Simonas Masiulis & Patricia M. G. E. Brown & Ioana T. Grigoras & Lina Malinauskaite & Tomas Malinauskas & Jonas Miehling & Tomasz Ucha, 2020. "Single-particle cryo-EM at atomic resolution," Nature, Nature, vol. 587(7832), pages 152-156, November.
  • Handle: RePEc:nat:nature:v:587:y:2020:i:7832:d:10.1038_s41586-020-2829-0
    DOI: 10.1038/s41586-020-2829-0
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    Citations

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    Cited by:

    1. Alister Burt & Lorenzo Gaifas & Tom Dendooven & Irina Gutsche, 2021. "A flexible framework for multi-particle refinement in cryo-electron tomography," PLOS Biology, Public Library of Science, vol. 19(8), pages 1-16, August.
    2. Alex J. Flynn & Svetlana V. Antonyuk & Robert R. Eady & Stephen P. Muench & S. Samar Hasnain, 2023. "A 2.2 Å cryoEM structure of a quinol-dependent NO Reductase shows close similarity to respiratory oxidases," Nature Communications, Nature, vol. 14(1), pages 1-12, December.
    3. Ninghai Gan & Weizhong Zeng & Yan Han & Qingfeng Chen & Youxing Jiang, 2024. "Structural mechanism of proton conduction in otopetrin proton channel," Nature Communications, Nature, vol. 15(1), pages 1-13, December.
    4. Lars V. Bock & Helmut Grubmüller, 2022. "Effects of cryo-EM cooling on structural ensembles," Nature Communications, Nature, vol. 13(1), pages 1-13, December.
    5. Simon A. Fromm & Kate M. O’Connor & Michael Purdy & Pramod R. Bhatt & Gary Loughran & John F. Atkins & Ahmad Jomaa & Simone Mattei, 2023. "The translating bacterial ribosome at 1.55 Å resolution generated by cryo-EM imaging services," Nature Communications, Nature, vol. 14(1), pages 1-9, December.
    6. Sheng Chen & Sen Zhang & Xiaoyu Fang & Liang Lin & Huiying Zhao & Yuedong Yang, 2024. "Protein complex structure modeling by cross-modal alignment between cryo-EM maps and protein sequences," Nature Communications, Nature, vol. 15(1), pages 1-14, December.
    7. Sriram Aiyer & Philip R. Baldwin & Shi Min Tan & Zelin Shan & Juntaek Oh & Atousa Mehrani & Marianne E. Bowman & Gordon Louie & Dario Oliveira Passos & Selena Đorđević-Marquardt & Mario Mietzsch & Jos, 2024. "Overcoming resolution attenuation during tilted cryo-EM data collection," Nature Communications, Nature, vol. 15(1), pages 1-19, December.
    8. Berk Küçükoğlu & Inayathulla Mohammed & Ricardo C. Guerrero-Ferreira & Stephanie M. Ribet & Georgios Varnavides & Max Leo Leidl & Kelvin Lau & Sergey Nazarov & Alexander Myasnikov & Massimo Kube & Jul, 2024. "Low-dose cryo-electron ptychography of proteins at sub-nanometer resolution," Nature Communications, Nature, vol. 15(1), pages 1-10, December.
    9. Hongcheng Fan & Bo Wang & Yan Zhang & Yun Zhu & Bo Song & Haijin Xu & Yujia Zhai & Mingqiang Qiao & Fei Sun, 2021. "A cryo-electron microscopy support film formed by 2D crystals of hydrophobin HFBI," Nature Communications, Nature, vol. 12(1), pages 1-13, December.
    10. Rebeccah A. Warmack & Ailiena O. Maggiolo & Andres Orta & Belinda B. Wenke & James B. Howard & Douglas C. Rees, 2023. "Structural consequences of turnover-induced homocitrate loss in nitrogenase," Nature Communications, Nature, vol. 14(1), pages 1-10, December.
    11. Andrew Muenks & Samantha Zepeda & Guangfeng Zhou & David Veesler & Frank DiMaio, 2023. "Automatic and accurate ligand structure determination guided by cryo-electron microscopy maps," Nature Communications, Nature, vol. 14(1), pages 1-10, December.
    12. Victoria I. Cushing & Adrian F. Koh & Junjie Feng & Kaste Jurgaityte & Alexander Bondke & Sebastian H. B. Kroll & Marion Barbazanges & Bodo Scheiper & Ash K. Bahl & Anthony G. M. Barrett & Simak Ali &, 2024. "High-resolution cryo-EM of the human CDK-activating kinase for structure-based drug design," Nature Communications, Nature, vol. 15(1), pages 1-17, December.
    13. Jing Cheng & Tong Liu & Xin You & Fa Zhang & Sen-Fang Sui & Xiaohua Wan & Xinzheng Zhang, 2023. "Determining protein structures in cellular lamella at pseudo-atomic resolution by GisSPA," Nature Communications, Nature, vol. 14(1), pages 1-9, December.
    14. Jiahua He & Tao Li & Sheng-You Huang, 2023. "Improvement of cryo-EM maps by simultaneous local and non-local deep learning," Nature Communications, Nature, vol. 14(1), pages 1-16, December.
    15. Mitsuyoshi Kamba & Ryoga Shimizu & Kiyotaka Aikawa, 2023. "Nanoscale feedback control of six degrees of freedom of a near-sphere," Nature Communications, Nature, vol. 14(1), pages 1-9, December.

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