Reconstitution of RNA cap methylation reveals different features of SARS‐CoV‐2 and SARS‐CoV methyltransferases

Cap RNA methylations play important roles in the replication, evasion of host RNA sensor recognition, and pathogenesis. Coronaviruses possess both guanine N7‐ and 2′‐O‐ribose methyltransferases (N7‐MTase and 2′‐O‐MTase) encoded by nonstructural protein (nsp) 14 and nsp16/10 complex, respectively. In this study, we reconstituted the two‐step RNA methylations of N7‐MTase and 2′‐O‐MTase of severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) in vitro and demonstrated its common and different features in comparison with that of SARS‐CoV. We revealed that the nsp16/10 2′‐O‐MTase of SARS‐CoV‐2 has a broader substrate selectivity than the counterpart of SARS‐CoV and can accommodate both unmethylated and uncapped RNA substrates in a sequence‐independent manner. Most intriguingly, the substrate selectivity of nsp16/10 complex is not determined by the apoenzyme of nsp16 MTase but by its cofactor nsp10. These results provide insight into the unique features of SARS‐CoV‐2 MTases and may help develop strategies to precisely intervene in the methylation pathway and pathogenesis of SARS‐CoV‐2.