RT Journal Article
SR Electronic
T1 Allosteric activation of SARS-CoV-2 RdRp by remdesivir triphosphate and other phosphorylated nucleotides
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 2021.01.24.428004
DO 10.1101/2021.01.24.428004
A1 Wang, Bing
A1 Svetlov, Vladimir
A1 Wolf, Yuri I
A1 Koonin, Eugene V
A1 Nudler, Evgeny
A1 Artsimovitch, Irina
YR 2021
UL http://biorxiv.org/content/early/2021/04/28/2021.01.24.428004.abstract
AB The catalytic subunit of SARS-CoV-2 RNA-dependent RNA polymerase (RdRp), Nsp12, has a unique NiRAN domain that transfers nucleoside monophosphates to the Nsp9 protein. The NiRAN and RdRp modules form a dynamic interface distant from their catalytic sites and both activities are essential for viral replication. We report that codon-optimized (for the pause-free translation) Nsp12 exists in inactive state in which NiRAN/RdRp interactions are broken, whereas translation by slow ribosomes and incubation with accessory Nsp7/8 subunits or NTPs partially rescue RdRp activity. Our data show that adenosine and remdesivir triphosphates promote synthesis of A-less RNAs, as does ppGpp, while amino acid substitutions at the NiRAN/RdRp interface augment activation, suggesting that ligand binding to the NiRAN catalytic site modulates RdRp activity. The existence of allosterically-linked nucleotidyl transferase sites that utilize the same substrates has important implications for understanding the mechanism of SARS-CoV-2 replication and design of its inhibitors.HighlightsCodon-optimization of Nsp12 triggers misfolding and activity lossSlow translation, accessory Nsp7 and Nsp8 subunits, and NTPs rescue Nsp12Non-substrate nucleotides activate RNA chain synthesis, likely via NiRAN domainCrosstalk between two Nsp12 active sites that bind the same ligandsCompeting Interest StatementThe authors have declared no competing interest.