Abstract
Objective: We aimed to determine the abilities of several drugs to block the second methylation process of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA with non-structural protein 16 (NSP16) and expose the virus to the innate immune mechanism of the host for the purpose of improving infection control and drug development for COVID-19. Methods: Recombinant prokaryotic expression plasmids PET30a-NSP16 and PET15b-NSP10 and a plasmid for preserving the untranslated region (UTR) sequences, pUC57-UTR, were constructed. The obtained UTR template was transcribed in vitro to obtain RNAs. Then, bioluminescence was used to determine the Km values of NSP16 and non-structural protein 10 (NSP10) and study the inhibition effects of four clinical drugs - cladribine, didanosine, sin efungin and ebselen - on SARS-CoV-2 NSP16 2'-O-MTase. Results: The catalytic subunit NSP16 and stimulatory subunit NSP10 of SARS-COV-2 2'-O-MTase were successfully expressed. The Km values of the substrates of NSP16, including SAM and Cap0-RNA, were also determined. Among the four drugs, sinefungin exhibited the strongest inhibitory effect and ebselen ranked second, while cladribine and didanosine showed no significant inhibitory effects according to the luminescence data. Conclusion: Four drugs with potential inhibitory activity were examined. Among them, cladribine and didanosine have weak inhibitory effects on SARS-CoV-2 NSP16 and, therefore, are not suitable for clinical application. Sinefungin has the strongest inhibitory effect, and ebselen ranks second. Therefore, they can be regarded as qualified clinical candidates for SARS-CoV-2 treatment.