Subunit phosphorylation and activation of phosphorylase kinase in perfused rat hearts.
The potential correlations between phosphorylase kinase subunit phosphorylation and activation have been examined using 32P-perfused rat hearts exposed to a variety of hormonal stimuli. Phosphate incorporation was measured after isolation of the enzyme by immunoprecipitation from heart extracts. Time courses of catecholamine or glucagon treatment produced a rapid rise in both the activity and the beta subunit phosphorylation of the enzyme, and a slightly slower increase in alpha' subunit phosphorylation. For short durations of catecholamine stimulation, the ratio of phosphate in the alpha' versus beta subunit was dependent upon hormone dose. After removal of hormone, both inactivation and alpha' subunit dephosphorylation were fairly slow, while the beta subunit was dephosphorylated more rapidly. For all of the above conditions, activation correlated with both alpha' and beta subunit phosphorylation. The maximum level of phosphate incorporation observed in response to hormonal stimulation is estimated to be approximately 1.3-1.7 mol of [32P]phosphate/mol of (alpha' beta gamma delta)4, divided about equally between the alpha' and beta subunits. When hearts were treated with hormone either in the absence of added calcium or in the presence of a calcium channel blocker, the time courses of subunit phosphorylation and activation were similar to those seen with standard perfusion conditions, suggesting that if any Ca2+-dependent autophosphorylation of phosphorylase kinase were occurring it does not make a major contribution to the observed hormonal responses. The complicated relationships observed here between phosphorylase kinase subunit phosphorylation and activation for the most part provide physiological affirmation of the patterns observed in vitro, but they also show some possible differences of potential interest.
D000818:Animals; D002118:Calcium / Q000502:physiology; D000242:Cyclic AMP / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D004837:Epinephrine / Q000494:pharmacology; D005934:Glucagon / Q000494:pharmacology; D007545:Isoproterenol / Q000494:pharmacology; D007700:Kinetics; D008297:Male; D009206:Myocardium / Q000201:enzymology*; D010477:Perfusion; D010764:Phosphorylase Kinase / Q000378:metabolism*; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism; D051381:Rats; D011919:Rats, Inbred Strains; D014700:Verapamil / Q000494:pharmacology
D005934:Glucagon; D014700:Verapamil; D000242:Cyclic AMP; D011494:Protein Kinases; D010764:Phosphorylase Kinase; D007545:Isoproterenol; D002118:Calcium; D004837:Epinephrine