Metal displacement effects on monoesterase activity of calf intestinal alkaline phosphatase

Abstract

The mechanism of modulation of alkaline phosphatase activity by metal ions has not been fully elucidated. We investigated the time-dependent modulatory effects of Mg²⁺ and Zn²⁺ in promoting the hydrolysis of para-nitrophenyl phosphate (monoesterase reaction) by calf intestinal alkaline phosphatase (CIAP) and the effects of addition of the activating metal ions to the metal-inhibited enzyme. The CIAP was affected by changes in pre-incubation time in the presence of the two metal cofactors. Both Mg²⁺ (0.1 – 0.25 mM) and Zn²⁺ (0.1 – 5 mM) modulated Zn²⁺- and Mg²⁺- inhibited monoesterase activity of CIAP. The CIAP activity was inhibited when the enzyme was pre-incubated with 1 mM Ca²⁺. Further addition of Mg²⁺ (0.1 – 2 mM) did not completely restore the activity though partly relieved the inhibition caused by Ca²⁺- pre-incubated enzyme. Again, addition of 2mM Zn²⁺ to Ca²⁺- pre-incubated alkaline phosphatase completely restored the activity of the enzyme. This study suggests that Mg²⁺ and Zn²⁺ regulate the catalytic property of each other and modulate the inhibitory effect of Ca²⁺ in alkaline phosphatase catalysis through a displacement effect. The modulation of Ca²⁺-inhibited CIAP activity by Mg²⁺ and Zn²⁺ may be explored in the treatment of disorders of bone mineralization especially those arising from inhibited alkaline phosphatase activity.