title: Imidazole chloride and tris-chloride substitute for sodium chloride in inducing high-affinity AdoPP[NH]P binding to (Na+ + K+)-ATPase.
abstract: Optimal binding of [2,8-3H]AdoPP[NH]P to (Na+ + K+)-ATPase requires 25 mM Na+ (Cl-), 50 mM imidazole+ (Cl-) or 50 mM Tris+ (Cl-). Chloride is essential as counterion. We conclude that imidazole+ and Tris+ are able to bind to the Na+ site, and recommend the use of dilute buffers for studying the partial reactions of (Na+ + K+)-ATPase. In NaCl or the substituting buffers the dissociation constant for the enzyme-AdoPP[NH]P complex at 0 degrees C and pH 7.25 is 0.4 microM, whereas in millimolar MgCl2 it is about 2 microM. These distinct levels in affinity with MgCl2 as compared to NaCl, together with the MgCl2-dependence of photolabelling of the enzyme with ATP analogues (Rempeters, G. and Schoner, W. (1981) Eur. J. Biochem. 121, 131-137), suggest significant changes within the substrate site of (Na+ + K+)-ATPase upon binding of Mg2+ (Cl-)2.
mesh_terms: D000255:Adenosine Triphosphate / Q000031:analogs & derivatives*; D000266:Adenylyl Imidodiphosphate / Q000378:metabolism*; D000818:Animals; D007093:Imidazoles / Q000494:pharmacology*; D007668:Kidney / Q000201:enzymology; D007700:Kinetics; D009994:Osmolar Concentration; D011485:Protein Binding; D012965:Sodium Chloride / Q000494:pharmacology*; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism*; D013552:Swine; D014325:Tromethamine / Q000494:pharmacology*
chemical_list: D007093:Imidazoles; D014325:Tromethamine; D000266:Adenylyl Imidodiphosphate; D012965:Sodium Chloride; D000255:Adenosine Triphosphate; D000254:Sodium-Potassium-Exchanging ATPase