Phosphorylation of Adenohypophyseal Plasma Membranes and Properties of Associated Protein Kinase

Plasma membranes isolated from bovine anterior pituitary gland are self-phosphorylated in the presence of [γ-32P]ATP by an endogenous protein kinase. Eighty per cent of protein kinase activity is solubilized by treatment of the plasma membranes with 1.0 n NH4Cl or 0.5% Triton X-100. The presence of latent protein kinase activity is shown by an approximately 50% increase of total enzymatic activity upon addition of the detergent. The enzyme associated with the plasma membrane is Mg2+-dependent and its activity is inhibited by Ca2+ at all concentrations studied. [32P]Phosphate is incorporated into phosphoserine and phosphothreonine. Only 13% of the incorporated 32P can be removed by washing the membranes in a high salt solution (0.5 m KCl). As resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis at pH 7.1, adenohypophyseal plasma membranes show a consistent pattern of 36 bands, of which 11 act as substrate for protein kinase. Cyclic adenosine 3' : 5'-monophosphate leads to a 40 to 100% increased incorporation of 32P into nine bands. Changes of the level of phosphorylation of components of the plasma membranes and of the membranes of the secretory granules could lead to altered rates of membrane fusion-fission processes accompanying exocytosis.