A natural product compound inhibits coronaviral replication in vitro by binding to the conserved Nsp9 SARS-CoV-2 protein

Littler, Dene R., Liu, Miaomiao, McAuley, Julie L., Lowery, Shea A., Illing, Patricia T., Gully, Benjamin S., Purcell, Anthony W., Chandrashekaran, Indu R., Perlman, Stanley, Purcell, Damian F.J., Quinn, Ronald J. and Rossjohn, Jamie ORCID: https://orcid.org/0000-0002-2020-7522 2021. A natural product compound inhibits coronaviral replication in vitro by binding to the conserved Nsp9 SARS-CoV-2 protein. Journal of Biological Chemistry 297 (6) , 101362. 10.1016/j.jbc.2021.101362

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Abstract

The Nsp9 replicase is a conserved coronaviral protein that acts as an essential accessory component of the multi-subunit viral replication/transcription complex. Nsp9 is the predominant substrate for the essential nucleotidylation activity of Nsp12. Compounds specifically interfering with this viral activity would facilitate its study. Using a native mass-spectrometry-based approach to screen a natural product library for Nsp9 binders, we identified an ent-kaurane natural product, oridonin, capable of binding to purified SARS-CoV-2 Nsp9 with micromolar affinities. By determining the crystal structure of the Nsp9-oridonin complex, we showed that oridonin binds through a conserved site near Nsp9’s C-terminal GxxxG-helix. In enzymatic assays, oridonin’s binding to Nsp9 reduces its potential to act as substrate for Nsp12’s Nidovirus RdRp-Associated Nucleotidyl transferase (NiRAN) domain. We also showed using in vitro cellular assays oridonin, while cytotoxic at higher doses has broad antiviral activity, reducing viral titer following infection with either SARS-CoV-2 or, to a lesser extent, MERS-CoV. Accordingly, these preliminary findings suggest that the oridonin molecular scaffold may have the potential to be developed into an antiviral compound to inhibit the function of Nsp9 during coronaviral replication.

Item Type:	Article
Date Type:	Publication
Status:	Published
Schools:	Medicine
Additional Information:	This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
Publisher:	American Society for Biochemistry and Molecular Biology
ISSN:	0021-9258
Date of First Compliant Deposit:	26 January 2022
Date of Acceptance:	18 October 2021
Last Modified:	21 May 2023 08:00
URI:	https://orca.cardiff.ac.uk/id/eprint/146956

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