State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University , Nanjing 210093, China.
Anal Chem. 2017 Dec 5;89(23):12924-12929. doi: 10.1021/acs.analchem.7b03780. Epub 2017 Nov 15.
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.
基于酶反应的电化学生物传感器已经应用于广泛的领域。随着这一趋势的持续发展，这些生物传感器已经接近物理和化学所施加的限制。为了进一步提高生物传感器的性能，在生物传感器的设计中应考虑质量传输和酶反应动力学之间的相互作用，特别是在酶级联系统中。在此，我们提出了一种简单的方法来研究质量传输和酶分子运动对酶级联反应动力学的影响。β-半乳糖苷酶（β-Gal）和葡萄糖氧化酶（GOx）的酶级联反应通过共价附着分别精确地固定在旋转环盘电极（RRDE）的盘和环电极上。在低转速（<600 rpm）下，对流传输促进了酶级联反应。当转速高于 600 rpm 时，级联反应变为动力学控制。进一步提高转速会导致反应速率缓慢下降，这可能是由于产物抑制效应。此外，还应考虑在较高离心力下酶构象变化对酶活性的影响。本研究将阐明对流力对酶级联反应动力学调节的影响，为研究其他酶级联反应提供理想的平台，并为设计高性能生物传感器、生物燃料电池和生物电子学提供基础。
