2.7 Å cryo-EM structure of rotavirus core protein VP3, a unique capping machine with a helicase activity

Kumar, Dilip; Yu, Xinzhe; Crawford, Sue E.; Moreno, Rodolfo; Jakana, Joanita; Sankaran, Banumathi; Anish, Ramakrishnan; Kaundal, Soni; Hu, Liya; Estes, Mary K.; Wang, Zhao; Prasad, B. V. Venkataram

doi:10.1126/sciadv.aay6410

Title: 2.7 Å cryo-EM structure of rotavirus core protein VP3, a unique capping machine with a helicase activity

Journal Article · Wed Apr 15 00:00:00 EDT 2020 · Science Advances

DOI:https://doi.org/10.1126/sciadv.aay6410· OSTI ID:1631645

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Baylor College of Medicine, Houston, TX (United States)
Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States)

In many viruses, including rotavirus (RV), the major pathogen of infantile gastroenteritis, capping of viral messenger RNAs is a pivotal step for efficient translation of the viral genome. In RV, VP3 caps the nascent transcripts synthesized from the genomic dsRNA segments by the RV polymerase VP1 within the particle core. Here, from cryo–electron microscopy, x-ray crystallography, and biochemical analyses, we show that VP3 forms a stable tetrameric assembly with each subunit having a modular domain organization, which uniquely integrates five distinct enzymatic steps required for capping the transcripts. In addition to the previously known guanylyl- and methyltransferase activities, we show that VP3 exhibits hitherto unsuspected RNA triphosphatase activity necessary for initiating transcript capping and RNA helicase activity likely required for separating the RNA duplex formed transiently during endogenous transcription. From our studies, we propose a new mechanism for how VP3 inside the virion core caps the nascent transcripts exiting from the polymerase.

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Research Organization:: Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States)

Sponsoring Organization:: USDOE Office of Science (SC); National Institutes of Health (NIH)

Grant/Contract Number:: AC02-05CH11231; AI36040; P30 GM124169

OSTI ID:: 1631645

Journal Information:: Science Advances, Vol. 6, Issue 16; ISSN 2375-2548

Publisher:: AAASCopyright Statement

Country of Publication:: United States

Language:: English

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Cited by: 6 works

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References (51)

