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An Improved Ratiometric FRET Biosensor with Higher Affinity for Extracellular ATP
Autumn Cholger1, Jason M Conley1, Elaine Colomb1
1Department of Chemistry & Interdisciplinary Life Science Program, Purdue University, West Lafayette, IN 47907, USA.
Sensors (Basel, Switzerland)
|September 27, 2025
Summary
Researchers developed ECATS2, a new biosensor for extracellular adenosine triphosphate (ATP). This improved tool offers higher affinity for detecting purinergic signaling and ATP release in cells.
Area of Science:
- Biochemistry
- Cell Biology
- Neuroscience
Background:
- Extracellular adenosine triphosphate (ATP) acts as a key purinergic signaling molecule.
- Previous genetically encoded Förster Resonance Energy Transfer (FRET) biosensors could detect micromolar levels of extracellular ATP.
Purpose of the Study:
- To develop a second-generation biosensor with enhanced affinity for extracellular ATP.
- To optimize the biosensor for improved detection of purinergic signaling dynamics.
Main Methods:
- Site-directed mutagenesis of the ATP binding site.
- Optimization of cell-surface display and tether length.
- Development of the ECATS2 FRET biosensor.
Main Results:
- The ECATS2 biosensor exhibits over three-fold higher affinity for extracellular ATP compared to previous sensors.
- Tether length was identified as a critical factor for biosensor performance optimization.
- ECATS2 successfully detected extracellular ATP release in cultured astrocytes under hypoosmotic stress.
Conclusions:
- ECATS2 represents a significant advancement in tools for detecting extracellular ATP.
- The enhanced sensor enables more sensitive ratiometric detection of purinergic signaling.
- This tool facilitates the study of extracellular ATP dynamics in various biological contexts.
