Fig. 1.
SARS-CoV-2 3CLpro expression and characterization. (a) SDS-PAGE of 3CLpro; the calculated molecular weight of the 3CLpro is 33,796 Da. (b) Michaelis-Menten plot of 15 nM 3CLpro with various concentrations of FRET substrate (c) Initial velocity plot of enzyme reaction with various concentrations of 3CLpro. (d) Principle of the enzymatic FRET-based assay used to monitor 3CLpro activity. Once the fluorogenic substrate is cleaved by the enzyme, the fluorophore (Edans) and the fluorescence quencher (Dabcyl) are spatially separated, resulting in an increase in fluorescence which is proportional to the enzyme activity. (e) SARS-CoV-2 3CLpro inhibition by reference compounds. 3CLpro was pre-incubated 30 min with various concentration of GC-376 or boceprevir before FRET substrate addition and initial velocity measurement. Data are shown as mean ± SD of duplicates from representative experiments.