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

This report describes the substrate and phosphoryl donor specificities of solubilized erythrocyte membrane cyclic adenosine 3',5'-monophosphate (cAMP)-independent protein kinases toward human and rabbit erythrocyte membrane proteins. Three types of substrate preparations have been 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, catalyzes the phosphorylation of heat-inactivated membranes in the presence of ATP. The resulting phosphorylation profile is analogous to that of the autophosphorylation of membranes with ATP (in the absence of cAMP). These kinases also phosphorylate 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 second kinase, which is 100 000 daltons and derived from rabbit erythrocyte membranes, uses 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.