Sensitive multiplexed analysis of kinase activities and activity-based kinase identification. Kubota, K., Anjum, R., Yu, Y., Kunz, R. C, Andersen, J. N, Kraus, M., Keilhack, H., Nagashima, K., Krauss, S., Paweletz, C., Hendrickson, R. C, Feldman, A. S, Wu, C., Rush, J., Villén, J., & Gygi, S. P Nature biotechnology, 27(10):933–940, October, 2009. ![Sensitive multiplexed analysis of kinase activities and activity-based kinase identification [link]](https://bibbase.org/img/filetypes/link.svg "link")
Paper doi abstract bibtex Constitutive activation of one or more kinase signaling pathways is a hallmark of many cancers. Here we extend the previously described mass spectrometry–based KAYAK approach by monitoring kinase activities from multiple signaling pathways simultaneously. This improved single-reaction strategy, which quantifies the phosphorylation of 90 synthetic peptides in a single mass spectrometry run, is compatible with nanogram to microgram amounts of cell lysate. Furthermore, the approach enhances kinase monospecificity through substrate competition effects, faithfully reporting the signatures of many signaling pathways after mitogen stimulation or of basal pathway activation differences across a panel of well-studied cancer cell lines. Hierarchical clustering of activities from related experiments groups peptides phosphorylated by similar kinases together and, when combined with pathway alteration using pharmacological inhibitors, distinguishes underlying differences in potency, off-target effects and genetic backgrounds. Finally, we introduce a strategy to identify the kinase, and even associated protein complex members, responsible for phosphorylation events of interest.
@article{kubota_sensitive_2009,
title = {Sensitive multiplexed analysis of kinase activities and activity-based kinase identification},
volume = {27},
issn = {1087-0156},
url = {https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3129615/},
doi = {10.1038/nbt.1566},
abstract = {Constitutive activation of one or more kinase signaling pathways is a hallmark of many cancers. Here we extend the previously described mass spectrometry–based KAYAK approach by monitoring kinase activities from multiple signaling pathways simultaneously. This improved single-reaction strategy, which quantifies the phosphorylation of 90 synthetic peptides in a single mass spectrometry run, is compatible with nanogram to microgram amounts of cell lysate. Furthermore, the approach enhances kinase monospecificity through substrate competition effects, faithfully reporting the signatures of many signaling pathways after mitogen stimulation or of basal pathway activation differences across a panel of well-studied cancer cell lines. Hierarchical clustering of activities from related experiments groups peptides phosphorylated by similar kinases together and, when combined with pathway alteration using pharmacological inhibitors, distinguishes underlying differences in potency, off-target effects and genetic backgrounds. Finally, we introduce a strategy to identify the kinase, and even associated protein complex members, responsible for phosphorylation events of interest.},
number = {10},
urldate = {2022-01-30},
journal = {Nature biotechnology},
author = {Kubota, Kazuishi and Anjum, Rana and Yu, Yonghao and Kunz, Ryan C and Andersen, Jannik N and Kraus, Manfred and Keilhack, Heike and Nagashima, Kumiko and Krauss, Stefan and Paweletz, Cloud and Hendrickson, Ronald C and Feldman, Adam S and Wu, Chin-Lee and Rush, John and Villén, Judit and Gygi, Steven P},
month = oct,
year = {2009},
pmid = {19801977},
pmcid = {PMC3129615},
pages = {933--940},
}
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