Illustrating the Mass-Transport Effect on Enzyme Cascade Reaction Kinetics by Use of a Rotating Ring-Disk Electrode.

Abstract

Electrochemical biosensors based on enzymatic reaction have been applied to a wide range of fields. As the trend continues to grow, these biosensors are approaching the limit imposed by physics and chemistry. To further improve the performance of biosensors, the interplay of mass transport and enzymatic reaction kinetics, especially in enzyme cascade systems, should be considered in the design of biosensors. Herein, we propose a simple approach to studying the influence of mass transport and enzyme molecule motion on the kinetics of enzyme cascade reactions. β-Galactosidase (β-Gal) and glucose oxidase (GOx) of the enzyme cascade reaction are precisely immobilized onto the disk and ring electrodes, respectively, of a rotating ring-disk electrode (RRDE) via covalent attachment. At a low rotating speed (<600 rpm), convective transport promotes the enzyme cascade reaction. When the rotating speed is higher than 600 rpm, the cascade reaction becomes kinetically controlled. Further increase of the rotating speed results in a slow decline in reaction rate, possibly due to the production inhibition effect. In addition, the effect of conformation change of the enzyme at higher centrifugal forces on enzyme activity should be considered. This study would shine light on the effect of convective force on regulation of kinetics of enzyme cascade reaction, offering an ideal platform for studying other enzyme cascade reactions and providing fundamentals to design high-performance biosensors, biofuel cells, and bioelectronics.