Immobilization of β-Galactosidase on an Insoluble Carrier with a Polyisocyanate Polymer. II. Kinetics and Stability

Abstract

The kinetic properties of commercial fl-galactosidase (Escherichia coli K-12) bonded to .8 cm x 1.3 cm Teflon stirring bars via a polyisocyanate polymer were compared to those of the soluble enzyme Assays of immobilized fl-galactosidase were performed with test tubes placed in a 37 C water-jacketed container positioned on top of a stirring motor set at 80 -4"_ 5 rpm. The Michaelis-Menten constants for soluble and immobilized flgalactosidase at pH 6.5 were 13.1 mM and 22.1 raM. The pH optimum for immobilized fl-galactosidase shifted upward .8 pH units relative to the soluble enzyme. Activation energies for hydrolysis of lactose were 15,200 cal/mole and 12,500 cal/mole for soluble and immobilized /3-galactosidase. The immobilized fl-galactosidase exhibited good stability, retaining 65.6% of the original activity after intermittent use for 118 days. When not in use, the immobilized fl-galaetosidase was stored at 4 C in .1 M phosphate buffer (pH 6.5).