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Updated: Mar 31, 2026

Kinase Inhibitor Screening In Self-assembled Human Protein Microarrays

Kinase Inhibitor Screening In Self-assembled Human Protein Microarrays

Published on: October 23, 2019

Screening One-Bead-One-Compound Peptide Libraries for Optimal Kinase Substrates.

Thi B Trinh¹, Dehua Pei²

¹Department of Chemistry and Biochemistry, The Ohio State University, 578 Biosciences Building, 484 West 12th Avenue, Columbus, OH, 43210, USA.

Methods in Molecular Biology (Clifton, N.J.)

|October 27, 2015

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This study introduces a novel high-throughput method to profile protein kinase sequence specificity. The technique rapidly identifies optimal kinase substrates, enabling detailed analysis of sequence context effects for serine, threonine, and tyrosine kinases.

Area of Science:

Biochemistry
Molecular Biology
Enzymology

Background:

Protein kinases regulate cellular functions by phosphorylating target proteins.
Kinase substrate specificity is often determined by the amino acid sequence surrounding the phosphorylation site.
Understanding kinase sequence specificity is crucial for deciphering cellular signaling pathways.

Purpose of the Study:

To develop and validate a robust, high-throughput method for profiling the sequence specificity of protein kinases.
To enable the identification of optimal substrates and sequence contextual effects for various kinases.

Main Methods:

Rapid synthesis of up to 10^7 unique peptides on PEGA beads using a one-bead-one-compound approach.
Kinase reactions utilizing [γ-S]ATP, followed by fluorescent labeling of thiophosphorylated peptides via disulfide exchange.

Keywords:

One-bead-one-compound library Peptide library Protein kinase Sequence specificity Substrate specificity

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Identification of Kinase-substrate Pairs Using High Throughput Screening

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Last Updated: Mar 31, 2026

Kinase Inhibitor Screening In Self-assembled Human Protein Microarrays

Kinase Inhibitor Screening In Self-assembled Human Protein Microarrays

Published on: October 23, 2019

Identification of Kinase-substrate Pairs Using High Throughput Screening

Identification of Kinase-substrate Pairs Using High Throughput Screening

Published on: August 29, 2015

Kinetic Screening of Nuclease Activity using Nucleic Acid Probes

Kinetic Screening of Nuclease Activity using Nucleic Acid Probes

Published on: November 1, 2019

Identification of active substrates using partial Edman degradation-mass spectrometry (PED-MS).

Main Results:

Successful profiling of protein kinase sequence specificity with high throughput.
Identification of optimal kinase substrates and non-permissive residues.
Demonstration of the method's applicability to protein serine, threonine, and tyrosine kinases.

Conclusions:

The developed method provides a powerful tool for comprehensive analysis of protein kinase substrate specificity.
This technique facilitates the discovery of sequence contextual effects and non-permissive residues, advancing our understanding of kinase-mediated signaling.
The method is versatile and applicable across different classes of protein kinases.