Bayfield, OW, Kilmuk, E, Winkler, DC et al. (9 more authors) (2019) Cryo-EM structure and in vitro DNA packaging of a thermophilic virus with supersized T=7 capsids. Proceedings of the National Academy of Sciences of the United States of America, 116 (9). pp. 3556-3561. ISSN 0027-8424
Abstract
Double-stranded DNA viruses, including bacteriophages and herpesviruses, package their genomes into preformed capsids, using ATP-driven motors. Seeking to advance structural and mechanistic understanding, we established in vitro packaging for a thermostable bacteriophage, P23-45 of Thermus thermophilus. Both the unexpanded procapsid and the expanded mature capsid can package DNA in the presence of packaging ATPase over the 20 °C to 70 °C temperature range, with optimum activity at 50 °C to 65 °C. Cryo-EM reconstructions for the mature and immature capsids at 3.7-Å and 4.4-Å resolution, respectively, reveal conformational changes during capsid expansion. Capsomer interactions in the expanded capsid are reinforced by formation of intersubunit β-sheets with N-terminal segments of auxiliary protein trimers. Unexpectedly, the capsid has T=7 quasi-symmetry, despite the P23-45 genome being twice as large as those of known T=7 phages, in which the DNA is compacted to near-crystalline density. Our data explain this anomaly, showing how the canonical HK97 fold has adapted to double the volume of the capsid, while maintaining its structural integrity. Reconstructions of the procapsid and the expanded capsid defined the structure of the single vertex containing the portal protein. Together with a 1.95-Å resolution crystal structure of the portal protein and DNA packaging assays, these reconstructions indicate that capsid expansion affects the conformation of the portal protein, while still allowing DNA to be packaged. These observations suggest a mechanism by which structural events inside the capsid can be communicated to the outside.
Metadata
Item Type: | Article |
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Authors/Creators: |
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Copyright, Publisher and Additional Information: | © 2019 the Author(s). Published by PNAS. This open access article is distributed under Creative Commons Attribution License 4.0 (CC BY) |
Keywords: | virus assembly; DNA packaging; capsid; portal protein; cryo-EM |
Dates: |
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Institution: | The University of Leeds |
Academic Units: | The University of Leeds > Faculty of Biological Sciences (Leeds) > School of Molecular and Cellular Biology (Leeds) > Cryo EM, Image Processing (Leeds) |
Depositing User: | Symplectic Publications |
Date Deposited: | 14 Feb 2019 10:40 |
Last Modified: | 25 Jun 2023 21:42 |
Status: | Published |
Publisher: | National Academy of Sciences |
Identification Number: | 10.1073/pnas.1813204116 |
Open Archives Initiative ID (OAI ID): | oai:eprints.whiterose.ac.uk:142474 |