Novel Grafted Agar Disks for the Covalent Immobilization of β-D-Galactosidase

Novel grafted agar disks were prepared for the covalent immobilization of beta-D-galactosidase (beta-gal). The agar disks were activated through reacting with ethylenediamine or different molecular weights of Polyethyleneimine (PEI), followed by glutaraldehyde (GA). The modification of the agar gel and the binding of the enzyme were verified by Fourier Transform Infrared (FTIR) and elemental analysis. Moreover, the agar's activation process was optimized, and the amount of immobilized enzyme increased 3.44 folds, from 38.1 to 131.2 U/g gel, during the course of the optimization process. The immobilization of beta-gal onto the activated agar disks caused its optimum temperature to increase from 45 degrees C to 45-55 degrees C. The optimum pH of the enzyme was also shifted towards the acidic side (3.6-4.6) after its immobilization. Additionally, the Michaelis-Menten constant (K-m) increased for the immobilized beta-gal as compared to its free counterpart whereas the maximum reaction rate (V-max) decreased. The immobilized enzyme was also shown to retain 92.99% of its initial activity after being used for 15 consecutive times. (C) 2015 Wiley Periodicals, Inc.