Generic phosphatase activity detection using zinc mediated aggregation modulation of polypeptide-modified gold
Robert Selegård1, Karin Enander, Daniel Aili
1Division of Molecular Physics, Department of Physics, Chemistry and Biology, Linköping University, SE-583 36 Linköping, Sweden. daniel.aili@liu.se.
Nanoscale
|September 6, 2014
Summary
This study presents a novel, generic assay for detecting phosphatase activity. The method decouples enzyme recognition from nanoparticle changes, enabling broad application in drug screening and diagnostics.
Area of Science:
- Biochemistry
- Nanotechnology
- Assay Development
Background:
- Plasmonic nanoparticle assays often link colloidal stability changes directly to biomolecular recognition.
- This necessitates new assay strategies for each specific substrate and enzyme.
Purpose of the Study:
- To develop a generic strategy for monitoring phosphatase activity.
- To decouple substrate recognition from nanoparticle stability modulation for a universal detection scheme.
Main Methods:
- Utilized inorganic phosphate generation by phosphatase activity to form an insoluble complex with Zn(2+).
- Measured the reduction in dissolved Zn(2+) concentration, which impacts the aggregation of gold nanoparticles functionalized with a Zn(2+)-responsive polypeptide.
- Employed a single detection scheme adaptable to various enzymes and substrates.
Main Results:
- Demonstrated a rapid and sensitive readout of phosphatase activity through changes in nanoparticle stability.
- Successfully applied the assay to three distinct phosphatases and five different substrates.
- Showcased the assay's versatility and independence from specific enzyme-substrate pairs.
Conclusions:
- The developed assay provides a universal platform for phosphatase activity detection.
- This generic strategy is highly valuable for drug screening and diagnostic applications.
- Decoupling recognition from signal generation offers a robust and adaptable assay design.
