Dephosphorylation and activation of chicken liver acetyl-coenzyme-A carboxylase.
Studies were made on the dephosphorylation and activation of chicken liver acetyl-coenzyme-A carboxylase. The enzyme isolated by avidin-Sepharose affinity chromatography in the presence of protein phosphatase inhibitors contained 4.9 +/- 0.2 mol of alkali-labile phosphate/mol subunit and had a specific activity of 3.5 +/- 0.4 units/mg protein. The purified enzyme was dephosphorylated and activated concomitantly when incubated in the presence of protein phosphatase with a release of approximately 2 mol of phosphate/mol subunit. Limited tryptic digestion of the native and dephosphorylated forms of the enzyme (Mr 220 000) containing 4.9 and 2.9 mol of phosphate/mol of subunit, respectively, gave almost quantitatively similar polypeptides of Mr 215 000 containing 4.0 mol and 3.0 mol of phosphate per mol, respectively, which were indistinguishable by dodecylsulfate-polyacrylamide gel electrophoresis. A peptide of Mr approximately 5000 was lost from both enzymes. This result suggests that at least one of the two protein-phosphatase-labile phosphorylation sites is sensitive to trypsin. The native enzyme and those modified by protein phosphatase or by limited tryptic digestion exhibited a bi-phasic dependence on citrate. Activation of the native enzyme by protein phosphatase occurred at all the concentrations of citrate used. However, activation of the enzyme by limited tryptic digestion was found at concentrations greater than 5 mM citrate. The dephosphorylation by protein phosphatase caused an approximately fivefold activation in the enzyme activity when assayed at physiological concentrations of citrate (0.5-2.0 mM).
D000103:Acetyl-CoA Carboxylase / Q000302:isolation & purification / Q000378:metabolism*; D000469:Alkaline Phosphatase / Q000494:pharmacology; D000818:Animals; D002645:Chickens; D004789:Enzyme Activation / Q000187:drug effects; D005260:Female; D008025:Ligases / Q000378:metabolism*; D008099:Liver / Q000201:enzymology*; D008970:Molecular Weight; D010749:Phosphoprotein Phosphatases / Q000494:pharmacology; D010766:Phosphorylation; D014357:Trypsin / Q000494:pharmacology
D000469:Alkaline Phosphatase; D010749:Phosphoprotein Phosphatases; D014357:Trypsin; D008025:Ligases; D000103:Acetyl-CoA Carboxylase