Kinetic studies of acid-inactivation of alpha-amylase from Aspergillus oryzae

The stability of alpha-amylase from Aspergillus oryzae has been studied at different pH. The enzyme is extremely stable at neutral pH (pH 5-8), whereas outside this pH-range a substantial loss of activity is observed. The acid-inactivation of alpha-amylase from A. oryzae was monitored on-line by a FIA-system. At low pH (pH<4.0) the inactivation follows first-order kinetics, and the rate constant is given by the empirical expression k = 1.19 x 10(7) [H+](1.99)(h(-1)) illustrating that the inactivation process is highly dependent of the pH in the medium. At pH 6 the acid-inactivated enzyme regains part of its activity, and the reactivation process also follows first-order kinetics. The irreversible loss of activity is found not to result from a protease contamination of the protein samples. A proposed model, where irreversibly inactivated a-amylase is formed both directly from the active enzyme and via the reversibly inactivated enzyme, describes the experimental data very well.