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Low-dose cryo-electron ptychography of proteins at sub-nanometer resolution

Author

Listed:
  • Berk Küçükoğlu

    (Rte. de la Sorge)

  • Inayathulla Mohammed

    (Rte. de la Sorge)

  • Ricardo C. Guerrero-Ferreira

    (Rte. de la Sorge
    1521 Dickey Drive NE)

  • Stephanie M. Ribet

    (Lawrence Berkeley National Laboratory)

  • Georgios Varnavides

    (Lawrence Berkeley National Laboratory
    University of California)

  • Max Leo Leidl

    (Ernst Ruska-Centre for Microscopy and Spectroscopy with Electrons (ER-C-3): Structural Biology
    Butenandstr. 11)

  • Kelvin Lau

    (Rte Cantonale)

  • Sergey Nazarov

    (EPFL VPA DCI-Lausanne)

  • Alexander Myasnikov

    (EPFL VPA DCI-Lausanne)

  • Massimo Kube

    (Rte. de la Sorge)

  • Julika Radecke

    (Rte. de la Sorge)

  • Carsten Sachse

    (Ernst Ruska-Centre for Microscopy and Spectroscopy with Electrons (ER-C-3): Structural Biology
    Heinrich Heine University)

  • Knut Müller-Caspary

    (Butenandstr. 11)

  • Colin Ophus

    (Lawrence Berkeley National Laboratory)

  • Henning Stahlberg

    (Rte. de la Sorge)

Abstract

Cryo-transmission electron microscopy (cryo-EM) of frozen hydrated specimens is an efficient method for the structural analysis of purified biological molecules. However, cryo-EM and cryo-electron tomography are limited by the low signal-to-noise ratio (SNR) of recorded images, making detection of smaller particles challenging. For dose-resilient samples often studied in the physical sciences, electron ptychography – a coherent diffractive imaging technique using 4D scanning transmission electron microscopy (4D-STEM) – has recently demonstrated excellent SNR and resolution down to tens of picometers for thin specimens imaged at room temperature. Here we apply 4D-STEM and ptychographic data analysis to frozen hydrated proteins, reaching sub-nanometer resolution 3D reconstructions. We employ low-dose cryo-EM with an aberration-corrected, convergent electron beam to collect 4D-STEM data for our reconstructions. The high frame rate of the electron detector allows us to record large datasets of electron diffraction patterns with substantial overlaps between the interaction volumes of adjacent scan positions, from which the scattering potentials of the samples are iteratively reconstructed. The reconstructed micrographs show strong SNR enabling the reconstruction of the structure of apoferritin protein at up to 5.8 Å resolution. We also show structural analysis of the Phi92 capsid and sheath, tobacco mosaic virus, and bacteriorhodopsin at slightly lower resolutions.

Suggested Citation

  • 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.
  • Handle: RePEc:nat:natcom:v:15:y:2024:i:1:d:10.1038_s41467-024-52403-5
    DOI: 10.1038/s41467-024-52403-5
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    References listed on IDEAS

    as
    1. 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.
    2. Benedikt Diederichs & Ziria Herdegen & Achim Strauch & Frank Filbir & Knut Müller-Caspary, 2024. "Exact inversion of partially coherent dynamical electron scattering for picometric structure retrieval," Nature Communications, Nature, vol. 15(1), pages 1-9, December.
    3. Philipp M. Pelz & Sinéad M. Griffin & Scott Stonemeyer & Derek Popple & Hannah DeVyldere & Peter Ercius & Alex Zettl & Mary C. Scott & Colin Ophus, 2023. "Solving complex nanostructures with ptychographic atomic electron tomography," Nature Communications, Nature, vol. 14(1), pages 1-9, December.
    4. Ka Man Yip & Niels Fischer & Elham Paknia & Ashwin Chari & Holger Stark, 2020. "Atomic-resolution protein structure determination by cryo-EM," Nature, Nature, vol. 587(7832), pages 157-161, November.
    5. Xudong Pei & Liqi Zhou & Chen Huang & Mark Boyce & Judy S. Kim & Emanuela Liberti & Yiming Hu & Takeo Sasaki & Peter D. Nellist & Peijun Zhang & David I. Stuart & Angus I. Kirkland & Peng Wang, 2023. "Cryogenic electron ptychographic single particle analysis with wide bandwidth information transfer," Nature Communications, Nature, vol. 14(1), pages 1-10, December.
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