Selective phosphorylation of erythrocyte membrane proteins by the solubilized membrane protein kinases

Abstract

The substrate and phosphoryl donor specificities of solubilized erythrocyte [rabbit, human] membrane cyclic [c] independent protein kinases toward human and rabbit erythrocyte membrane proteins were described. Three types of substrate preparations were utilized: heat-inactivated ghosts, isolated spectrin and 2,3-dimethylmaleic anhydride (DMMA)-extracted membranes. A 30,000-dalton protein kinase, extracted from either human or rabbit erythrocyte membranes, catalyze the phosphorylation of heat-inactivated membranes in the presence of ATP. The resulting phosphorylation profile was analogous to that of the autophosphorylation of membranes with ATP (in the absence of cAMP). These kinases also phosphorylated band 2 of isolated spectrin and band 3, but not glycophorin, in the DMMA-extracted ghosts. The ability of the 30,000-dalton kinases to use GTP as a phosphoryl donor appears to be related to the substrate or some other membrane factor. A 2nd kinase, which was 100,000 daltons and derived from rabbit erythrocyte membranes, used ATP or GTP to phosphorylate membrane proteins 2, 2.1, 2.9-3 in heat-inactivated ghosts, band 2 in isolated spectrin, glycophorin and to a lesser extent, band 3 in the DMMA-extracted ghosts.