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Cryo-EM structures of the SARS-CoV-2 endoribonuclease Nsp15 reveal insight into nuclease specificity and dynamics.

Published version
Peer-reviewed

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Authors

Frazier, Meredith N 
Dillard, Lucas B 
Williams, Jason G 
Kocaman, Seda 

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.

Description

Keywords

Amino Acid Sequence, Catalytic Domain, Cryoelectron Microscopy, Endoribonucleases, Models, Chemical, Models, Molecular, SARS-CoV-2, Uridine Triphosphate, Viral Nonstructural Proteins

Journal Title

Nat Commun

Conference Name

Journal ISSN

2041-1723
2041-1723

Volume Title

12

Publisher

Springer Science and Business Media LLC