Negative or positive cooperation in calcium binding to detergent-solubilized ATPase of the sarcoplasmic reticulum. Its modulation by a high concentration of ATP.
Two different conformations of chemically equivalent Ca(2+)-ATPase molecules in the sarcoplasmic reticulum have been shown to non- and positive cooperatively bind two calcium ions, respectively (Nakamura, J. (1994) J. Biol. Chem. 269, 30822-30827). At pH 7.40, these ATPase molecules split into E1 (high affinity state for calcium), and E2 (low affinity state for calcium), respectively, before calcium binding. At this pH, calcium binding to the monomeric ATPase, solubilized with dodecyloctaethylenglycol monoether, was studied by examining 45Ca2+ binding to the ATPase and calcium dependence of its phosphorylation, fluorescence intensity, ATP-hydrolysis at a low (5 microM) concentration of ATP, and acetyl phosphate hydrolysis. The results suggest that the solubilized ATPase molecules predominantly preexist in E2 and negative cooperatively (the Hill value (nH) = 0.5-0.6) bind 2 mol of calcium/mol of the ATPase with an apparent calcium affinity (K0.5) of 3-5 microM. The nonequivalences of calcium bindings at the membranous ATPase molecules seem to result from the intermolecular interaction of the molecules. A high concentration (5 mM) of ATP modulated the binding manner so that it became positively cooperative (nH approximately 2) and increased the K0.5 to 0.1 microM.
Nakamura
J
J
Biological Institute, Faculty of Science, Tohoku University, Miyagi, Japan.
Tajima
G
G
D000255:Adenosine Triphosphate / Q000378:metabolism / Q000494:pharmacology*; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D000252:Calcium-Transporting ATPases / Q000737:chemistry / Q000378:metabolism*; D003902:Detergents / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D010766:Phosphorylation; D011487:Protein Conformation; D011817:Rabbits; D012519:Sarcoplasmic Reticulum / Q000201:enzymology*
D003902:Detergents; D000255:Adenosine Triphosphate; D000252:Calcium-Transporting ATPases; D002118:Calcium