Interfering with nucleotide excision by the coronavirus 3'-to-5' exoribonuclease

Rukesh Chinthapatla, Mohamad Sotoudegan, Pankaj Srivastava, Thomas K. Anderson, Ibrahim M. Moustafa, Kellan T. Passow, Samantha A. Kennelly, Ramkumar Moorthy, David Dulin, Joy Y. Feng, Daniel A. Harki, Robert N. Kirchdoerfer, Craig E. Cameron, Jamie J. Arnold

Research output: Contribution to journalArticlepeer-review

6 Scopus citations

Abstract

Some of the most efficacious antiviral therapeutics are ribonucleos(t)ide analogs. The presence of a 3'-to-5' proofreading exoribonuclease (ExoN) in coronaviruses diminishes the potency of many ribonucleotide analogs. The ability to interfere with ExoN activity will create new possibilities for control of SARS-CoV-2 infection. ExoN is formed by a 1:1 complex of nsp14 and nsp10 proteins. We have purified and characterized ExoN using a robust, quantitative system that reveals determinants of specificity and efficiency of hydrolysis. Double-stranded RNA is preferred over single-stranded RNA. Nucleotide excision is distributive, with only one or two nucleotides hydrolyzed in a single binding event. The composition of the terminal basepair modulates excision. A stalled SARS-CoV-2 replicase in complex with either correctly or incorrectly terminated products prevents excision, suggesting that a mispaired end is insufficient to displace the replicase. Finally, we have discovered several modifications to the 3'-RNA terminus that interfere with or block ExoN-catalyzed excision. While a 3'-OH facilitates hydrolysis of a nucleotide with a normal ribose configuration, this substituent is not required for a nucleotide with a planar ribose configuration such as that present in the antiviral nucleotide produced by viperin. Design of ExoN-resistant, antiviral ribonucleotides should be feasible.

Original languageEnglish (US)
Pages (from-to)315-336
Number of pages22
JournalNucleic acids research
Volume51
Issue number1
DOIs
StatePublished - Jan 11 2023

All Science Journal Classification (ASJC) codes

  • Genetics

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