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. 2020 Dec;35(1):145-151.
doi: 10.1080/14756366.2019.1690480.

Inhibition of SARS-CoV 3CL protease by flavonoids

Seri Jo  1 , Suwon Kim  1 , Dong Hae Shin  1 , Mi-Sun Kim  1
Affiliations

Inhibition of SARS-CoV 3CL protease by flavonoids

Seri Jo et al. J Enzyme Inhib Med Chem. 2020 Dec.

Abstract

There were severe panics caused by Severe Acute Respiratory Syndrome (SARS) and Middle-East Respiratory Syndrome-Coronavirus. Therefore, researches targeting these viruses have been required. Coronaviruses (CoVs) have been rising targets of some flavonoids. The antiviral activity of some flavonoids against CoVs is presumed directly caused by inhibiting 3C-like protease (3CLpro). Here, we applied a flavonoid library to systematically probe inhibitory compounds against SARS-CoV 3CLpro. Herbacetin, rhoifolin and pectolinarin were found to efficiently block the enzymatic activity of SARS-CoV 3CLpro. The interaction of the three flavonoids was confirmed using a tryptophan-based fluorescence method, too. An induced-fit docking analysis indicated that S1, S2 and S3' sites are involved in binding with flavonoids. The comparison with previous studies showed that Triton X-100 played a critical role in objecting false positive or overestimated inhibitory activity of flavonoids. With the systematic analysis, the three flavonoids are suggested to be templates to design functionally improved inhibitors.

Keywords: FRET; SARS-CoV; SARS-CoV 3CLpro; flavonoid; inhibitory compounds.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1.
Figure 1.
The basic skeleton structures of flavonoids and their scaffolds. Basic representative structures of the most common flavonoids classified in this study were drawn with rings and numbered positions.
Figure 2.
Figure 2.
Results from the FRET method. Each data point represents the effect of each inhibitory compound against SARS-CoV 3CLpro compared to the control. The RFU are plotted against the log-concentration of inhibitory compounds. Each dot is expressed as the mean ± standard error of the mean (n = 3). RFU: Relative Fluorescence Units.
Figure 3.
Figure 3.
Fluorescence quenching spectra of SARS-CoV 3CLpro. A solution containing 1 μM SARS-CoV 3CLpro showed a strong fluorescence emission (the solid line) with a peak at 340 nm at the excitation wavelength of 290 nm. After adding 40 μM each inhibitory compound such as herbacetin (the dashed line), rhoifolin (one dashed one dotted line) and pectolinarin (one dotted line), fluorescence quenching spectra were obtained.
Figure 4.
Figure 4.
Predicted complexes of flavonoids in the catalytic site of SARS-CoV 3CLpro. Docking poses of (A) herbacetin, kaempferol and morin and (B) rhoifolin and pectolinarin were depicted on the electrostatic surface potential of SARS-CoV 3CLpro (red, negative; blue, positive; white, uncharged). Flavonoids were predicted to occupy the active site of SARS-CoV 3CLpro. The 2D schematic representations of the interactions of five flavonoids were also drawn. Figures were created with Maestro v11.5.011. S1 represents the polar S1 site of SARS-CoV 3CLpro, S2 for the hydrophobic S2 site, and the S3′ site with no strong tendency. The pink arrows represent hydrogen bond interaction.
Figure 5.
Figure 5.
The effect of Triton X-100 on flavonoids. Each of two bars represents the inhibitory activity of compounds w/wt 0.01% Triton X-100. The first bar (shaded) represents the control. Inhibitory compounds were used at 40 μM concentration. Each bar is expressed as the mean ± standard error of the mean (n = 3). RFU: Relative Fluorescence Units.
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Grants and funding

This work was supported by the Basic Science Research Programmes, 2018R1D1A1B07050781 to DHS and 2018R1D1A1B07050942 to MK, funded by the National Research Foundation of Korea grant granted by the Ministry of Education, Science and Technology, Republic of Korea (MEST). S. Jo was supported by Brain Korea 21 (BK21) Project.