Enhancing azo dye degradation using dual enzyme systems immobilized on a mesoporous silica scaffold

Highly efficient degradation of the azo dye methyl red has been achieved through a coupled enzymatic reaction using mesoporous silica as an immobilization scaffold for two different enzymes. The dispersion of the non-immobilized free azoreductase (AzoR) and glucose dehydrogenase (GDH) in the reaction solution caused vigorous aggregation of these heterologous enzymes, resulting in their deactivation. In the present study, we aimed to overcome this limitation by fusing Si-tag, a silica-binding protein, to both enzymes and simultaneously immobilizing them on the surface of highly ordered pores of mesoporous silica. Consequently, the degradation ratio of methyl red significantly increased with immobilization on the mesoporous silica compared to that from immobilization on a non-porous silica support. However, the immobilization of the AzoR and GDH with and without the Si-tag, respectively, on silica markedly decreased the enzyme activity during the reusability test owing to the desorption of GDH involved in coenzyme regeneration. By contrast, the activities of the two enzymes immobilized on the mesoporous silica surface markedly increased upon fusion of both with Si-tag. Furthermore, these dual enzyme-mesoporous silica composites prepared at high salt and surfactant concentrations exhibited higher durability and repeatability during methyl red degradation than when using non-porous silica.