Related Experiment Video
Updated: Jun 26, 2026
09:30
Open Source High Content Analysis Utilizing Automated Fluorescence Lifetime Imaging Microscopy
Published on: January 18, 2017
A dual-mode fluorescence strategy for screening HAT modulators.
Nan Xie1, Emilia N Elangwe, Sabrina Asher
1Department of Chemistry, Georgia State University, PO Box 4098, Atlanta, Georgia 30302, USA.
Bioconjugate Chemistry
|January 17, 2009
Summary
This study introduces a new dual fluorescence assay to identify histone acetyltransferase (HAT) inhibitors. This method effectively detects and characterizes inhibitors for epigenetic enzymes, aiding in anticancer drug discovery.
Area of Science:
- Biochemistry
- Epigenetics
- Enzymology
Background:
- Histone acetyltransferases (HATs) are crucial epigenetic enzymes regulating gene transcription through chromatin modification.
- Dysregulation of HATs is implicated in various diseases, including cancer, making them important therapeutic targets.
Purpose of the Study:
- To develop a novel, dual-mode fluorescence assay for identifying and characterizing HAT inhibitors.
- To validate the assay's effectiveness for detecting inhibitors targeting PCAF and p300 HATs.
Main Methods:
- Utilized expressed protein ligation (EPL) to label HATs (PCAF, p300) with FRET acceptors (Dabcyl).
- Employed fluorescently labeled HAT substrate analogues (H3CoA20Mca, LysCoAMca) as FRET donors.
- Integrated fluorescence resonance energy transfer (FRET) and fluorescence polarization (anisotropy) measurements in a single assay system.
Main Results:
- Demonstrated that FRET and anisotropy measurements effectively detect HAT inhibitors.
- Showcased the assay's ability to characterize ligand-protein interactions with HATs.
- Observed decreased donor fluorescence intensity due to FRET quenching and increased fluorescence anisotropy upon ligand binding.
Conclusions:
- The developed dual fluorescence assay provides a robust platform for HAT inhibitor discovery.
- This strategy is valuable for identifying novel anticancer agents targeting HATs.
- The assay can also be applied to mechanistic studies of HAT enzymes.
