Long-term stabilization and crystallization of (Ca2+ + Mg2+)-ATPase of detergent-solubilized erythrocyte plasma membrane

Abstract Conditions which were optimal for the stabilization of Ca 2+ -transporting ATPase in solubilized sarcoplasmic reticulum membranes (Pikula, S., Mullner, N., Dux, L. and Martonosi, A. (1988) J. Biol. Chem. 263, 5277–5286) were also found conductive for preservation of (Ca 2+ + Mg 2+ )-ATPase activity in detergent-solubilized erythrocyte plasma membrane for up to 60 days. Of particular importance for the stabilization of calmodulin-stimulated Ca 2+ -dependent activity of (Ca 2+ + Mg 2+ )-ATPase of solubilized erythrocyte plasma membrane was the presence of Ca 2+ (10–20 mM), glycerol, anti-oxidants, proteinase inhibitors and appropriate detergents. Among eight detergents tested octaethylene glycol dodecyl ether, polyoxyethylene glycol(10) lauryl alcohol and polydocanol were found to be promotive in long-term preservation of the enzyme activity. Under these conditions (Ca 2+ + Mg 2+ )-ATPase of erythrocyte ghosts became highly stable and developed microcrystalline arrays after storage for 35 days. Electron micrographs of the negatively stained and thin sectioned material indicated that crystals of purified, detergent-solubilized, lipid-stabilized erythrocyte (Ca 2+ + Mg 2+ )-ATPase differ from those of Ca 2+ -ATPase of detergent-solubilized sarcopiasmic reticulum microsomes.