Abstract
The human proteome is known to contain >500 protein kinases, which regulate almost all facets of cellular biology by the post-translational attachment of a phosphate moiety to serine, threonine, or tyrosine residues within a substrate protein. Most protein kinases remain poorly characterized and, as a result, current studies are directed toward defining their target substrates experimentally to gain a comprehensive view of the signaling proteins and pathways modulated by these kinases. Herein, we describe a rapid and convenient method for elucidating the consensus substrate motif for phosphorylation by a protein kinase using peptide SPOT arrays that are custom-synthesized on a cellulose membrane support. The definition of the target consensus motif provides an important starting point for the identification of physiologically relevant kinase substrates.
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Acknowledgments
We thank C.S. Ho and I.M. Blasutig for helpful comments on the manuscript, and T. Pawson for use of his Multipep SPOT synthesizer. This work was supported by grants from the National Cancer Institute of Canada and the Canadian Institutes of Health Research to F.S., and from the Terry Fox Foundation through an award from National Cancer Institute of Canada to GCL. JMM is a recipient of a CJ Martin (Biomedical) Fellowship from the National Health and Medical Research Council of Australia.
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© 2009 Humana Press, a part of Springer Science+Business Media, LLC
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Leung, G.C., Murphy, J.M., Briant, D., Sicheri, F. (2009). Characterization of Kinase Target Phosphorylation Consensus Motifs Using Peptide SPOT Arrays. In: Cretich, M., Chiari, M. (eds) Peptide Microarrays. Methods in Molecular Biology™, vol 570. Humana Press. https://doi.org/10.1007/978-1-60327-394-7_7
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DOI: https://doi.org/10.1007/978-1-60327-394-7_7
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