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).
Original language | English (US) |
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Pages (from-to) | 147-159 |
Number of pages | 13 |
Journal | Methods in Molecular Biology |
Volume | 1493 |
DOIs | |
State | Published - 2017 |
Keywords
- Kinase assay
- PKA
- Phosphorylation
- Plexin
ASJC Scopus subject areas
- Molecular Biology
- Genetics