Fusion of proteoliposomes and cells. ATP-dependent Ca2+ uptake into erythrocytes catalyzed by Ca2+-ATPase from skeletal muscle.
Purified Ca2+-ATPase from rabbit skeletal muscle has been incorporated into intact erythrocyte membranes by a two-step procedure. The isolated protein was reconstituted into proteoliposomes composed of phosphatidylethanolamine, phosphatidylcholine, and cardiolipin (50:20:30%, respectively). The resulting proteoliposomes were fused with erythrocytes in presence of La3+, Ca2+, or Mg2+. Subsequently, 45Ca uptake into the cells could be demonstrated. It was dependent on externally added ATP, inhibited by N-ethylmaleimide and p-hydroxymercuribenzoate, and enhanced by inactivation of the endogenous Ca2+-ATPase which catalyzes Ca2+ extrusion from the cells. The insertion of the protein did not induce cell lysis, but the cells did become more fragile. Functional insertion of isolated membrane proteins into cell membranes allows a new approach to research of plasma membranes.
D000818:Animals; D001693:Biological Transport, Active / Q000187:drug effects; D002118:Calcium / Q000097:blood*; D000252:Calcium-Transporting ATPases / Q000378:metabolism*; D004912:Erythrocytes / Q000378:metabolism*; D007700:Kinetics; D007811:Lanthanum / Q000494:pharmacology; D008081:Liposomes; D008274:Magnesium / Q000494:pharmacology; D009132:Muscles / Q000201:enzymology*; D010743:Phospholipids; D011485:Protein Binding; D011510:Proteolipids; D011817:Rabbits
D008081:Liposomes; D010743:Phospholipids; D011510:Proteolipids; C020485:proteoliposomes; D007811:Lanthanum; D000252:Calcium-Transporting ATPases; D008274:Magnesium; D002118:Calcium