Using Sugar and Amino Acid Additives to Stabilize Enzymes within Sol−Gel Derived Silica

The inclusion of additives during the immobilization of proteins into sol−gel processed materials has been widely explored as a route to stabilize proteins against the denaturing stresses encountered upon entrapment. In this report, we explore the effects of sorbitol and N-methylglycine (collectively referred to as osmolytes) on both the conformational stability and biological activity of the enzymes α-chymotrypsin and ribonuclease T1 in solution and when entrapped into sol−gel derived silica. In each case, the encapsulation of the enzymes into sol−gel derived silica in the absence of additives led to a moderate decrease in the thermodynamic stability of the proteins. However, entrapment in the presence of the osmolytes produced significant increases in the thermal stability and biological activity of the encapsulated proteins. We show that the observed enhancements in enzyme stability are likely based on a combination of increases in the pore size of the silica material (which improves substrate delivery and thus activity) and changes in the thermal stability of the entrapped enzymes in the presence of osmolytes. Our results suggest that these additives stabilize the two proteins by altering the hydration of the entrapped protein, hence this stabilization method may prove to be applicable to a wide variety of proteins.