Control of Phosphorylase Activity in a Muscle Glycogen Particle

This study describes a new type of inhibition of phosphorylase phosphatase that strongly suggests that the activity of this enzyme is modulated during regulation of the phosphorylase system. Phosphorylase phosphatase included in a muscle protein-glycogen complex along with phosphorylase, phosphorylase kinase, and other enzymes of glycogen metabolism undergoes a reversible inhibition when phosphorylase is activated. This inhibition requires free Ca2f in addition to Mg-ATP, i.e. the same conditions that trigger phosphorylase activation; it is observed only in the intact protein-glycogen complex. It is not caused by AMP or IMP generated from the rapid breakdown of ATP, two nucleotides known to inhibit the phosphorylase phosphatase reaction by binding to the substrate phosphorylase u. (a) Addition of AMP or IMP to the particulate system is without effect; (b) phosphatase activity returns to normal at the end of an activation cycle of phosphorylase when concentration of IMP is maximum; and (c) the same inhibition pattern is observed when a phosphopeptide derived from phosphorylase a is used as substrate, a reaction not affected by AMP or IMP. Digestion of glycogen in the complex by a-amylase abolishes the Cazt-dependent inhibition of phosphorylase phosphatase, but the activity of this enzyme is still predominantly unaffected by nucleotides. On the other hand, dissociation of the protein-glycogen complex by dilution brings about both a loss of the Ca2f-dependent inhibition of the phosphatase and the reappearance of its sensitivity to AMP or IMP. Phosphorylase phosphatase included in the complex acts on phosphorylase (I endogenously produced by addition of Ca2f, Mg2f, an d ATP 16-fold faster than on soluble phosphorylase Q added to the system. Nevertheless, the phosphatase reaction on the exogenous substrate still remains unaffected by high concentrations of AMP or IMP. No evidence was obtained that during inhibition the enzyme was covalently