Journal of Biological Chemistry · 1981 · 83 citations · 20 references
Life SciencesBiochemistryMolecular BiologyEnzymologyMetabolismProtein BiochemistryEnergy MetabolismGlucagon Signaling In MetabolismProtein PhosphorylationProtein SynthesisCamp-protein Kinase A SignalingDigestionKinase Substrate SpecificityPhosphopeptide IdentificationPhosphorylation Site MappingProtein Phosphorylation AnalysisProtein Phosphorylation In MetabolismProtein Phosphosite Mapping
[32P]ATP-citrate lyase phosphorylated by the cAMP-dependent protein kinase was partially digested by trypsin. Two tryptic 32P-labeled phosphopeptides containing more than 90% of the 32P radioactivity present on the phosphorylated enzyme were purified and found to have overlapping amino acid sequences around the same phosphorylated site (Thr-Ala-Ser(32P)-Phe-Ser-Glu-Ser-Arg). Tryptic digestion of 32P-labeled ATP-citrate lyase purified from 32P-labeled hepatocytes exposed to glucagon yielded a major 32P-labeled peptide of identical amino acid composition with that indicated above. Thus, the site on ATP-citrate lyase phosphorylated by the cAMP-dependent protein kinase in vitro resides on the same octapeptide as the site of glucagon-stimulated phosphorylation in intact hepatocytes.
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