Inhibitory Effect of 0.19 α-Amylase Inhibitor from Wheat Kernel on the Activity of Porcine Pancreas α-Amylase and Its Thermal Stability

Abstract

The inhibitory effect of 0.19 α-amylase inhibitor (0.19 AI) from wheat kernel on the porcine pancreas α-amylase (PPA)-catalyzed hydrolysis of p-nitrophenyl-α-D-maltoside (pNP-G2) was examined. 0.19 AI is a homodimer of 26.6 kDa with 13.3-kDa subunits under the conditions used. The elution behaviors in gel filtration HPLC of PPA and 0.19 AI indicated that a PPA molecule bound with a 0.19 AI molecule (homodimer) at a molar ratio of 1:1. 0.19 AI inhibited PPA activity in a competitive manner with an inhibitor constant, K_i, of 57.3 nM at pH 6.9, 30°C, and the binding between them was found to be endothermic and entropy-driven. The activation energy for the thermal inactivation of 0.19 AI was determined to be 87.0 kJ/mol, and the temperature, T₅₀, giving 50% inactivation in a 30-min incubation at pH 6.9 was 88.1°C. The high inhibitory activity of 0.19 AI against PPA and its high thermal stability suggest its potential for use in the prevention and therapy of obesity and diabetes.