SARS-CoV-2 nsp10-nsp16 complex is a 2'-O-methyltransferase (MTase) involved in viral RNA capping, enabling the virus to evade the immune system in humans. It has been considered a valuable target in the discovery of antiviral therapeutics, as the RNA cap formation is crucial for viral propagation. Through cross-screening of the inhibitors that we previously reported for SARS-CoV-2 nsp14 MTase activity against nsp10-nsp16 complex, we identified two compounds (SS148 and WZ16) that also inhibited nsp16 MTase activity. To further enable the chemical optimization of these two compounds towards more potent and selective dual nsp14/nsp16 MTase inhibitors, we determined the crystal structure of nsp10-nsp16 in complex with each of SS148 and WZ16. As expected, the structures revealed the binding of both compounds to S-adenosyl-L-methionine (SAM) binding pocket of nsp16. However, our structural data along with the biochemical mechanism of action determination revealed an RNA-dependent SAM-competitive pattern of inhibition for WZ16, clearly suggesting that binding of the RNA first may help the binding of some SAM competitive inhibitors. Both compounds also showed some degree of selectivity against human protein MTases, an indication of great potential for chemical optimization towards more potent and selective inhibitors of coronavirus MTases.

• Keywords
• COVID-19, SARS-CoV-2, SS148, WZ16, nsp10, nsp16,
• MeSH
• COVID-19 Drug Treatment * MeSH
• Humans MeSH
• Methyltransferases chemistry   MeSH
• RNA, Viral metabolism   MeSH
• SARS-CoV-2 * MeSH
• Viral Nonstructural Proteins chemistry   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Methyltransferases MeSH
• RNA, Viral MeSH
• Viral Nonstructural Proteins MeSH

Coronaviral methyltransferases (MTases), nsp10/16 and nsp14, catalyze the last two steps of viral RNA-cap creation that takes place in cytoplasm. This cap is essential for the stability of viral RNA and, most importantly, for the evasion of innate immune system. Non-capped RNA is recognized by innate immunity which leads to its degradation and the activation of antiviral immunity. As a result, both coronaviral MTases are in the center of scientific scrutiny. Recently, X-ray and cryo-EM structures of both enzymes were solved even in complex with other parts of the viral replication complex. High-throughput screening as well as structure-guided inhibitor design have led to the discovery of their potent inhibitors. Here, we critically summarize the tremendous advancement of the coronaviral MTase field since the beginning of COVID pandemic.

• MeSH
• Amino Acids chemistry   MeSH
• Coronavirus drug effects   enzymology   genetics   MeSH
• Humans MeSH
• Methyltransferases antagonists & inhibitors   chemistry   metabolism   MeSH
• Methylation MeSH
• Molecular Conformation MeSH
• Models, Molecular MeSH
• Molecular Structure MeSH
• Drug Discovery MeSH
• RNA, Viral chemistry   genetics   metabolism   MeSH
• Amino Acid Sequence MeSH
• Protein Binding MeSH
• Binding Sites MeSH
• Structure-Activity Relationship MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Amino Acids MeSH
• Methyltransferases MeSH
• RNA, Viral MeSH