Cell
Volume 185, Issue 23, 10 November 2022, Pages 4347-4360.e17
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Article
A mechanism for SARS-CoV-2 RNA capping and its inhibition by nucleotide analog inhibitors

https://doi.org/10.1016/j.cell.2022.09.037Get rights and content
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Highlights

  • SARS-CoV-2 polymerase NiRAN domain catalyzes nsp9 RNAylation

  • The RNAylated nsp9 serves as an intermediate to form GpppA with GTP catalyzed by NiRAN

  • Nucleotide analog inhibitors inhibit nsp9 RNAylation and GpppA formation

  • An “induce-and-lock” mechanism for inhibitor design is proposed

Summary

Decoration of cap on viral RNA plays essential roles in SARS-CoV-2 proliferation. Here, we report a mechanism for SARS-CoV-2 RNA capping and document structural details at atomic resolution. The NiRAN domain in polymerase catalyzes the covalent link of RNA 5′ end to the first residue of nsp9 (termed as RNAylation), thus being an intermediate to form cap core (GpppA) with GTP catalyzed again by NiRAN. We also reveal that triphosphorylated nucleotide analog inhibitors can be bonded to nsp9 and fit into a previously unknown “Nuc-pocket” in NiRAN, thus inhibiting nsp9 RNAylation and formation of GpppA. S-loop (residues 50-KTN-52) in NiRAN presents a remarkable conformational shift observed in RTC bound with sofosbuvir monophosphate, reasoning an “induce-and-lock” mechanism to design inhibitors. These findings not only improve the understanding of SARS-CoV-2 RNA capping and the mode of action of NAIs but also provide a strategy to design antiviral drugs.

Keywords

SARS-CoV-2
capping
nucleotide analog inhibitor
cryo-EM

Data and code availability

The cryo-EM density maps and the structures were deposited into the Electron Microscopy Data Bank (EMDB) and Protein Data Bank (PDB) with the accession numbers 34308 and 8GWB for E-RTC:RNA-nsp9, 34310 and 8GWE for E-RTC:RNA-nsp9:GMPPNP, 34311 and 8GWF for E-RTC:RNA-nsp9:GTP, 34314 and 8GWK for E-RTC:RMP-nsp9:GMPPNP, 34317 and 8GWN for E-RTC:ATMP-nsp9:GMPPNP, 34316 and 8GEM for E-RTC:MMP-nsp9:GMPPNP, 34312 and 8GWG for E-RTC:SMP-nsp9:GMPPNP, 34313 and 8GWI for E-RTC:SMP-nsp9:GTP, 34302 and 8GW1 for E-RTC:UMP-nsp9, 34318 for 8GWO for E-RTC:UMP-nsp9:GMPPNP.

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These authors contributed equally

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