The Mg2+-dependent ATPase from the erythrocyte plasma membrane of the flounder Platichthys flesus L. General properties and some observations on the steady state kinetics.

1. The plasma membrane of the flounder erythrocyte contains a Mg2+-dependent ATPase which is insensitive to ouabain. Mg2+ is part of the substrate, Mg-ATP, and Mg2+ also functions as a nonessential activator. 2. Ca2+, Mn2+ and Co2+ can replace Mg2+ as an activator of ATP hydrolysis. Cu2+ and Zn2+ abolish the Mg-dependent activity. It is shown that Ca2+ and Mg2+ activate the same enzyme and that Mg-ATP and Ca-ATP are mutually competitive. 3. The hydrolysis of ATP obeys Michaelis-Menten kinetics whether or not the Mg2+-ATPase is fully activated by Mg2+. The KM values for Mg-ATP were found to be 0.13 and 0.43 mM respectively. 4. Free ATP acts as a competitive inhibitor towards Mg-ATP and the dissociation constant for the enzyme-ATP complex was determined to be about 0.55 mM. 5. The Mg2+ -ATPase has a low specificity and reacts with the common nucleoside triphosphates GTP, ITP, UTP and CTP. 6. The enzyme has a broad pH optimum ranging from 6.5 to 7.2 and an energy of activation of 13.5 kcal/mol between 0 and 30 degrees C. 7. The effect of some activators and inhibitors of membrane-bound ATPases are reported.