Conventional and unconventional mechanisms for capping viral mRNA Decroly, Etienne; Ferron, François; Lescar, Julien Nature Reviews Microbiology, Vol. 10, Issue 1 https://doi.org/10.1038/nrmicro2675	journal	December 2011
Electrostatics of nanosystems: Application to microtubules and the ribosome Baker, N. A.; Sept, D.; Joseph, S. Proceedings of the National Academy of Sciences, Vol. 98, Issue 18, p. 10037-10041 https://doi.org/10.1073/pnas.181342398	journal	August 2001
mRNA capping: biological functions and applications Ramanathan, Anand; Robb, G. Brett; Chan, Siu-Hong Nucleic Acids Research, Vol. 44, Issue 16 https://doi.org/10.1093/nar/gkw551	journal	June 2016
Structural insights into the coupling of virion assembly and rotavirus replication Trask, Shane D.; McDonald, Sarah M.; Patton, John T. Nature Reviews Microbiology, Vol. 10, Issue 3 https://doi.org/10.1038/nrmicro2673	journal	January 2012
Homologous 2',5'-phosphodiesterases from disparate RNA viruses antagonize antiviral innate immunity Zhang, R.; Jha, B. K.; Ogden, K. M. Proceedings of the National Academy of Sciences, Vol. 110, Issue 32 https://doi.org/10.1073/pnas.1306917110	journal	July 2013
In situ Structure of Rotavirus VP1 RNA-Dependent RNA Polymerase Jenni, Simon; Salgado, Eric N.; Herrmann, Tobias Journal of Molecular Biology, Vol. 431, Issue 17 https://doi.org/10.1016/j.jmb.2019.06.016	journal	August 2019
Dali server update Holm, Liisa; Laakso, Laura M. Nucleic Acids Research, Vol. 44, Issue W1 https://doi.org/10.1093/nar/gkw357	journal	April 2016
Size-Distribution Analysis of Proteins by Analytical Ultracentrifugation: Strategies and Application to Model Systems Schuck, Peter; Perugini, Matthew A.; Gonzales, Noreen R. Biophysical Journal, Vol. 82, Issue 2 https://doi.org/10.1016/S0006-3495(02)75469-6	journal	February 2002
Structure of the guanylyltransferase domain of human mRNA capping enzyme Chu, C.; Das, K.; Tyminski, J. R. Proceedings of the National Academy of Sciences, Vol. 108, Issue 25 https://doi.org/10.1073/pnas.1106610108	journal	June 2011
Atomic-accuracy models from 4.5-Å cryo-electron microscopy data with density-guided iterative local refinement DiMaio, Frank; Song, Yifan; Li, Xueming Nature Methods, Vol. 12, Issue 4 https://doi.org/10.1038/nmeth.3286	journal	February 2015
High resolution single particle refinement in EMAN2.1 Bell, James M.; Chen, Muyuan; Baldwin, Philip R. Methods, Vol. 100 https://doi.org/10.1016/j.ymeth.2016.02.018	journal	May 2016
Software tools for macromolecular microscopy Carragher, Bridget; Potter, Clinton S.; Sigworth, Fred J. Journal of Structural Biology, Vol. 157, Issue 1 https://doi.org/10.1016/j.jsb.2006.11.001	journal	January 2007
Quantitative analysis of cryo-EM density map segmentation by watershed and scale-space filtering, and fitting of structures by alignment to regions Pintilie, Grigore D.; Zhang, Junjie; Goddard, Thomas D. Journal of Structural Biology, Vol. 170, Issue 3 https://doi.org/10.1016/j.jsb.2010.03.007	journal	June 2010
An atomic model of brome mosaic virus using direct electron detection and real-space optimization Wang, Zhao; Hryc, Corey F.; Bammes, Benjamin Nature Communications, Vol. 5, Issue 1 https://doi.org/10.1038/ncomms5808	journal	September 2014
MotionCor2: anisotropic correction of beam-induced motion for improved cryo-electron microscopy Zheng, Shawn Q.; Palovcak, Eugene; Armache, Jean-Paul Nature Methods, Vol. 14, Issue 4 https://doi.org/10.1038/nmeth.4193	journal	February 2017
Rotavirus VP3 expressed in insect cells possesses guanylyltransferase activity Liu, Ming; Mattion, Nora M.; Estes, Mary K. Virology, Vol. 188, Issue 1 https://doi.org/10.1016/0042-6822(92)90736-9	journal	May 1992
Real-space refinement in PHENIX for cryo-EM and crystallography Afonine, Pavel V.; Poon, Billy K.; Read, Randy J. Acta Crystallographica Section D Structural Biology, Vol. 74, Issue 6 https://doi.org/10.1107/S2059798318006551	journal	May 2018
