Journal
NATURE BIOTECHNOLOGY
Volume 18, Issue 3, Pages 313-316Publisher
NATURE AMERICA INC
DOI: 10.1038/73767
Keywords
phosphorylation; FRET; green fluorescent protein; imaging; PKA
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We have developed a method for visualizing phosphorylation of proteins in living cells using a novel fluorescent indicator composed of two green fluorescent protein (GFP) variants joined by the kinase-inducible domain (KID) of the transcription factor cyclic adenosine monophosphate (cAMP)-responsive element binding protein (CREB). Phosphorylation of KID by the cAMP-dependent protein kinase A (PKA) decreased the fluorescence resonance energy transfer (FRET) among the flanking GFPs, By transfecting COS-7 cells with an expression vector encoding this indicator protein (termed ART for cAMP-responsive tracer), we were able to visualize activation dynamics of PKA in living cells.
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