Cryo-EM structures of the SARS-CoV-2 endoribonuclease Nsp15 reveal insight into nuclease specificity and dynamics
Open Access
- 27 January 2021
- journal article
- research article
- Published by Springer Science and Business Media LLC in Nature Communications
- Vol. 12 (1), 1-12
- https://doi.org/10.1038/s41467-020-20608-z
Abstract
Nsp15, a uridine specific endoribonuclease conserved across coronaviruses, processes viral RNA to evade detection by host defense systems. Crystal structures of Nsp15 from different coronaviruses have shown a common hexameric assembly, yet how the enzyme recognizes and processes RNA remains poorly understood. Here we report a series of cryo-EM reconstructions of SARS-CoV-2 Nsp15, in both apo and UTP-bound states. The cryo-EM reconstructions, combined with biochemistry, mass spectrometry, and molecular dynamics, expose molecular details of how critical active site residues recognize uridine and facilitate catalysis of the phosphodiester bond. Mass spectrometry revealed the accumulation of cyclic phosphate cleavage products, while analysis of the apo and UTP-bound datasets revealed conformational dynamics not observed by crystal structures that are likely important to facilitate substrate recognition and regulate nuclease activity. Collectively, these findings advance understanding of how Nsp15 processes viral RNA and provide a structural framework for the development of new therapeutics.This publication has 43 references indexed in Scilit:
- Bovine Pancreatic Ribonuclease: Fifty Years of the First Enzymatic Reaction MechanismBiochemistry, 2011
- Presenting your structures: theCCP4mgmolecular-graphics softwareActa crystallographica. Section D, Structural biology, 2011
- Features and development of CootActa crystallographica. Section D, Structural biology, 2010
- PHENIX: a comprehensive Python-based system for macromolecular structure solutionActa crystallographica. Section D, Structural biology, 2010
- Biochemical Characterization of Arterivirus Nonstructural Protein 11 Reveals the Nidovirus-Wide Conservation of a Replicative EndoribonucleaseJournal of Virology, 2009
- Biochemical and Genetic Analyses of Murine Hepatitis Virus Nsp15 EndoribonucleaseJournal of Virology, 2007
- Crystal Structure of a Monomeric Form of Severe Acute Respiratory Syndrome Coronavirus Endonuclease nsp15 Suggests a Role for Hexamerization as an Allosteric SwitchJournal of Virology, 2007
- New Antiviral Target Revealed by the Hexameric Structure of Mouse Hepatitis Virus Nonstructural Protein nsp15Journal of Virology, 2006
- Crystal structure and mechanistic determinants of SARS coronavirus nonstructural protein 15 define an endoribonuclease familyProceedings of the National Academy of Sciences of the United States of America, 2006
- UCSF Chimera?A visualization system for exploratory research and analysisJournal of Computational Chemistry, 2004