Effects of Crowding and Confinement of Enzyme in Mesoporous Silicates on Thermal Stability

Abstract

Since the enzyme encapsulation in mesoporous silica could mimic the confined and crowded protein environments in a living cell, the immobilized enzymes in mesoporous silicates were investigated as a possible experimental system for estimating the effects of macromolecular crowding in a confined space on protein thermal stability. Glucose oxidase (GOX) was encapsulated in the both mesoporous silicates at the first, MSU-F and SBA-15, as a model enzyme. The pore diameters of both MSU-F (30 nm) and SBA-15 (8 nm) are large enough to encapsulate GOX. SBA-15 is channel type structure while MSU-F has the structure of cellular form as well. Therefore, GOX molecules (5.2 nm × 6.0 nm × 7.7 nm) can be confined in channel type pore structure of SBA-15 as well as crowded in those inside the mesopore structures. The amount of enzyme loading on the silica supports was controlled by varying initial concentrations of enzyme solution at the adsorption step. During pre-determined time of thermal incubation, the silicate entrapping more GOX molecules retained higher activity than that of less enzyme molecules. As the concentrations of free GOX solutions were increased, thermal stability of enzyme inside the pore was also enhanced presumably in virtue of the crowding effect as well as confinement effect. This result suggests that more confined and crowded environment can enhance the thermal stability of enzymes.