The thermal stabilization of the native protein structure upon ligand binding was monitored by differential scanning fluorimetry, performed according to the method described previously [34] with some modifications. Reaction mixture (25 μL) consisted of 20 mM HEPES (pH 7.5), 150 mM NaCl, 1X SYPRO orange dye (Invitrogen), 5 μM SARS-CoV nsp14 protein and 1 mM compound previously dissolved in 100% DMSO or water (SAM and Sinefungin). The protein was first mixed with compound before the addition of the dye and incubated in 96-well white microplates (Bio-Rad). Fluorescence intensities were measured from 20 to 95 °C with a ramp rate of 0.2 °C/min using a Real-Time PCR detection system (Bio-Rad). Control reactions were performed in the presence of 5% DMSO. Fluorescence intensities were plotted as a function of temperature and melting temperature (T m) was determined by curve-fitting using Prism (GraphPad software). The apparent affinity constant (KD) for compound 13 was calculated by nonlinear regression analysis with one site-specific binding equation Hill slope using Prism (GraphPad software).