Effect of the pH value on the thermal stability of alkaline phosphatase

The thermal stability of bovine and chicken intestinal alkaline phosphatase (IAP) was studied at 60°C over a pH range from 6.7 to 9.0. It was shown that the peaks of stability (at pH 7.5) and activity (at pH 9.0) do not coincide. The pH dependence of the elementary rate constants of dissociative thermal inactivation of the IAP dimers was determined: k ₁ (the rate constant of dissociation of active dimer E₂) and k _d (the rate constant of denaturation of the inactive monomers). At pH 7.5, the stability IAP attains its highest level. As the pH increases, k ₁ increases drastically while k _d does so only slightly. A comparison of the rate constants of dissociation k ₁ and association k ₁ showed that the interrelation between these parameters explains why the stability of the active enzyme is lower at acidic and basic pH values. The pH produces a weaker effect on the stability of the inactive monomers. An analysis of the thermal stability of chicken IAP at an optimum pH value and 55–60°C showed that the thermal inactivation is a three-stage process (including dissociation and denaturation) with an induction period. Measuring the induction period makes it possible to determine the minimum number of latent stages preceding the dissociation of E₂: 6, 4, and 3 at 55, 58, and 60°C, respectively.