The pyrophosphatase activity of pig kidney alkaline phosphatase and its inhibition by magnesium ions and excess of pyrophosphate

Abstract

1. Pig kidney enzyme resembles other non-specific alkaline phosphatases in its ability to hydrolyse inorganic pyrophosphate (PPi). 2. Studies of enzyme velocity as a function of PPi concentration show that Michaelis–Menten kinetics are obeyed when a constant PPi/Mg2+ concentration ratio is maintained, but velocity–substrate concentration curves are sigmoid when the concentration of PPi is increased but that of Mg2+ is kept constant. The enzyme is inhibited when the total PPi concentration is greater than the total concentration of Mg2+. Pyrophosphatase activity is activated by Mg2+, but if the concentration of the metal ion is increased to a value in excess of the total PPi concentration Mg2+ is then strongly inhibitory. 4. It appears that the enzyme is most active towards the complex ion MgPPi2−. The enzyme probably hydrolyses PPi4− also, but this is a poorer substrate and its competition with MgPPi2− leads to inhibition. At high Mg2+ concentrations Mg2PPi is formed. This complex appears to be a potent inhibitor. 5. Sigmoid plots of v against s and of v against i result from interactions occurring between Mg2+ and PPi4− leading to MgPPi2− and Mg2PPi, and are not indicative of allosteric behaviour.