Abstract
Protein phosphorylation is one of the widely used posttranslational modifications that alter protein function in vivo. We recently showed phosphorylation of Drosophila Plexin A by cyclic adenosine monophosphate (cAMP)-dependent protein kinase (PKA) and subsequent inhibition of plexin-mediated repulsive guidance. This phosphorylation occurs in the active site of the plexin GTPase-activating protein (GAP) domain, which in turn inhibits endogenous GAP activity toward Ras/Rap family small GTP-binding proteins by recruiting the phospho-serine/threonine-binding protein 14-3-3ε. Here we describe how phosphorylation of Plexin A can be detected and quantitated using an in vitro kinase assay and radioactive [γ-P32] adenosine 5′-triphosphate (ATP).
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Acknowledgements
This work was supported by NIH (MH085923) and Welch Foundation (I-1749) grants to Jonathan Terman.
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Yang, T., Terman, J.R. (2017). Characterizing PKA-Mediated Phosphorylation of Plexin Using Purified Proteins. In: Terman, J. (eds) Semaphorin Signaling. Methods in Molecular Biology, vol 1493. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6448-2_10
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DOI: https://doi.org/10.1007/978-1-4939-6448-2_10
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