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Updated: Jan 23, 2026

Use of the Protease Fluorescent Detection Kit to Determine Protease Activity
Published on: August 4, 2009
Simultaneous assay for protease activities of hepatitis C virus and human immunodeficiency virus based on
Tsutomu Kabashima1, Keiko Tonooka2, Makoto Takada3
1Faculty of Pharmaceutical Sciences, Nagasaki International University, 2825-7 Huis Ten Bosch machi, Sasebo, Nagasaki, 859-3298, Japan. kabashima@niu.ac.jp.
Abstract:
Hepatitis C virus protease (HCV-PR) and human immunodeficiency virus protease (HIV-PR) are important for virus maturation, and thus can be used as potential target molecules for the development of antiviral drugs for the treatment of viral infections. In this study, a novel assay was developed to determine HCV-PR activity. This assay is based on a fluorogenic reaction, in which peptide fragments generated from an acetyl peptide substrate by HCV-PR can be selectively converted into a fluorescent derivative, and quantified by high-performance liquid chromatography (HPLC) with fluorescent detection. Herein, several acetyl-peptides can be used as substrates for HPLC. The application of this assay was further validated by simultaneous detection of HCV-PR and HIV-PR in a reaction mixture. The proposed method can differentiate the enzyme activities of HCV-PR and HIV-PR in a sample using their corresponding substrates. The results suggest that this assay can detect various proteases by employing set of substrate peptides under the same reaction conditions.
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