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| | SARS-CoV-2 Mpro enzyme assay |
| Description: | The activity of SARS-Cov-2 Mpro was determined in a Fluorescence Resonance Energy Transfer (FRET)-based enzymatic assay using FRET Substrate Dabcyl-KTSAVLQSGFRKM-E(Edans)-Amide. In brief, 5 pL of test compounds (concentrations ranging from 100 pM to 0.0017 pM) was preincubated with 5 pL of 20 nM (final concentration) Mpro enzyme for 30 min at 30 °C in an assay buffer containing 20 mM HEPES, 120 mM NaCI, 0.4 mM EDTA, and 4 mM DTT and 20% glycerol. Reaction was initiated by addition of 10 pL of 20 pM (final concentration) of FRET substrate (Dabcyl-KTSAVLQSGFRKM-E (Edans)-Amide). The reaction was incubated for 1 h and the resulting fluorescent intensity was measured at Ex=360 nm/Em=490 nm at 30 °C using a SPARK 20M plate reader (Tecan). Boceprevir was used a reference standard compound. pICso and pKi were determined using 4PL GraphPad Prism and data were represented as a mean n=2± SD. |
| Affinity data for this assay | |
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