Kinetics and thermodynamics of catalysis by inorganic pyrophosphatase

Combined evidence obtained from measurements of pyrophosphate hydrolysis and synthesis, oxygen exchange between phosphate and water and enzyme-bound pyrophosphate formation was used to determine forward and reverse rate constants for the four reaction steps (binding/release of pyrophosphate, hydrolysis/synthesis of pyrophosphate and successive binding/release of two phosphates) catalyzed by yeast and Escherichia coli pyrophosphatase. Catalysis by the E. coli enzyme occurs via a single pathway, which requires the binding of Mg 2+ to four sites, while the yeast enzyme operates via two parallel pathways involving three or four Mg 2+ ions per one pyrophosphate molecule. The rate of synthesis is almost entirely determined by the rate of pyrophosphate release from the active site