Inactivation of detergent-solubilized sarcoplasmic reticulum ATPase.
Inactivation of sarcoplasmic ATPase in the solubilized state was studied in the absence and presence of Ca2+, Mg2+ and glycerol. The effects of the detergents octa(ethyleneglycol) mono-n-dodecyl ether (C12E8), 1-O-tetradecylpropanediol-(1,3)-3-phosphorylcholine and myristoylglycerophosphocholine were compared. All three detergents caused a rapid decline of the dinitrophenyl phosphatase activity of the unprotected enzyme. The stabilizing effect of Ca2+ ions was kinetically analysed. It was found that the stability of the solubilized enzyme depends on the Ca2+ concentration in a manner which is best explained by assuming rapid inactivation of Ca2+-free enzyme accompanied by slow inactivation of a calcium-enzyme complex (E1Ca). The apparent affinity constants obtained are in the order of 10(6)M-1, suggesting that high-affinity Ca2+ binding must be involved. No indications of a contribution were found, either of low-affinity Ca2+-binding sites of the conformational state E2 or of the high-affinity calcium complex E1Ca2. If Ca2+ was replaced by Mg2+, which exerts a weaker protection, the apparent affinity constants for Mg2+ are in the range of 1 mM-1. The stoichiometry of the effect of Mg2+ depends on the detergent.
Swoboda
G
G
Max-Planck-Institut für Medizinische Forschung, Heidelberg, Federal Republic of Germany.
Hasselbach
W
W
D009597:4-Nitrophenylphosphatase / Q000032:analysis; D000251:Adenosine Triphosphatases / Q000037:antagonists & inhibitors*; D000818:Animals; D002118:Calcium / Q000494:pharmacology; D003902:Detergents / Q000494:pharmacology*; D005990:Glycerol / Q000494:pharmacology; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D011817:Rabbits; D012519:Sarcoplasmic Reticulum / Q000201:enzymology*; D012995:Solubility; D013501:Surface-Active Agents / Q000494:pharmacology*
D003902:Detergents; D013501:Surface-Active Agents; D009597:4-Nitrophenylphosphatase; D000251:Adenosine Triphosphatases; D008274:Magnesium; D005990:Glycerol; D002118:Calcium