Protein kinase activity and endogenous protein phosphorylation in rat liver plasma membranes

When rat liver plasma membranes were incubated with [gamma-32P]ATP radio-labelled 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. Cyclic AMP did not affect the incorporation of phosphate. The protein kinase activity was extracted from the membranes by 1% Triton X-100 and a high concentration of KCl. The solubilised enzyme resolved into two fractions on DEAE-cellulose chromatography. One enzyme fraction had the same properties as the catalytic subunit of cytosolic cyclic AMP-dependent protein kinases. Endogenously phosphorylated proteins were resolved by SDS-polyacrylamide gel electrophoresis into five 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% Triton X-100 and 1M KCl.