Stimulation of phosphorylase kinase autophosphorylation by peptide analogs of phosphorylase.
Autoactivation of phosphorylase kinase in the presence of substrates has been studied to determine the cause of the hysteresis, or lag, in the phosphorylase kinase reaction. Peptide analogs corresponding to the convertible serine region of phosphorylase have been used as low molecular weight alternative substrates. Autophosphorylation of the kinase molecule was measured under conditions that favored autoactivation. Phosphorylase b and a tetradecapeptide, which was found to be a good model of phosphorylase, stimulated autoactivation by 86- and 37-fold, respectively. The tetradecapeptide also stimulated autophosphorylation of subunits A and B of the kinase molecule. This increased autophosphorylation coincided with an increased ability to convert phosphorylase. This finding supports the hypothesis that autophosphorylation is responsible for the lag in the phosphorylase kinase reaction. No evidence was obtained to suggest that the lag could be due to dissociation of the kinase. The stoichiometry of phosphate incorporation into phosphorylase kinase subunits by autophosphorylation was much greater than that reported to occur by protein kinase phosphorylation. Multiple phosphorylation sites in subunit A accounted for most of the phosphate incorporation during autophosphorylation. Saturating levels of hexa- and octapeptide analogs also caused stimulation of autophosphorylation. Possible mechanisms and experimental implications of substrate-stimulated autophosphorylation are discussed. Consideration also is given to the possible role of effectors in autophosphorylation in vivo.
D000818:Animals; D002118:Calcium / Q000494:pharmacology; D004789:Enzyme Activation; D007700:Kinetics; D009132:Muscles / Q000201:enzymology; D009842:Oligopeptides / Q000494:pharmacology; D010446:Peptide Fragments / Q000494:pharmacology; D010764:Phosphorylase Kinase / Q000302:isolation & purification / Q000378:metabolism*; D006005:Phosphorylases / Q000378:metabolism*; D011817:Rabbits; D013329:Structure-Activity Relationship
D009842:Oligopeptides; D010446:Peptide Fragments; D006005:Phosphorylases; D010764:Phosphorylase Kinase; D002118:Calcium