3.4 Anti-SARS-CoV-2 Mpro effects of the
newly identified covalent binders
Next, the anti-Mpro effects of these newly identified
covalent binders from LJ were validated by using the standards or the
purified compounds. Herein, a total of 20 naturally occurring
Mpro binders isolated from LJ were collected and their
anti-Mpro effects were tested one by one. As
demonstrated in Fig. 4 , among all tested natural covalent
binders, nine compounds exhibited relatively strong inhibition to the
proteolytic activity of Mpro in both time- and dose-
dependent manners. Notably, gallic acid and quercetin exhibited the
highest anti-Mpro potency with an IC50value of 6.18 μM and 9.44 μM after preincubation for 63 min. The
IC50 values for the other compounds and the positive
inhibitor (myricetin) are listed in Table 2 . The structures of
all the above constituents are shown in Fig. S4 .
3.5 Inactivation kinetics of gallic acid and quercetin
against SARS-CoV-2
Mpro
Next, the inactivation kinetics of gallic acid and quercetin were
studied via performing a suit of SARS-CoV-2
Mpro inhibition assays. The residual activity of
gallic acid and quercetin were measured by varying inhibitor
concentrations at various time intervals. The obtained data was analyzed
to determine the rate and mechanism of inactivation. As illustrated inFig. 5 , gallic acid and quercetin could inactivate the activity
of SARS-CoV-2 Mpro in dose- and time- dependent
manners. The KI value of gallic acid was
determined to be 5.63 μM, and the Kinact value
was calculated to be 0.05 min-1.
In contrast, the inactivation of
Mpro by quercetin was relatively weak with theKI value of 75.98 μM and theKinact value of 0.05 min-1.