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Substrate Specificity of SARS-CoV-2 Nsp10-Nsp16 Methyltransferase
R. Benoni, P. Krafcikova, MR. Baranowski, J. Kowalska, E. Boura, H. Cahová
Language English Country Switzerland
Document type Journal Article, Research Support, Non-U.S. Gov't
Grant support
(No. CZ.02.1.01/0.0/0.0/16_019/0000729
European Regional Development Fund
E-resources NLK Online Full text
Directory of Open Access Journals from 2009Free Medical Journals from 2009
PubMed Central from 2009
Europe PubMed Central from 2009
ProQuest Central from 2009-01-01
Open Access Digital Library from 2009-01-01
Open Access Digital Library from 2009-01-01
Health & Medicine (ProQuest) from 2009-01-01
ROAD: Directory of Open Access Scholarly Resources from 2009
Links
PubMed
34578302
DOI
10.3390/v13091722
Knihovny.cz E-resources
- MeSH
- Chromatography, Liquid MeSH
- COVID-19 virology MeSH
- Mass Spectrometry MeSH
- Humans MeSH
- Methyltransferases genetics metabolism MeSH
- Methylation MeSH
- Gene Expression Regulation, Viral MeSH
- RNA Caps MeSH
- RNA, Viral genetics MeSH
- SARS-CoV-2 enzymology genetics MeSH
- Substrate Specificity MeSH
- Viral Nonstructural Proteins genetics metabolism MeSH
- Viral Regulatory and Accessory Proteins genetics metabolism MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
The ongoing COVID-19 pandemic exemplifies the general need to better understand viral infections. The positive single-strand RNA genome of its causative agent, the SARS coronavirus 2 (SARS-CoV-2), encodes all viral enzymes. In this work, we focused on one particular methyltransferase (MTase), nsp16, which, in complex with nsp10, is capable of methylating the first nucleotide of a capped RNA strand at the 2'-O position. This process is part of a viral capping system and is crucial for viral evasion of the innate immune reaction. In light of recently discovered non-canonical RNA caps, we tested various dinucleoside polyphosphate-capped RNAs as substrates for nsp10-nsp16 MTase. We developed an LC-MS-based method and discovered four types of capped RNA (m7Gp3A(G)- and Gp3A(G)-RNA) that are substrates of the nsp10-nsp16 MTase. Our technique is an alternative to the classical isotope labelling approach for the measurement of 2'-O-MTase activity. Further, we determined the IC50 value of sinefungin to illustrate the use of our approach for inhibitor screening. In the future, this approach may be an alternative technique to the radioactive labelling method for screening inhibitors of any type of 2'-O-MTase.
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