Production and immobilization of β-galactosidase isolated from Enterobacter aerogenes KCTC2190 by entrapment method...

In the present study, Enterobacter aerogenes KCTC2190 was isolated from soil around a cattle shed area, which was capable of producing intracellular β-galactosidase. Partially purified β-galactosidase was immobilized by entrapment method in agar-agar gel matrix. Agar-agar entrapped beads were prepared by dropping the enzyme-agar solution to ice-cooled toluene-chloroform ((3:1 (v/v)). 45.88±0.11% activity of partially purified β-galactosidase was retained after immobilization (bead shape). Maximum immobilization yield was observed in the presence of 2.5% agar-agar concentration. After immobilization, optimum temperature required for the enzyme-substrate reaction was shifted from 50 to 60 °C and the optimum reaction time was shifted from 15 to 25 min. The optimum pH for both free and immobilized β-galactosidase was pH 7. Free enzyme showed lower activation energy in comparison with the immobilized one. For free as well as immobilized β-galactosidase thermal deactivation, rate constant (k_d) increased with increasing temperature while the values of decimal reduction time (D-values) and half-lives (t_1/2) decreased. Immobilization process increased the t_1/2 and D-values of β-galactosidase while it decreased the k_d. Thermostability of immobilized β-galactosidase was higher as they showed higher enthalpy (ΔΗ⁰) and Gibb's free energy (ΔG⁰)value than those of the free β-galactosidase. The negative entropy (ΔS⁰) of free and immobilized β-galactosidase established that both were in a more ordered state within the temperature range (50 to 70 °C) studied. Immobilized β-galactosidase was able to retain 51.65±1.61% of its initial activity after 7 batches of enzyme-substrate reaction. Immobilized β-galactosidase showed 78.09±3.69% of its initial activity even after 40 days of storage at 4 °C.