| Assay Method Information | |
| | Inhibition Test of 3CL Protease by Some Compounds of the Invention |
| Description: | The fluorescence resonance energy transfer method reported in the literature is adopted (J in et al 0.2020. Structure of Mprofrom SARS-CoV-2 and discovery of its inhibitors. Nature, 582: 289-293), the enzyme inhibitory activity of the compound is determined. The catalytic activity and initial rate of 3CL enzyme are determined by enzyme kinetics using commercially available fluorescence-labeled polypeptide MCA-AVLQSGFR-Lys (Dmp)-Lys-NH2 as substrate (GLBiochem, Shanghai). The incubation system contained 3CL protease of 2019-nCoV (0.2 μM), fluorescently labeled peptides (20 μM) and a series of concentrations of compounds to be tested (0-20 μM). The fluorescence intensity of the system during incubation for 2-3 minutes is measured by enzyme marker, and the excitation wavelength and detection wavelength are 320 mm and 405 mm, respectively. According to the change rate of the initial velocity of substrate Hydrolysis catalyzed by enzyme after the addition of inhibitor, the enzyme inhibition rate of the tested substance at different concentrations was calculated. All experiments are repeated three times, and IC 50 values of inhibitory enzymes are calculated by Prism5 software. |
| Affinity data for this assay | |
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