Kinase inhibition profiles as a tool to identify kinases for specific phosphorylation sites.

Nikolaus A Watson, Tyrell N Cartwright, Conor Lawless, Marcos Cámara-Donoso, Onur Sen, Kosuke Sako, Toru Hirota, Hiroshi Kimura, Jonathan M G Higgins

Nature communications 2020 Apr 03

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There are thousands of known cellular phosphorylation sites, but the paucity of ways to identify kinases for particular phosphorylation events remains a major roadblock for understanding kinase signaling. To address this, we here develop a generally applicable method that exploits the large number of kinase inhibitors that have been profiled on near-kinome-wide panels of protein kinases. The inhibition profile for each kinase provides a fingerprint that allows identification of unknown kinases acting on target phosphosites in cell extracts. We validate the method on diverse known kinase-phosphosite pairs, including histone kinases, EGFR autophosphorylation, and Integrin β1 phosphorylation by Src-family kinases. We also use our approach to identify the previously unknown kinases responsible for phosphorylation of INCENP at a site within a commonly phosphorylated motif in mitosis (a non-canonical target of Cyclin B-Cdk1), and of BCL9L at S915 (PKA). We show that the method has clear advantages over in silico and genetic screening.

Citation

Nikolaus A Watson, Tyrell N Cartwright, Conor Lawless, Marcos Cámara-Donoso, Onur Sen, Kosuke Sako, Toru Hirota, Hiroshi Kimura, Jonathan M G Higgins. Kinase inhibition profiles as a tool to identify kinases for specific phosphorylation sites. Nature communications. 2020 Apr 03;11(1):1684