Rotavirus Open Cores Catalyze 5′-Capping and Methylation of Exogenous RNA: Evidence That VP3 Is a Methyltransferase Chen, Dayue; Luongo, Cindy L.; Nibert, Max L. Virology, Vol. 265, Issue 1 https://doi.org/10.1006/viro.1999.0029	journal	December 1999
Inference of Macromolecular Assemblies from Crystalline State Krissinel, Evgeny; Henrick, Kim Journal of Molecular Biology, Vol. 372, Issue 3 https://doi.org/10.1016/j.jmb.2007.05.022	journal	September 2007
UCSF Chimera?A visualization system for exploratory research and analysis Pettersen, Eric F.; Goddard, Thomas D.; Huang, Conrad C. Journal of Computational Chemistry, Vol. 25, Issue 13 https://doi.org/10.1002/jcc.20084	journal	January 2004
Quantifying the local resolution of cryo-EM density maps Kucukelbir, Alp; Sigworth, Fred J.; Tagare, Hemant D. Nature Methods, Vol. 11, Issue 1 https://doi.org/10.1038/nmeth.2727	journal	November 2013
In situ structures of rotavirus polymerase in action and mechanism of mRNA transcription and release Ding, Ke; Celma, Cristina C.; Zhang, Xing Nature Communications, Vol. 10, Issue 1 https://doi.org/10.1038/s41467-019-10236-7	journal	May 2019
Gctf: Real-time CTF determination and correction Zhang, Kai Journal of Structural Biology, Vol. 193, Issue 1 https://doi.org/10.1016/j.jsb.2015.11.003	journal	January 2016
RNA-Binding and Capping Activities of Proteins in Rotavirus Open Cores Patton, John T.; Chen, Dayue Journal of Virology, Vol. 73, Issue 2 https://doi.org/10.1128/JVI.73.2.1382-1391.1999	journal	February 1999
RNA Synthesis in a Cage—Structural Studies of Reovirus Polymerase λ3 Tao, Yizhi; Farsetta, Diane L.; Nibert, Max L. Cell, Vol. 111, Issue 5 https://doi.org/10.1016/S0092-8674(02)01110-8	journal	November 2002
Use of knowledge-based restraints in phenix.refine to improve macromolecular refinement at low resolution Headd, Jeffrey J.; Echols, Nathaniel; Afonine, Pavel V. Acta Crystallographica Section D Biological Crystallography, Vol. 68, Issue 4 https://doi.org/10.1107/S0907444911047834	journal	March 2012
Histidine Triad-like Motif of the Rotavirus NSP2 Octamer Mediates both RTPase and NTPase Activities Vasquez-Del Carpio, Rodrigo; Gonzalez-Nilo, Fernando D.; Riadi, Gonzalo Journal of Molecular Biology, Vol. 362, Issue 3 https://doi.org/10.1016/j.jmb.2006.07.050	journal	September 2006
In situ structures of RNA-dependent RNA polymerase inside bluetongue virus before and after uncoating He, Yao; Shivakoti, Sakar; Ding, Ke Proceedings of the National Academy of Sciences, Vol. 116, Issue 33 https://doi.org/10.1073/pnas.1905849116	journal	July 2019
RNA Cap Methyltransferase Activity Assay Trotman, Jackson; Schoenberg, Daniel BIO-PROTOCOL, Vol. 8, Issue 6 https://doi.org/10.21769/BioProtoc.2767	journal	January 2018
MolProbity : all-atom structure validation for macromolecular crystallography Chen, Vincent B.; Arendall, W. Bryan; Headd, Jeffrey J. Acta Crystallographica Section D Biological Crystallography, Vol. 66, Issue 1 https://doi.org/10.1107/S0907444909042073	journal	December 2009
Deciphering key features in protein structures with the new ENDscript server Robert, Xavier; Gouet, Patrice Nucleic Acids Research, Vol. 42, Issue W1 https://doi.org/10.1093/nar/gku316	journal	April 2014
Rotavirus Vaccination and the Global Burden of Rotavirus Diarrhea Among Children Younger Than 5 Years Troeger, Christopher; Khalil, Ibrahim A.; Rao, Puja C. JAMA Pediatrics, Vol. 172, Issue 10 https://doi.org/10.1001/jamapediatrics.2018.1960	journal	October 2018
RELION: Implementation of a Bayesian approach to cryo-EM structure determination Scheres, Sjors H. W. Journal of Structural Biology, Vol. 180, Issue 3, p. 519-530 https://doi.org/10.1016/j.jsb.2012.09.006	journal	December 2012
Features and development of Coot Emsley, P.; Lohkamp, B.; Scott, W. G. Acta Crystallographica Section D Biological Crystallography, Vol. 66, Issue 4 https://doi.org/10.1107/S0907444910007493	journal	March 2010
Real-time monitoring of RNA helicase activity using fluorescence resonance energy transfer in vitro Tani, Hidenori; Fujita, Osamu; Furuta, Atsushi Biochemical and Biophysical Research Communications, Vol. 393, Issue 1 https://doi.org/10.1016/j.bbrc.2010.01.100	journal	February 2010
