Inactivation of Glycogen Synthetase and Activation of Phosphorylase Kinase by Muscle Adenosine 3',5'-Monophosphate-dependent Protein Kinases

Abstract

Rabbit skeletal muscle glycogen synthetase I has been purified and obtained essentially free of phosphorylase, phosphorylase kinase, and glycogen synthetase kinase. Using the purified glycogen synthetase as substrate, it was determined that two separable adenosine 3',5'-monophosphate (cyclic AMP)-dependent protein kinase fractions from skeletal muscle each catalyze the conversion of glycogen synthetase I to glycogen synthetase D; activity changes during the reaction correlate closely with the uptake of phosphate by glycogen synthetase. These same protein kinase fractions also catalyze the cyclic AMP-dependent phosphorylation and activation of phosphorylase kinase. The specific activities of the two protein kinase fractions towards phosphorylation of glycogen synthetase and phosphorylase kinase were enriched equally during purification. Further evidence that phosphorylase kinase phosphorylation and glycogen synthetase phosphorylation are common activities of the same enzyme was obtained by studying heat inactivation of the protein kinase, destabilization by cyclic AMP, inhibition by a protein inhibitor, and the cyclic nucleotide specificity. Evidence is presented that the cyclic AMP-dependent protein kinase acts directly on glycogen synthetase I and not through a second kinase as occurs in the phosphorylase activation system. Rabbit skeletal muscle glycogen synthetase I sediments as a single peak with an s20, w of 14.3. Preliminary studies indicate that the enzyme probably exists as a tetramer composed of subunits having a molecular weight between 90,000 and 100,000. On conversion of the I form to the D form, the enzyme incorporates 1 mole of phosphate per 91,000 g of protein.