Phospholipid-sensitive Ca2+-dependent protein kinase from heart. II. Substrate specificity and inhibition by various agents.

Abstract

The specificity of substrates (including both phosphate donors and acceptors) for phospholipid-sensitive Ca2'-dependent protein kinase purified (80-95% homogeneous) from bovine heart and inhibition of its activity by various agents were investigated.The apparent K,,, for ATP, using histone H1 as substrate, of the enzyme was 4.4 p ~. Phosphorylation by ATP was inhibited most markedly by a,P-methylene ATP and, to a lesser extent, by adenosine 5'-0-(thiotriphosphate), 2'-deoxy ATP, and P,y-methylene ATP.The enzyme was able to utilize adenosine 5'-0-(thiotriphosphate) to thiophosphorylate histone H1.Among histone subfractions, peptides, and proteins examined, histone H1 (apparent K , = 0.6 PM; v , , = 0.83 pmol/min/mg of enzyme) and myelin basic protein (apparent K , = 0.3 PM;V , , = 0.83 pmol/min/mg of enzyme) were the best substrates for the enzyme.The enzyme partially purified from rat brain and pig spleen also effectively phosphorylated myelin basic protein, with apparent K , val-