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Kinase Inhibitors

Volume 795 of the series Methods in Molecular Biology pp 179-190

Date:

Covalent Cross-Linking of Kinases with Their Corresponding Peptide Substrates

  • Alexander V. StatsukAffiliated withDepartment of Chemistry, Northwestern University
  • , Kevan M. ShokatAffiliated withDepartment of Cellular and Molecular Pharmacology, University of California San Francisco Email author 

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Abstract

Protein phosphorylation represents the most dominant and evolutionary conserved posttranslational modification for information transfer in cells and organisms. The human genome encodes >500 protein kinases, and thousands of phosphorylation sites are present in mammalian proteome. To develop a global view of phosphorylation network, there is a need to map the connectivity between kinases and phosphoproteome. We developed a chemical kinase–substrate cross-linker 1 that converts transient kinase–substrate interactions into a covalently linked kinase–substrate complex in vitro and in the presence of cell lysates. The method can be applied to identify unknown upstream kinases responsible for phosphorylation events in cell lysates.

Key words

Kinase Substrate Kinase inhibitor Thiophene-2,3-dialdehyde Covalent cross-link