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Challenges in solving structures from radiation-damaged tomograms of protein nanocrystals assessed by simulation

Peck, Ariana and Yao, Qing and Brewster, Aaron S. and Zwart, Petrus H. and Heumann, John M. and Sauter, Nicholas K. and Jensen, Grant J. (2020) Challenges in solving structures from radiation-damaged tomograms of protein nanocrystals assessed by simulation. . (Unpublished)

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Structure determination methods are needed to resolve the atomic details that underlie protein function. X-ray crystallography has provided most of our knowledge of protein structure but is constrained by the need for large, well-ordered crystals and the loss of phase information. The rapidly developing methods of serial femtosecond crystallography, micro-electron diffraction, and single-particle reconstruction circumvent the first of these limitations by enabling data collection from nanocrystals or purified proteins. However, the first two methods also suffer from the phase problem, while many proteins fall below the molecular weight threshold required by single-particle reconstruction. Cryo-electron tomography of protein nanocrystals has the potential to overcome these obstacles of mainstream structure determination methods. Here we present a data processing scheme that combines routines from X-ray crystallography and new algorithms we developed to solve structures from tomograms of nanocrystals. This pipeline handles image processing challenges specific to tomographic sampling of periodic specimens and is validated using simulated crystals. We also assess the tolerance of this workflow to the effects of radiation damage. Our simulations indicate a trade-off between a wider tilt-range to facilitate merging data from multiple tomograms and a smaller tilt increment to improve phase accuracy. Since phase errors but not merging errors can be overcome with additional datasets, these results recommend distributing the dose over a wide angular range rather than using a finer sampling interval to solve the protein structure.

Item Type:Report or Paper (Discussion Paper)
Related URLs:
URLURL TypeDescription Paper ItemCode
Peck, Ariana0000-0002-5940-3897
Yao, Qing0000-0003-3575-9909
Jensen, Grant J.0000-0003-1556-4864
Additional Information:The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC-ND 4.0 International license. This version posted September 20, 2020. We thank Lauren Ann Metskas and Florian Schur for valuable discussions, in addition to David Stokes and Steven Ludtke for advice on the phase splitting phenomenon. A.P. is The Mark Foundation for Cancer Research Fellow of the Damon Runyon Cancer Research Foundation (DRG 2361-19). This work was supported by NIH grants R35 GM122588 (to G.J.J), AI150464 (to G.J.J), and GM117126 (to N.K.S). Code availability: The code developed to process tomograms of nanocrystals is available at
Funding AgencyGrant Number
Damon Runyon Cancer Research FoundationDRG 2361-19
NIHR35 GM122588
Record Number:CaltechAUTHORS:20200922-102800972
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Official Citation:Challenges in solving structures from radiation-damaged tomograms of protein nanocrystals assessed by simulation Ariana Peck, Qing Yao, Aaron S. Brewster, Petrus H. Zwart, John M. Heumann, Nicholas K. Sauter, Grant J. Jensen bioRxiv 2020.09.18.298562; doi:
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:105468
Deposited By: Tony Diaz
Deposited On:22 Sep 2020 17:43
Last Modified:18 Nov 2020 00:08

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