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Updated: Jun 4, 2026

Identification of Kinase-substrate Pairs Using High Throughput Screening
Published on: August 29, 2015
Using genetically engineered kinases to screen for novel protein kinase substrates: phosphorylation of
Abstract:
INTRODUCTIONThis protocol describes a method for detection of direct substrates of a protein kinase in cell lysates or fractions. The approach involves identification of kinase-associated substrates by immunoprecipitating a tagged form of the mutant kinase from transfected COS-1 cells and performing a kinase reaction by the addition of [γ-(32)P]ATP analog. This technique has been used for the phosphorylation of extracellular signal-regulated kinase 2 (ERK2) substrates; however, the methodology can be applied to other protein kinases as well.
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