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Assaying Protein Kinase Activity with Radiolabeled ATP
Published on: May 26, 2017
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Probing Protein Kinase-ATP Interactions Using a Fluorescent ATP Analog
Leslie E W LaConte1, Sarika Srivastava1, Konark Mukherjee2,3
1Virginia Tech Carilion Research Institute, Roanoke, 2 Riverside Circle, Roanoke, VA, 24016, USA.
Methods in Molecular Biology (Clifton, N.J.)
|August 16, 2017
Summary
This study presents a simple fluorescence assay to characterize adenosine triphosphate (ATP) binding to eukaryotic protein kinases. This method aids in understanding kinase function and developing targeted molecular inhibitors.
Area of Science:
- Biochemistry
- Enzymology
- Molecular Biology
Background:
- Eukaryotic protein kinases are crucial enzymes and significant drug targets.
- Understanding adenosine triphosphate (ATP) binding regulation is vital for kinase function and inhibitor design.
Purpose of the Study:
- To provide a general, adaptable procedure for characterizing ATP binding to eukaryotic protein kinases.
- To identify conditions that activate specific protein kinases.
Main Methods:
- Utilizes a simple fluorescence-based assay.
- Employs fluorescent ATP analogs (e.g., TNP-ATP, MANT-ATP).
- Monitors changes in fluorescence to assess nucleotide binding.
Main Results:
- The assay is straightforward to perform and interpret.
- It allows for the optimization of conditions affecting kinase-nucleotide interactions.
- Provides a method to characterize ATP binding kinetics.
Conclusions:
- This fluorescence assay offers a practical approach to study eukaryotic protein kinase activity.
- The method facilitates the discovery of kinase-specific inhibitors.
- It is adaptable for probing various activation conditions for kinases.
