Demonstration of a phosphopeptide intermediate in the Mg ++ -dependent, Na + - and K + -stimulated adenosine triphosphatase reaction of the erythrocyte membrane.
Human erythrocyte membranes are phosphorylated by [gamma-(32)P]ATP in association with the Mg(++)-dependent, Na(+) and K(+)-stimulated ATPase (EC 3.1.6.3) reaction. To delineate the membrane species involved, phosphorylated membranes were analyzed by polyacrylamide gel electrophoresis in sodium dodecyl sulfate, under conditions that minimize hydrolysis of acyl phosphate linkages. Three radioactive components were detected, of which only one was a phosphopeptide, of apparent molecular weight 105,000. The phosphate bound to this peptide undergoes rapid turnover and is discharged by hydroxylamine. In the presence of Mg(++), the phosphorylation of this peptide is specifically stimulated by Na(+) and blocked by ethylene diamine tetraacetate; its dephosphorylation is stimulated by K(+) and blocked by ouabain. We conclude that this phosphopeptide is an intermediate in the Mg(++)-dependent, Na(+)- and K(+)-stimulated ATPase reaction of the erythrocyte membrane.
D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000494:pharmacology; D001693:Biological Transport, Active / Q000187:drug effects; D001797:Blood Protein Electrophoresis; D002462:Cell Membrane / Q000032:analysis / Q000187:drug effects / Q000201:enzymology*; D004492:Edetic Acid / Q000494:pharmacology; D004912:Erythrocytes / Q000201:enzymology*; D006801:Humans; D006898:Hydroxylamines / Q000494:pharmacology; D008274:Magnesium / Q000494:pharmacology*; D010042:Ouabain / Q000494:pharmacology; D010085:Oxidative Phosphorylation / Q000187:drug effects; D010748:Phosphopeptides / Q000302:isolation & purification / Q000378:metabolism*; D010759:Phosphorus Isotopes; D011188:Potassium / Q000494:pharmacology*; D011485:Protein Binding / Q000187:drug effects; D012964:Sodium / Q000378:metabolism / Q000494:pharmacology*
D006898:Hydroxylamines; D010748:Phosphopeptides; D010759:Phosphorus Isotopes; D010042:Ouabain; D000255:Adenosine Triphosphate; D004492:Edetic Acid; D012964:Sodium; D000251:Adenosine Triphosphatases; D008274:Magnesium; D011188:Potassium