Relationship of major phosphorylation reactions and MgATPase activities to ATP-dependent shape change of human erythrocyte membranes.
Human erythrocyte ghosts prepared by hemolysis and washing in hypotonic Tris are crenated by salt and divalent cations, but undergo shape change to smooth biconcave discs and stomatocytic forms when incubated with MgATP at 37 degrees C. This is normally accompanied by protein and lipid phosphorylations in which the major phosphate acceptors are the spectrin beta-chain and inositol phospholipids, respectively. The system was manipulated in several ways to demonstrate the independence of ATP-dependent shape change from the major phosphorylation reactions. Salt-extracted membranes incubated with adenosine, an inhibitor of spectrin and phosphatidylinositol kinases, underwent normal shape change despite reductions of greater than 90% in spectrin and phospholipid labeling by [gamma-32P]ATP. ATP-dependent shape change was blocked by vanadate at micromolar concentrations (half-maximal inhibition at less than 1 microM), but vanadate did not inhibit membrane autophosphorylation reactions or turnover of spectrin- or lipid-bound phosphate. Vanadate inhibited part of the ATP hydrolysis that accompanies shape change and is expressed in the presence of ouabain and EGTA. The vanadate-sensitive MgATPase activity was approximately 3 nmol Pi X min-1 X mg of protein-1. The results implicate it in ATP-dependent shape change.
D000241:Adenosine / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000097:blood*; D017301:Ca(2+) Mg(2+)-ATPase / Q000097:blood*; D003035:Cobalt / Q000378:metabolism; D004910:Erythrocyte Membrane / Q000201:enzymology / Q000648:ultrastructure*; D006801:Humans; D006868:Hydrolysis; D007700:Kinetics; D050356:Lipid Metabolism; D008345:Manganese / Q000378:metabolism; D010716:Phosphatidylinositols / Q000378:metabolism; D010766:Phosphorylation; D013049:Spectrin / Q000378:metabolism; D014638:Vanadates; D014639:Vanadium / Q000494:pharmacology
D010716:Phosphatidylinositols; D014639:Vanadium; D013049:Spectrin; D003035:Cobalt; D014638:Vanadates; D008345:Manganese; D000255:Adenosine Triphosphate; D017301:Ca(2+) Mg(2+)-ATPase; D000241:Adenosine