Analysis of mechanism and kinetics of thermal inactivation of enzymes: Evaluation of multitemperature data applied to inactivation of yeast invertase

Abstract

A method is presented that substantially improves the analysis of the inactivation mechanism when compared to the conventional evaluation of isothermal data. The method utilizes a simultaneous fit of inactivation data obtained at several temperatures with the kinetic models containing the temperature dependence of rate constants in the form of the Arrhenius equation. In a case study of thermal inactivation of yeast invertase in the temperature range of 40–60°C, it was demonstrated that the multitemperature evaluation method significantly diminished the lumped character of inactivation kinetics. While the isothermal evaluation could provide only a formal two-step kinetic equation without verifying a particular mechanism, the presented method efficiently discriminated among different two- and three-reaction mechanisms. The general assessment of the method was made by comparing the modeling results in relation to the degree of complexity and structure of the examined mechanisms. Based on the modeling results, the inactivation of invertase was assumed to proceed in at least three steps which were discussed in relation to the available structural data on the mechanism of yeast invertase inactivation.