Nanoparticle-supported multi-enzyme biocatalysis with in situ cofactor regeneration

Although there have been a long history of studying and using immobilized enzymes, little has been reported regarding the nature of immobilized cofactors. Herein we report that cofactor NAD(H) covalently attached to silica nanoparticles successfully coordinated with particle-immobilized enzymes and enabled multistep biotransformations. Specifically, silica nanoparticle-attached glutamate dehydrogenase (GLDH), lactate dehydrogenase (LDH) and NAD(H) were prepared and applied to catalyze the coupled reactions for production of α-ketoglutarate and lactate with the cofactor regenerated within the reaction cycle. It appeared that particle–particle collision driven by Brownian motion of the nanoparticles provided effective interactions among the catalytic components, and thus realized a dynamic shuttling of the particle-supported cofactor between the two enzymes to keep the reaction cycles continuing. Total turnover numbers (TTNs) as high as 20,000 h−1 were observed for the cofactor. It appeared to us that the use of particle-attached cofactor promises a new biochemical processing strategy for cofactor-dependent biotransformations.