ACTIVATION OF PROTEIN PHOSPHATASE 2A BY CAMP-DEPENDENT PROTEIN KINASE-CATALYZED PHOSPHORYLATION OF THE 74-KDA B'' (DELTA)-REGULATORY SUBUNIT IN-VITRO AND IDENTIFICATION OF THE PHOSPHORYLATION SITES

Human erythrocyte protein phosphatase 2A, which comprises a 34-kDa cat alytic C subunit, a 63-kDa regulatory A subunit and a 74-kDa regulator y B '' (delta) subunit, was phosphorylated at serine residues of B '' in vitro by cAMP-dependent protein kinase (A-kinase), In the presence and absence of 0.5 mu M okadaic acid (OA), A-kinase gave maximal incor poration of 1.7 and 1.0 mol of phosphate per mol of B '', respectively . The K-m value of A-kinase for CAB '' was 0.17 +/- 0.01 mu M in the p resence of OA, The major in vitro phosphorylation sites of B '' were i dentified as Ser-60, -75 and -573 in the presence of OA. and Ser-75 an d -573 in the absence of OA, Phosphorylation of B '' did not dissociat e B '' from CA, and stimulated the molecular activity of CAB '' toward phosphorylated H1 and H2B histones, 3.8- and 1.4-fold. respectively, but not toward phosphorylase a. (C) 1998 Federation of European Bioche mical Societies.