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UVC inactivation of pathogenic samples suitable for cryo-EM analysis

Depelteau, Jamie S. and Renault, Ludovic and Althof, Nynke and Cassidy, C. Keith and Mendonça, Luiza M. and Jensen, Grant J. and Resch, Guenter P. and Briegel, Ariane (2022) UVC inactivation of pathogenic samples suitable for cryo-EM analysis. Communications Biology, 5 . Art. No. 29. ISSN 2399-3642. doi:10.1038/s42003-021-02962-w.

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Cryo-electron microscopy has become an essential tool to understand structure and function of biological samples. Especially for pathogens, such as disease-causing bacteria and viruses, insights gained by cryo-EM can aid in developing cures. However, due to the biosafety restrictions of pathogens, samples are often treated by chemical fixation to render the pathogen inert, affecting the ultrastructure of the sample. Alternatively, researchers use in vitro or ex vivo models, which are non-pathogenic but lack the complexity of the pathogen of interest. Here we show that ultraviolet-C (UVC) radiation applied at cryogenic temperatures can be used to eliminate or dramatically reduce the infectivity of Vibrio cholerae and the bacterial virus, the ICP1 bacteriophage. We show no discernable structural impact of this treatment of either sample using two cryo-EM methods: cryo-electron tomography followed by sub-tomogram averaging, and single particle analysis (SPA). Additionally, we applied the UVC irradiation to the protein apoferritin (ApoF), which is a widely used test sample for high-resolution SPA studies. The UVC-treated ApoF sample resulted in a 2.1 Å structure indistinguishable from an untreated published map. This research demonstrates that UVC treatment is an effective and inexpensive addition to the cryo-EM sample preparation toolbox.

Item Type:Article
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URLURL TypeDescription Paper
Depelteau, Jamie S.0000-0001-8983-2855
Renault, Ludovic0000-0001-6464-8808
Cassidy, C. Keith0000-0001-8106-1187
Jensen, Grant J.0000-0003-1556-4864
Briegel, Ariane0000-0003-3733-3725
Additional Information:© The Author(s) 2022. This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit Received 24 July 2021; Accepted 07 December 2021; Published 11 January 2022. We are grateful to Davi Ortega for helpful discussions. We also wish to thank Weng Yang and Willem Noteborn of NeCEN for assistance during data collection, and Gert Koning (Department of Fine Mechanicsl) for assistance with the construction of the prototype UVC inactivation box. This work was funded in part by the Building Blocks of Life Grant 737.016.004 from the Netherlands Organization for Scientific Research (A.B.) and Instruct-ULTRA (Horizon 2020 Coordination and Support action Number ID: 731005 (A.B. and NeCEN). Funding was also provided to C.K.C. by UK Biotechnology and Biological Sciences Research Council grant number BB/S003339/1. G.J.J. and L.M.M. was funded by NIH grant AI150464. Data availability: The structures resulting from single particle analysis of the untreated and UVC-treated ICP1 bacteriophage data (EMD-13403, EMD-13402), and the UVC-treated ApoF (EMD-13364; PDB ID: 7PF1) data have been deposited in the EMDB. Author Contributions: J.S.D., L.M.M., G.J.J., G.P.R., and A.B. designed the research, J.S.D., L.R., N.A., performed experiments, J.S.D., L.R., N.A., and C.K.C. analyzed data. J.S.D., L.R., N.A., C.K.C., and A.B. wrote the manuscript. All authors read and commented on the manuscript. The authors declare no competing interests. Peer review information: Communications Biology thanks Koji Yonekura, Andreas Boland and the other, anonymous, reviewer for their contribution to the peer review of this work. Primary Handling Editors: Janesh Kumar and Caitlin Karniski.
Funding AgencyGrant Number
Nederlandse Organisatie voor Wetenschappelijk Onderzoek (NWO)737.016.004
European Research Council (ERC)731005
Biotechnology and Biological Sciences Research Council (BBSRC)BB/S003339/1
Subject Keywords:Cryoelectron microscopy; Microbiology
Record Number:CaltechAUTHORS:20210715-140139799
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Official Citation:Depelteau, J.S., Renault, L., Althof, N. et al. UVC inactivation of pathogenic samples suitable for cryo-EM analysis. Commun Biol 5, 29 (2022).
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:109830
Deposited By: Tony Diaz
Deposited On:15 Jul 2021 17:17
Last Modified:18 Jan 2022 22:48

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