B-galactosidase of kluyveromyces lactis: Immobilization, characterization and hydrolysis behavior of enzyme

Abstract

B-galactosidase (lactase) enzyme is of great industrial interest, since it can be used to solve problems associated with whey disposal and lactose crystallization in sweetened and frozen dairy products such as ice cream. All over the world, many people suffer from lactose intolerance and lactase preparations are used as supplements for these persons. B-galactosidase is also used to produce prebiotics since this enzyme hydrolyses lactose into galactooligosaccharides. Immobilized B-galactosidase preparations are preferred in many processes because they can be recycled and maintain their activities for a long time without loosing their chemical stability.Novel cross-linked chitosan-hydroxyapatite composite support has beenprepared, lactase from Kluyveromyces lactis was immobilized onto these beads. Lactase immobilization mechanism and effect of factors such as initial glutaraldehyde concentration, temperature, pH, initial lactase concentration, solid-liquid ratio on immobilzation mechanism were studied.Optimum cross-linking was obtained at the glutaraldehyde concentration of 100 mg/L. The optimum values of temperature, pH and solid/liquid ratio on lactase/HAChitosan were found to be 200C, pH 7.5 and 0.3g/ml Vg/Vl, respectively. The pH and thermal stabilities of free and immobilized enzymes were also investigated and it was observed that the relative activity remained above 83.2% within pH 6-7.5 and maximum activity yield was obtained at 370C for free and all immobilized enzymes. The Michaelis constant Km and Vmax of immobilized and free enzyme on chitosanhydroxyapatite composite beads were found to be 9.5 mM and Vm 454.5 .mol ONP min.1 mg.1 protein and 1.011 mM and 1098.9 .mol ONP min.1 mg.1 protein, respectively.