A glucose biosensor based on glucose oxidase fused to a carbohydrate binding module family 2 tag that specifically binds to the cellulose-modified electrode

Weili Gong; Qingye Han; Yanru Chen; Binglian Wang; Jianguo Shi; Lushan Wang; Lei Cai; Qingjun Meng; Zhenyu Zhang; Qingai Liu; Yan Yang; Junhui Yang; Lan Zheng; Yiwei Li; Yaohong Ma

doi:10.1016/j.enzmictec.2021.109869

A glucose biosensor based on glucose oxidase fused to a carbohydrate binding module family 2 tag that specifically binds to the cellulose-modified electrode

Enzyme Microb Technol. 2021 Oct:150:109869. doi: 10.1016/j.enzmictec.2021.109869. Epub 2021 Jul 12.

Authors

Weili Gong¹, Qingye Han¹, Yanru Chen¹, Binglian Wang¹, Jianguo Shi¹, Lushan Wang², Lei Cai¹, Qingjun Meng¹, Zhenyu Zhang¹, Qingai Liu¹, Yan Yang¹, Junhui Yang¹, Lan Zheng¹, Yiwei Li¹, Yaohong Ma³

Affiliations

¹ Shandong Provincial Key Laboratory of Biosensors, Biology Institute, Qilu University of Technology (Shandong Academy of Sciences), No. 28789, Jingshi East Road, Licheng District, Jinan, Shandong, 250103, China.
² State Key Laboratory of Microbial Technology, Institute of Microbial Technology, Shandong University, Qingdao, Shandong, 266237, China.
³ Shandong Provincial Key Laboratory of Biosensors, Biology Institute, Qilu University of Technology (Shandong Academy of Sciences), No. 28789, Jingshi East Road, Licheng District, Jinan, Shandong, 250103, China. Electronic address: mayaohong@126.com.

PMID: 34489028
DOI: 10.1016/j.enzmictec.2021.109869

Abstract

The method of immobilization of glucose oxidase (GOD) on electrodes is especially important for the fabrication and performance of glucose biosensors. In this study, a carbohydrate binding module family 2 (CBM2) was successfully fused to the C terminal of GOD with a natural linker (NL) in endo-β-xylanase by genetic recombination, and a fusion GOD (GOD-NL-CBM2) was obtained. The CBM2 was used as an affinity adsorption tag for immobilization of the GOD-NL-CBM2 on a cellulose modified electrode. The specific activity of GOD-NL-CBM2 was comparable to that of the wild type GOD. In addition, the CBM2 tag of fusion GOD almost maintained its highest binding capacity under optimal catalytic conditions (pH 5.0, 50 °C). The morphology and composition analysis of the cellulose film reacted with and without GOD or GOD-NL-CBM2 confirmed the immobilization of GOD-NL-CBM2. The electrochemical properties of the GOD-NL-CBM2/cellulose film bioelectrode, with a characteristic peak of H₂O₂ at +0.6 V in the presence of glucose, revealed the capability of the immobilized GOD-NL-CBM2 to efficiently catalyze glucose and produce H₂O₂. Additionally, the current signal response of the biosensor to glucose was linear in the concentration range from 1.25 to 40 mM (r² ≥ 0.99). The sensitivity and detection limit of the GOD-NL-CBM2/cellulose film bioelectrode were 466.7 μA mol^-1 L cm^-2 and 0.475 mM (S/N = 3), respectively. Moreover, the glucose biosensor exhibited a rapid current change (< 5 s), high reproducibility (Relative standard deviation, RSD < 5%), substrate selectivity and stability, and retained about 80 % of the original current response after 2 months. The affinity adsorption-based immobilization strategy for GOD provides a promising approach to develop a high performance glucose biosensor.

Keywords: Affinity adsorption; Biosensor; Carbohydrate binding module family 2 tag; Cellulose film; Glucose oxidase.

MeSH terms

Biosensing Techniques*
Cellulose
Electrodes
Enzymes, Immobilized
Glucose
Glucose Oxidase*
Hydrogen Peroxide
Reproducibility of Results

Substances

Enzymes, Immobilized
Cellulose
Hydrogen Peroxide
Glucose Oxidase
Glucose