Crystallization of Ca2+-ATPase in detergent-solubilized sarcoplasmic reticulum.
Microcrystalline arrays of Ca2+-transporting ATPase (EC 3.6.1.38) develop in detergent-solubilized sarcoplasmic reticulum upon exposure to 10-20 mM CaCl2 at pH 6.0 for several weeks at 2 degrees C, in a crystallization medium that preserves the ATPase activity for several months. Of 48 detergents tested, optimal crystallization was obtained with Brij 36T, Brij 56, and Brij 96 at a detergent:protein weight ratio of 4:1 and with octaethylene glycol dodecyl ether at a ratio of 2:1. Similar Ca2+-induced crystalline arrays were obtained with the purified or delipidated Ca2+-ATPase of sarcoplasmic reticulum but at lower detergent:protein ratios. The crystals are stabilized by fixation with glutaraldehyde and persist even after the removal of phospholipids by treatment with phospholipases A or C and by extraction with organic solvents. The crystals obtained so far can be used only for electron microscopy, but ongoing experiments suggest that under similar conditions large ordered arrays may develop that are suitable for x-ray diffraction analysis.
D000252:Calcium-Transporting ATPases / Q000302:isolation & purification* / Q000378:metabolism; D003460:Crystallization; D003840:Deoxycholic Acid; D003902:Detergents; D005614:Freeze Fracturing; D007700:Kinetics; D008854:Microscopy, Electron; D012519:Sarcoplasmic Reticulum / Q000201:enzymology* / Q000648:ultrastructure; D012995:Solubility; D014961:X-Ray Diffraction
D003902:Detergents; D003840:Deoxycholic Acid; D000252:Calcium-Transporting ATPases