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. 2009 Dec 14;285(7):5106–5116. doi: 10.1074/jbc.M109.070995

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FIGURE 3. — Effects of recombinant Mys on activity of purified PKA in vitro and effects of ectopic expression of Mys on protein expression in zebrafish embryos. A, effect of recombinant Mys on the activity of PKA holoenzyme. PKA activity of the holoenzyme incubated with GST, (Full)Mys-GST, (ΔPRB)Mys-GST, and cAMP (mean ± S.D., n = 3). B, pull-down assay of (Full)Mys-GST and (ΔPRB)Mys-GST incubated with PKA holoenzyme showing interaction of (Full)Mys-GST with Prkar1 but not with Prkac. Immunoblotting of loading control (Load) and precipitations with glutathione-Sepharose beads (GSH) of a mixture of Mys-GST and PKA holoenzyme. C, effects of (Full)Mys-GST on PKA holoenzyme formation. Immunoprecipitations with anti-Prkac antibody probed with anti-Prkac (α-Prkac) and anti-Prkar1a (α-Prkar1a) antibodies showing that increasing the amounts of (Full)Mys-GST decreases the amounts of Prkar1a binding to Prkac. D, effects of ectopic expression of mutant Mys-GFP on expression of endogenous Mys in zebrafish embryos. Immunoblotting of extracts from wild-type embryos (−) and embryos injected with mys-(1–200)-gfp mRNA or mys-(201–435)-gfp mRNA showing reduction of endogenous Mys in embryos expressing Mys-(201–435)-GFP.