The Regulation of Skeletal Muscle Phosphorylase Kinase by Ca2+

Abstract

Abstract The interaction between highly purified rabbit muscle phosphorylase kinase and Ca2+ has been investigated and this metal has been shown to be necessary for enzyme activity. Utilizing calcium-free reagents, apparent Km values for Ca2+ were determined for the activated kinase (2 x 10-7 m at pH 8.2, 5 x 10-7 m at pH 6.8) and for the nonactivated kinase (3 x 10-6 m at pH 8.2, indeterminate at pH 6.8). Direct binding of Ca2+ to phosphorylase kinase was studied by the method of gel filtration equilibrium using 45Ca and an approximate minimal molar binding weight of 55,000 g was assigned. The dissociation constants of the various Ca2+-kinase complexes were in the same range as the above listed Km values. Sr2+ and Ba2+ were found to substitute for Ca2+ in the phosphorylase kinase reaction; the enzyme-bound Ca2+ was totally exchangeable with Sr2+. A freshly isolated fraction of sarcoplasmic reticulum from rabbit skeletal muscle completely inhibited nonactivated phosphorylase kinase at pH 7.6; this inhibition was reversed by Ca2+. The activated kinase was inhibited 80% under the same conditions. A model system is proposed for the regulation of skeletal muscle phosphorylase kinase by Ca2+ in vivo, and the consequent linking of the process of glycogenolysis to that of muscle contraction.