Kinetic Studies on the Interactions of Two Forms of Inorganic Pyrophosphatase of Heart Mitochondria with Physiological Ligands

Abstract

A scheme of interactions of Mg²⁺ ions and their 1 : 1 complex with PP_isub (PP_iMg′) with two forms of inorganic pyrophosphatase isolated from beef heart mitochondria has been deduced from the analysis of enzyme kinetics at pH varying from 5.6 to 8.5. The scheme implies the existence of two catalytically important metal-binding sites on the enzyme. The two enzyme forms differ in maximal velocity and affinity for the metal activator. The pH dependence of kinetic parameters suggests that the active form of the substrate is MgP₂O²⁻⁷. Ca²⁺ ions strongly inhibit pyrophosphatase activity and the corresponding Hill coefficient is 1.5. Phosphate and ATP are weak inhibitors of pyrophosphatase of the competitive and noncompetitive type respectively. The results show that these forms of mitochondria1 pyrophosphatase are similar to pyrophosphatases isolated from other sources.