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A fluorogenic histone deacetylase assay well suited for high-throughput activity screening
Dennis Wegener1, Frank Wirsching, Daniel Riester
1Department of Molecular Genetics and Preparative Molecular Biology, Institute for Microbiology and Genetics, Grisebachstrasse 8, 37077 Goettingen, Germany.
Chemistry & Biology
|February 8, 2003
Summary
Researchers developed a new fluorogenic assay to measure histone deacetylase (HDAC) activity. This assay enables sensitive detection and facilitates high-throughput screening for potential cancer drugs targeting HDACs.
Area of Science:
- Biochemistry
- Molecular Biology
- Enzymology
Background:
- Histone deacetylases (HDACs) are crucial enzymes regulating gene expression in eukaryotes.
- HDACs are increasingly recognized as potential therapeutic targets for cancer treatment.
- A sensitive assay for HDAC activity is needed for research and drug discovery.
Purpose of the Study:
- To develop a novel, sensitive, and homogeneous fluorogenic assay for measuring HDAC activity.
- To create a tool for expediting studies on HDACs in transcriptional regulation.
- To facilitate high-throughput screening of potential HDAC inhibitors for drug discovery.
Main Methods:
- Synthesis of novel fluorogenic substrates containing an epsilon-acetylated lysyl moiety and an MCA moiety.
- Assay principle relies on sequential deacetylation by HDACs and subsequent trypsin cleavage.
- Release of highly fluorescent AMC molecules upon enzymatic activity, measured by fluorescence intensity.
Main Results:
- The assay demonstrated a direct correlation between fluorescence intensity and HDAC activity.
- The developed fluorogenic substrates are effective for detecting HDAC activity.
- The assay is suitable for nonisotopic, homogeneous, and high-throughput screening.
Conclusions:
- A novel fluorogenic assay for HDAC activity has been successfully developed.
- This assay provides a sensitive and convenient method for studying HDACs.
- The assay is well-suited for high-throughput screening of HDAC inhibitors in drug discovery efforts.