Discovery of baicalin and baicalein as novel, natural product inhibitors of SARS-CoV-2 3CL protease in vitro

Haixia Su; Sheng Yao; Wenfeng Zhao; Minjun Li; Jia Liu; Weijuan Shang; Hang Xie; Changqiang Ke; Meina Gao; Kunqian Yu; Hong Liu; Jingshan Shen; Wei Tang; Leike Zhang; Jianping Zuo; Hualiang Jiang; Fang Bai; Yan Wu; Yang Ye; Yechun Xu

The amino sequence of SARS-CoV-2 3CLpro displays 96% sequence identity to SARS-CoV 3CLpro. There are 12 residues differed in two proteases and none of them participates in the direct contacts with baicalein. The high level of sequence identified between two proteases allows one to assume that baicalein will bind to the SARS-CoV 3CLpro in the same way as it does to SARS-CoV-2 3CLpro. The inhibition assay shows that baicalein can also inhibit SARS-CoV 3CLpro, with an IC50 of 1.18 ± 0.37 µM. Thus, a three-dimensional model of SARS-CoV 3CLpro in complex with baicalein was constructed by superimposing the crystal structure of SARS-CoV-2 3CLpro/baicalein with that of SARS-CoV 3CLpro/TG-0204998 (PDB code 2ZU4) (Fig. S4A ). The binding mode of baicalein with SARS-CoV 3CLpro is distinctively different from those of known inhibitors. All of the crystal structures of the inhibitor-bound SARS-CoV 3CLpro were collected for a comparison analysis (Fig. S4B ). If those peptidomimetic inhibitors are delineated like "swords" to compete with the binding of substrate, baicalein works as a "shield" in front of two catalytic dyads to prevent the approach of the substrate to the active site (Fig. S4B ). Such a unique binding mode in combination with its high ligand-binding efficiency and small molecular weight renders baicalein valuable for drug development.