Characterization of SARS-CoV main protease and identification of biologically active small molecule inhibitors using a continuous fluorescence-based assay

Richard Y. Kao; Amanda P.C. To; Louisa W.Y. Ng; Wayne H.W. Tsui; Terri S.W. Lee; Hoi Wah Tsoi; Kwok Yung Yuen

doi:10.1016/j.febslet.2004.09.026

Characterization of SARS-CoV main protease and identification of biologically active small molecule inhibitors using a continuous fluorescence-based assay

Richard Y. Kao, Amanda P.C. To, Louisa W.Y. Ng, Wayne H.W. Tsui, Terri S.W. Lee, Hoi Wah Tsoi, Kwok Yung Yuen

Research output: Contribution to journal › Article › peer-review

56 Citations (Scopus)

Abstract

Severe acute respiratory syndrome associated coronavirus main protease (SARS-CoV M^pro) has been proposed as a prime target for anti-SARS drug development. We have cloned and overexpressed the SARS-CoV M^pro in Escherichia coli, and purified the recombinant M^pro to homogeneity. The kinetic parameters of the recombinant SARS-CoV M^pro were characterized by high performance liquid chromatography-based assay and continuous fluorescence-based assay. Two novel small molecule inhibitors of the SARS-CoV M^pro were identified by high-throughput screening using an internally quenched fluorogenic substrate. The identified inhibitors have K _i values at low μM range with comparable anti-SARS-CoV activity in cell-based assays.

Original language	English
Pages (from-to)	325-330
Number of pages	6
Journal	FEBS Letters
Volume	576
Issue number	3
DOIs	https://doi.org/10.1016/j.febslet.2004.09.026
Publication status	Published - Oct 22 2004
Externally published	Yes

ASJC Scopus Subject Areas

Biophysics
Structural Biology
Biochemistry
Molecular Biology
Genetics
Cell Biology

Keywords

3C-like protease
Coronavirus
Fluorogenic substrate
Main protease
Severe acute respiratory syndrome
Small molecule inhibitor

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10.1016/j.febslet.2004.09.026

Cite this

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title = "Characterization of SARS-CoV main protease and identification of biologically active small molecule inhibitors using a continuous fluorescence-based assay",

abstract = "Severe acute respiratory syndrome associated coronavirus main protease (SARS-CoV Mpro) has been proposed as a prime target for anti-SARS drug development. We have cloned and overexpressed the SARS-CoV Mpro in Escherichia coli, and purified the recombinant Mpro to homogeneity. The kinetic parameters of the recombinant SARS-CoV Mpro were characterized by high performance liquid chromatography-based assay and continuous fluorescence-based assay. Two novel small molecule inhibitors of the SARS-CoV Mpro were identified by high-throughput screening using an internally quenched fluorogenic substrate. The identified inhibitors have K i values at low μM range with comparable anti-SARS-CoV activity in cell-based assays.",

keywords = "3C-like protease, Coronavirus, Fluorogenic substrate, Main protease, Severe acute respiratory syndrome, Small molecule inhibitor",

author = "Kao, {Richard Y.} and To, {Amanda P.C.} and Ng, {Louisa W.Y.} and Tsui, {Wayne H.W.} and Lee, {Terri S.W.} and Tsoi, {Hoi Wah} and Yuen, {Kwok Yung}",

year = "2004",

month = oct,

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doi = "10.1016/j.febslet.2004.09.026",

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journal = "FEBS Letters",

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AU - Kao, Richard Y.

AU - To, Amanda P.C.

AU - Ng, Louisa W.Y.

AU - Tsui, Wayne H.W.

AU - Lee, Terri S.W.

AU - Tsoi, Hoi Wah

AU - Yuen, Kwok Yung

PY - 2004/10/22

Y1 - 2004/10/22

N2 - Severe acute respiratory syndrome associated coronavirus main protease (SARS-CoV Mpro) has been proposed as a prime target for anti-SARS drug development. We have cloned and overexpressed the SARS-CoV Mpro in Escherichia coli, and purified the recombinant Mpro to homogeneity. The kinetic parameters of the recombinant SARS-CoV Mpro were characterized by high performance liquid chromatography-based assay and continuous fluorescence-based assay. Two novel small molecule inhibitors of the SARS-CoV Mpro were identified by high-throughput screening using an internally quenched fluorogenic substrate. The identified inhibitors have K i values at low μM range with comparable anti-SARS-CoV activity in cell-based assays.

AB - Severe acute respiratory syndrome associated coronavirus main protease (SARS-CoV Mpro) has been proposed as a prime target for anti-SARS drug development. We have cloned and overexpressed the SARS-CoV Mpro in Escherichia coli, and purified the recombinant Mpro to homogeneity. The kinetic parameters of the recombinant SARS-CoV Mpro were characterized by high performance liquid chromatography-based assay and continuous fluorescence-based assay. Two novel small molecule inhibitors of the SARS-CoV Mpro were identified by high-throughput screening using an internally quenched fluorogenic substrate. The identified inhibitors have K i values at low μM range with comparable anti-SARS-CoV activity in cell-based assays.

KW - 3C-like protease

KW - Coronavirus

KW - Fluorogenic substrate

KW - Main protease

KW - Severe acute respiratory syndrome

KW - Small molecule inhibitor

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Characterization of SARS-CoV main protease and identification of biologically active small molecule inhibitors using a continuous fluorescence-based assay

Abstract

ASJC Scopus Subject Areas

Keywords

Access to Document

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