Protein Kinase Activity and Endogenous Protein Phosphorylation in Rat Liver Plasma Membranes.

Abstract

When rat liver plasma membranes were incubated with [.gamma.-32P]ATP radio-labeled phosphate was incorporated into endogenous protein and exogenous substrate by a membrane-bound protein kinase activity. A high ATP/membrane protein ratio was required for optimum incorporation conditions. cAMP did not affect the incorporation of phosphate. The protein kinase activity was extracted from the membranes by 1% Triton X-100 and a high concentrations of KCl. The solubilized enzyme resolved into 2 fractions on DEAE-cellulose chromatography. One enzyme fraction had the same properties as the catalytic subunit of cytosolic cAMP-dependent protein kinases. Endogenously phosphorylated proteins were resolved by SDS[sodium dodecyl sulfate]-polyacrylamide gel electrophoresis into 5 major and additional minor phosphorylated components. Three of the major phosphorylated components were tightly bound to the membrane material and were not extracted by 1% Tritin X-100 and 1M KCl.