Subunit phosphorylation and activation of skeletal muscle phosphorylase kinase by the cAMP-dependent protein kinase. Divalent metal ion, ATP, and protein concentration dependence.
This report provides a characterization of the effects of varying the concentrations of Mg2+, ATP, phosphorylase kinase, and the cAMP-dependent protein kinase on the activation and phosphorylation of phosphorylase kinase. The results show the following. (a) The Km for MgATP2- for the cAMP-dependent protein kinase-catalyzed phosphorylation is decreased by increasing Mg2+, probably as a consequence of decreasing the free ATP:MgATP2- ratio and increasing free Mg2+. (b) Whereas beta subunit phosphorylation of phosphorylase kinase plays a prominent role in determining its activity, alpha subunit phosphorylation can also modulate activity. (c) The phosphorylation of the alpha subunit, which occurs following the initial cAMP-dependent phosphorylation of the beta subunit, is catalyzed by the cAMP-dependent protein kinase and is not a consequence of EGTA-insensitive (or EGTA-sensitive) autophosphorylation occurring as a result of the enhanced phosphorylase kinase activity. (d) The relationship between subunit phosphorylation and phosphorylase kinase activation is complex and particularly dependent upon concentrations of cAMP-dependent protein kinase and phosphorylase kinase in the activation reaction. The data suggest the possibilities that the pathway of phospho-intermediates involved in the activation process probably varies with the activation conditions, that the efficacy of a specific site to be covalently modified is dependent upon the phosphorylation status of other sites, and that the effect of phosphorylation in regulating activity may also be dependent on the phosphorylation status of other sites. It is clear from the data that the activation process for phosphorylase kinase can be very complex, and it is possible that this complexity might have significant physiological ramifications.
Pickett-Gies
C A
CA
Walsh
D A
DA
D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D000242:Cyclic AMP / Q000494:pharmacology*; D004789:Enzyme Activation; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D009132:Muscles / Q000378:metabolism*; D010764:Phosphorylase Kinase / Q000378:metabolism*; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism*; D011817:Rabbits
D000255:Adenosine Triphosphate; D000242:Cyclic AMP; D011494:Protein Kinases; D010764:Phosphorylase Kinase; D008274:Magnesium