Production of SARS-CoV-2 Omicron Variant Main Protease for Screening Approved Drugs as Its Potential Inhibitors
- VernacularTitle:新型冠状病毒奥密克戎变异株主蛋白酶的制备及其抑制剂的高通量筛选
- Author:
Haohao YAN
1
;
Rui ZHANG
1
;
Zhicheng LIU
1
;
Xiaoli LIU
1
;
Xiaoping LIU
1
;
Yunyu CHEN
1
Author Information
- Publication Type:Journal Article
- Keywords: SARS-CoV-2; Omicron; main protease inhibitor; fluorescence resonance energy transfer; cetylpyridinium chloride
- From: Chinese Journal of Modern Applied Pharmacy 2024;41(2):213-220
- CountryChina
- Language:Chinese
- Abstract: OBJECTIVE :To develop a high-throughput screening assay for the discovery of Omicron variant main protease(OM-Mpro) inhibitors based on the principle of fluorescence resonance energy transfer(FRET). METHODS The recombinant OM-Mpro enzyme was expressed in Escherichia coli Rosetta(DE3) cells, and further purified by a HisTrapTM chelating column. Subsequently, the enzymatic activity of OM-Mpro and wild type main protease(WT-Mpro) enzymes and inhibition of nirmatrelvir against both proteases were measured using FERT assay. With the FRET assay, OM-Mpro inhibitors were identified via high-throughput screening of an approved drug library. RESULTS The active OM-Mpro enzyme was successfully prepared from E. coli cells. OM-Mpro and WT-Mpro enzymes possessed the same enzymatic activity, and OM-Mpro remained susceptible to nirmatrelvir in vitro. Through high-throughput screening of the marketed drug library, it was found that cetylpyridinium chloride(CPC) is a mixed-type OM-Mpro inhibitor in vitro with an IC50 value of 8.76 μmol·L−1. CONCLUSION A robust FRET assay has been successfully developed based on the production of active OM-Mpro enzyme for screening of its inhibitors, and CPC is identified as a potential lead compound against OM-Mpro in vitro. This study provides a promising avenue for rapid discovery of broad-spectrum antivirals against coronavirus protease.
