Fusion protein and application thereof to construction of system for screening coronavirus 3CL protease inhibitor

Abstract

The invention discloses a fusion protein and an application thereof to construction of a system for screening a coronavirus 3CL protease inhibitor. The invention firstly provides the fusion protein. The fusion protein is formed by fusing a ubiquitination protein at an N end and a protein with a reporting function at a C end through a 3CL protease restriction enzyme cutting site amino acid fragment, wherein the 3CL protease restriction enzyme cutting site amino acid fragment is an amino acid fragment which can be recognized by 3CL protease and can be subjected to restriction enzyme cutting. A system for screening the coronavirus 3CL protease inhibitor is constructed by utilizing the fusion protein, an encoding gene of the fusion protein, a carrier containing the fusion protein or recombinant cells for expressing the fusion protein, and the system is convenient and rapid, is simple to operate and has good specificity and sensitivity; by utilizing the system, the coronavirus 3CL proteaseinhibitor can be conveniently and effectively screened, the screening mechanism is clear, the speed is high, the action effect is visual, the universality is high, the repeatability is good, the falsepositive rate is low, and the system has a good application prospect in screening of the coronavirus 3CL protease inhibitor.

Description

technical field [0001] The invention belongs to the technical field of medical virology. More specifically, it relates to a fusion protein and its application in constructing a system for screening coronavirus 3CL protease inhibitors. Background technique [0002] Coronaviruses are one of the main pathogens that infect the human respiratory system, causing severe acute respiratory syndrome (SARS), Middle East respiratory syndrome (MERS), and coronavirus disease 2019 (COVID-19). At present, there is still a lack of understanding of the mechanism of coronavirus-induced disease and the molecular and cellular biology of the disease, and there is no specific antiviral drug for this type of disease. [0003] The establishment of a drug screening model for coronavirus is an important basis for the pathogenic mechanism of coronavirus disease and its drug research. The existing drug screening model for coronavirus is mainly based on the in vitro enzyme activity detection system of t...

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Problems solved by technology

At present, the existing drug screening methods targeting 3CL protease are mostly based on computer analysis and the extracellular environment of the extracellular system (Yu-ChihLiu et al, Screening of drugs by FRET analysis identifies inhibitors of SARS-CoV 3CLprotease, 2005; Pamela Hamilletal, Development of a Red-Shifted Fluorescence-Based Assay for SARS-coronavirus 3CL Protease: Identification of a Novel Class of anti-SARS AgentsFrom the Tropical Marine Sponge Axinella Corrugata, 2006; Junwei Zhou et al, Identification of Novel Proteolytically Inactive Mutations in Coronavirus 3C-like Protease Using a Combined Approach, 2019), cannot truly reproduce the working state of 3CL protease in cells, thus limiting the process of screening highly effective inhibitory drugs and further verifying the activity of the screened drugs, making it difficult to obtain highly effective targeted inhibitory drugs

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