Kinetic modeling of lactose hydrolysis by a β-galactosidase from Kluyveromices fragilis

The kinetic model of lactose hydrolysis by means of a commercial beta-galactosidase from Kluyveromices fragilis provided by Novo nordisk has been determined using a wide range of the main variables: enzyme, substrate, and product concentrations and temperature. Lactose hydrolysis, which is of great interest due to physiological, nutritional, technological, and environmental reasons, has been performed in ct buffer whose salt composition is similar to that of milk. The effect of pH and temperature on enzyme activity and stability has been studied and it has been found that the optimal pH was 6.5. Temperatures over 45 degrees C cause a significant deactivation in few hours; thus a pH of 6.5 and a range of temperatures from 5-40 degrees C have been employed to accomplish the kinetic model discrimination. Five kinetic models described in the literature using different beta-galactosidases and reaction media have been considered. Substrate and product inhibition have been taken into account. Runs with different initial amounts of monosaccharides have been performed in order to discriminate among different kinetic models. Applying statistical and physical criteria, a Michaelis-Menten model with a competitive inhibition by galactose has been finally chosen, yielding a good fitting of the experimental data in the wide interval of variables studied. (C) 1998 Elsevier Science Inc.