Exploring high-resolution cryo-ET and subtomogram averaging capabilities of contemporary DEDs

Obr M, Hagen WJH, Dick RA, Yu L, Kotecha A, Schur FK. 2022. Exploring high-resolution cryo-ET and subtomogram averaging capabilities of contemporary DEDs. Journal of Structural Biology. 214(2), 107852.


Journal Article | Epub ahead of print | English

Scopus indexed
Author
Obr, MartinISTA; Hagen, Wim J.H.; Dick, Robert A.; Yu, Lingbo; Kotecha, Abhay; Schur, FlorianISTA
Department
Abstract
The potential of energy filtering and direct electron detection for cryo-electron microscopy (cryo-EM) has been well documented. Here, we assess the performance of recently introduced hardware for cryo-electron tomography (cryo-ET) and subtomogram averaging (STA), an increasingly popular structural determination method for complex 3D specimens. We acquired cryo-ET datasets of EIAV virus-like particles (VLPs) on two contemporary cryo-EM systems equipped with different energy filters and direct electron detectors (DED), specifically a Krios G4, equipped with a cold field emission gun (CFEG), Thermo Fisher Scientific Selectris X energy filter, and a Falcon 4 DED; and a Krios G3i, with a Schottky field emission gun (XFEG), a Gatan Bioquantum energy filter, and a K3 DED. We performed constrained cross-correlation-based STA on equally sized datasets acquired on the respective systems. The resulting EIAV CA hexamer reconstructions show that both systems perform comparably in the 4–6 Å resolution range based on Fourier-Shell correlation (FSC). In addition, by employing a recently introduced multiparticle refinement approach, we obtained a reconstruction of the EIAV CA hexamer at 2.9 Å. Our results demonstrate the potential of the new generation of energy filters and DEDs for STA, and the effects of using different processing pipelines on their STA outcomes.
Publishing Year
Date Published
2022-03-31
Journal Title
Journal of Structural Biology
Acknowledgement
This work was funded by the Austrian Science Fund (FWF) grant P31445 to F.K.M.S and the National Institute of Allergy and Infectious Diseases under awards R01AI147890 to R.A.D. This research was also supported by the Scientific Service Units (SSUs) of IST Austria through resources provided by Scientific Computing (SciComp), the Life Science Facility (LSF), and the Electron Microscopy Facility (EMF). We thank Dustin Morado for providing the software SubTOM for data processing. We also thank William Wan for critical reading of the manuscript and valuable feedback.
Volume
214
Issue
2
Article Number
107852
ISSN
IST-REx-ID

Cite this

Obr M, Hagen WJH, Dick RA, Yu L, Kotecha A, Schur FK. Exploring high-resolution cryo-ET and subtomogram averaging capabilities of contemporary DEDs. Journal of Structural Biology. 2022;214(2). doi:10.1016/j.jsb.2022.107852
Obr, M., Hagen, W. J. H., Dick, R. A., Yu, L., Kotecha, A., & Schur, F. K. (2022). Exploring high-resolution cryo-ET and subtomogram averaging capabilities of contemporary DEDs. Journal of Structural Biology. Elsevier. https://doi.org/10.1016/j.jsb.2022.107852
Obr, Martin, Wim J.H. Hagen, Robert A. Dick, Lingbo Yu, Abhay Kotecha, and Florian KM Schur. “Exploring High-Resolution Cryo-ET and Subtomogram Averaging Capabilities of Contemporary DEDs.” Journal of Structural Biology. Elsevier, 2022. https://doi.org/10.1016/j.jsb.2022.107852.
M. Obr, W. J. H. Hagen, R. A. Dick, L. Yu, A. Kotecha, and F. K. Schur, “Exploring high-resolution cryo-ET and subtomogram averaging capabilities of contemporary DEDs,” Journal of Structural Biology, vol. 214, no. 2. Elsevier, 2022.
Obr M, Hagen WJH, Dick RA, Yu L, Kotecha A, Schur FK. 2022. Exploring high-resolution cryo-ET and subtomogram averaging capabilities of contemporary DEDs. Journal of Structural Biology. 214(2), 107852.
Obr, Martin, et al. “Exploring High-Resolution Cryo-ET and Subtomogram Averaging Capabilities of Contemporary DEDs.” Journal of Structural Biology, vol. 214, no. 2, 107852, Elsevier, 2022, doi:10.1016/j.jsb.2022.107852.
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