The effects of macromolecular crowding and surface charge on the properties of an immobilized enzyme: activity, thermal stability, catalytic efficiency and reusability

Abstract

The microenvironment around an immobilized enzyme molecule significantly influences the properties of the immobilized enzyme. To explore the combined effects of macromolecular crowding and surface charge on the properties of an immobilized enzyme, epoxy resin was modified by different quality percentage concentrations of chitosan to obtain different crowding and charge conditions of the microenvironment. With this system, the catalytic features of immobilized 7α- and 7β-hydroxysteroid dehydrogenase (7α-, 7β-HSDH) were investigated. The results indicate that the activities and thermal stabilities of immobilized 7α-HSDH and 7β-HSDH can be improved by manipulating the crowding and surface charge effects. The relative activity of immobilized 7α-HSDH and 7β-HSDH on EP-0.5-C increased approximately 90% and 50% relative to the immobilization onto EP. The immobilization of 7α-HSDH and 7β-HSDH onto EP-0.5-C preserved a higher activity (approximately 73% and 88% of the initial activity) than that preserved by immobilization onto EP (approximately 29% and 40% of the initial activity) after incubation at 45 °C for 2 h. In addition, the catalytic efficiency and reusability of co-immobilized 7α-HSDH and 7β-HSDH was improved. The TCDCA conversion of the co-immobilized enzymes onto EP-0.5-C was increased approximately 45%, compared to the conversion of the co-immobilized enzymes onto EP. The TCDCA conversion of the co-immobilized enzymes on EP-0.5-C remained at 83.72 ± 0.66% after seven successive cycles. These findings suggest that crowding and charge of the microenvironment can significantly improve the properties of the immobilized enzyme.