The insulin-directed phosphorylation site on ATP-citrate lyase is identical with the site phosphorylated by the cAMP-dependent protein kinase in vitro.
32P-labeled ATP-citrate lyase isolated from 32P-labeled hepatocytes treated with insulin contained 1.6-1.8-fold greater 32P-radioactivity per mg protein than control enzyme. Both enzyme preparations were digested in parallel with trypsin until 94% of all 32P-radioactivity was rendered acid soluble. Quantitative high performance liquid chromatographic peptide mapping of the tryptic digests revealed a principal 32P-peptide which accounted for at least 80% of the insulin induced increment in 32P-radioactivity of native lyase. This peptide was purified, sequenced, and the site of 32P-phosphorylation assigned by two methods: electrophoresis (pH 6.5) of residual peptide after each step of Edman degradation and solid phase sequencing. The site of insulin-directed phosphorylation of ATP-citrate lyase (Thr-Ala-Ser(32P)-Phe-Ser-Glu-Ser-Arg) is the same as that directed by glucagon, and, in turn, identical with that phosphorylated by the cAMP-dependent protein kinase in vitro.
D001275:ATP Citrate (pro-S)-Lyase / Q000378:metabolism*; D000818:Animals; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000494:pharmacology; D007328:Insulin / Q000494:pharmacology*; D008099:Liver / Q000201:enzymology; D008297:Male; D010446:Peptide Fragments / Q000032:analysis; D010748:Phosphopeptides / Q000032:analysis; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains; D014357:Trypsin
D007328:Insulin; D010446:Peptide Fragments; D010748:Phosphopeptides; D000242:Cyclic AMP; D001275:ATP Citrate (pro-S)-Lyase; D011494:Protein Kinases; D014357:Trypsin