EMRinger: side chain–directed model and map validation for 3D cryo-electron microscopy Barad, Benjamin A.; Echols, Nathaniel; Wang, Ray Yu-Ruei Nature Methods, Vol. 12, Issue 10 https://doi.org/10.1038/nmeth.3541	journal	August 2015
Location of the dsRNA-Dependent Polymerase, VP1, in Rotavirus Particles Estrozi, Leandro F.; Settembre, Ethan C.; Goret, Gaël Journal of Molecular Biology, Vol. 425, Issue 1 https://doi.org/10.1016/j.jmb.2012.10.011	journal	January 2013
UCSF ChimeraX: Meeting modern challenges in visualization and analysis: UCSF ChimeraX Visualization System Goddard, Thomas D.; Huang, Conrad C.; Meng, Elaine C. Protein Science, Vol. 27, Issue 1 https://doi.org/10.1002/pro.3235	journal	September 2017
PHENIX: a comprehensive Python-based system for macromolecular structure solution Adams, Paul D.; Afonine, Pavel V.; Bunkóczi, Gábor Acta Crystallographica Section D Biological Crystallography, Vol. 66, Issue 2, p. 213-221 https://doi.org/10.1107/S0907444909052925	journal	January 2010
Identification and Characterization of a Transcription Pause Site in Rotavirus Lawton, Jeffrey A.; Estes, Mary K.; Prasad, B. V. Venkataram Journal of Virology, Vol. 75, Issue 4 https://doi.org/10.1128/JVI.75.4.1632-1642.2001	journal	February 2001
Detection of novel members, structure-function analysis and evolutionary classification of the 2H phosphoesterase superfamily Mazumder, R. Nucleic Acids Research, Vol. 30, Issue 23 https://doi.org/10.1093/nar/gkf645	journal	December 2002
Bluetongue virus VP4 is an RNA-capping assembly line Sutton, Geoff; Grimes, Jonathan M.; Stuart, David I. Nature Structural & Molecular Biology, Vol. 14, Issue 5 https://doi.org/10.1038/nsmb1225	journal	April 2007
NMRPipe: A multidimensional spectral processing system based on UNIX pipes Delaglio, Frank; Grzesiek, Stephan; Vuister, GeertenW. Journal of Biomolecular NMR, Vol. 6, Issue 3 https://doi.org/10.1007/BF00197809	journal	November 1995
T-Coffee: a web server for the multiple sequence alignment of protein and RNA sequences using structural information and homology extension Di Tommaso, P.; Moretti, S.; Xenarios, I. Nucleic Acids Research, Vol. 39, Issue suppl https://doi.org/10.1093/nar/gkr245	journal	May 2011
3V: cavity, channel and cleft volume calculator and extractor Voss, N. R.; Gerstein, M. Nucleic Acids Research, Vol. 38, Issue Web Server https://doi.org/10.1093/nar/gkq395	journal	May 2010
Predicted Structure and Domain Organization of Rotavirus Capping Enzyme and Innate Immune Antagonist VP3 Ogden, K. M.; Snyder, M. J.; Dennis, A. F. Journal of Virology, Vol. 88, Issue 16 https://doi.org/10.1128/JVI.00923-14	journal	June 2014
Automated electron microscope tomography using robust prediction of specimen movements Mastronarde, David N. Journal of Structural Biology, Vol. 152, Issue 1 https://doi.org/10.1016/j.jsb.2005.07.007	journal	October 2005
Structural Basis for 2′-5′-Oligoadenylate Binding and Enzyme Activity of a Viral RNase L Antagonist Ogden, Kristen M.; Hu, Liya; Jha, Babal K. Journal of Virology, Vol. 89, Issue 13 https://doi.org/10.1128/JVI.00701-15	journal	April 2015
Reaction in alphavirus mRNA capping: formation of a covalent complex of nonstructural protein nsP1 with 7-methyl-GMP. Ahola, T.; Kaariainen, L. Proceedings of the National Academy of Sciences, Vol. 92, Issue 2 https://doi.org/10.1073/pnas.92.2.507	journal	January 1995
Role of Divalent Metal Cations in ATP Hydrolysis Catalyzed by the Hepatitis C Virus NS3 Helicase: Magnesium Provides a Bridge for ATP to Fuel Unwinding Frick, David N.; Banik, Sukalyani; Rypma, Ryan S. Journal of Molecular Biology, Vol. 365, Issue 4 https://doi.org/10.1016/j.jmb.2006.10.023	journal	January 2007
Cryo-electron microscopy of vitrified specimens Dubochet, Jacques; Adrian, Marc; Chang, Jiin-Ju Quarterly Reviews of Biophysics, Vol. 21, Issue 2 https://doi.org/10.1017/S0033583500004297	journal	May 1988

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