β-galactosidase immobilization onto poly (acrylonitrile- co-methyl methacrylate) nanoparticles

In this study, β-galactosidase (Aspergillus oryzae) was covalently immobilized onto aminated Poly (Acrylonitrile-Co- Methyl Methacrylate), P(AN-co-MMA), nanoparticles via glutaraldehyde coupling. Effect of the immobilization conditions was investigated. The maximum expressed activity was found 22.35 (μmole min-1g-1). Biochemical characterization of the immobilized enzyme shows that the optimum pH was shifted to 4.4, while the optimum of the free enzyme was observed at 5.2. The optimum temperature for the immobilized and free forms was recognized at 60%C and 50°C, respectively. A change of the temperature profile of the immobilized enzyme below 50°C was observed where response to elevation temperature was less than the free counterpart which is an indication of acquired thermal stability at higher temperatures. The kinetic studies show that the Km values were estimated at 54.5 and 117.3 mM for the free and the immobilized β-galactosidase, respectively. The apparent Km value of the immobilized β-galactosidase was two times higher than that of the free enzyme. On the hand, the Vmax value of the immobilized enzyme increased about 3.5- fold, compared to the free enzyme. The maximum immobilization capacity was found 22350 activity units per Kg and the maximum retained activity about 85%. For practical applications, the thermal, pH and storage stabilities were evaluated. Finally, the immobilized enzyme kept about 50% of its initial activity after 10 runs.