Development of a colorimetric assay for the detection of SARS-CoV-2 3CLpro activity.

Authors
Garland, Gavin D 
Harvey, Robert F 
Mulroney, Thomas E 
Monti, Mie 
Fuller, Stewart 

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Type
Article
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Abstract

Diagnostic testing continues to be an integral component of the strategy to contain the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) global pandemic, the causative agent of Coronavirus Disease 2019 (COVID-19). The SARS-CoV-2 genome encodes the 3C-like protease (3CLpro) which is essential for coronavirus replication. This study adapts an in vitro colorimetric gold nanoparticle (AuNP) based protease assay to specifically detect the activity of SARS-CoV-2 3CLpro as a purified recombinant protein and as a cellular protein exogenously expressed in HEK293T human cells. We also demonstrate that the specific sensitivity of the assay for SARS-CoV-2 3CLpro can be improved by use of an optimised peptide substrate and through hybrid dimerisation with inactive 3CLpro mutant monomers. These findings highlight the potential for further development of the AuNP protease assay to detect SARS-CoV-2 3CLpro activity as a novel, accessible and cost-effective diagnostic test for SARS-CoV-2 infection at the point-of-care. Importantly, this versatile assay could also be easily adapted to detect specific protease activity associated with other viruses or diseases conditions.

Publication Date
2022-04-29
Online Publication Date
2022-04-21
Acceptance Date
2022-04-04
Keywords
COVID 19, Coronavirus, assay development, Antiviral Agents, COVID-19, Colorimetry, Coronavirus 3C Proteases, Gold, HEK293 Cells, Humans, Metal Nanoparticles, Peptide Hydrolases, Protease Inhibitors, SARS-CoV-2
Journal Title
Biochem J
Journal ISSN
0264-6021
1470-8728
Volume Title
479
Publisher
Portland Press Ltd.
Sponsorship
Medical Research Council (MR/P008801/1)