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Anal Chim Acta. 2015 Jul 16;884:83-9. doi: 10.1016/j.aca.2015.05.012. Epub 2015 May 12.

Amperometric L-glutamate biosensor based on bacterial cell-surface displayed glutamate dehydrogenase.

Author information

1
Laboratory for Biosensing, Key Laboratory of Biofuels, and Shandong Provinicial Key Laboratory of Energy Genetics, Qingdao Institute of Bioenergy & Bioprocess Technology, Chinese Academy of Sciences, 189 Songling Road, Qingdao 266101, China; University of Chinese Academy of Sciences, 19A Yuquan Road, Beijing 100049, China.
2
Laboratory for Biosensing, Key Laboratory of Biofuels, and Shandong Provinicial Key Laboratory of Energy Genetics, Qingdao Institute of Bioenergy & Bioprocess Technology, Chinese Academy of Sciences, 189 Songling Road, Qingdao 266101, China; Key Laboratory of Marine Chemistry Theory and Technology of Ministry of Education, Ocean University of China, 238 Songling Road, Qingdao 266100, China.
3
Laboratory for Biosensing, Key Laboratory of Biofuels, and Shandong Provinicial Key Laboratory of Energy Genetics, Qingdao Institute of Bioenergy & Bioprocess Technology, Chinese Academy of Sciences, 189 Songling Road, Qingdao 266101, China.
4
Key Laboratory of Marine Chemistry Theory and Technology of Ministry of Education, Ocean University of China, 238 Songling Road, Qingdao 266100, China.
5
Laboratory for Biosensing, Key Laboratory of Biofuels, and Shandong Provinicial Key Laboratory of Energy Genetics, Qingdao Institute of Bioenergy & Bioprocess Technology, Chinese Academy of Sciences, 189 Songling Road, Qingdao 266101, China; University of Chinese Academy of Sciences, 19A Yuquan Road, Beijing 100049, China. Electronic address: liuah@qibebt.ac.cn.

Abstract

A novel L-glutamate biosensor was fabricated using bacteria surface-displayed glutamate dehydrogenase (Gldh-bacteria). Here the cofactor NADP(+)-specific dependent Gldh was expressed on the surface of Escherichia coli using N-terminal region of ice nucleation protein (INP) as the anchoring motif. The cell fractionation assay and SDS-PAGE analysis indicated that the majority of INP-Gldh fusion proteins were located on the surface of cells. The biosensor was fabricated by successively casting polyethyleneimine (PEI)-dispersed multi-walled carbon nanotubes (MWNTs), Gldh-bacteria and Nafion onto the glassy carbon electrode (Nafion/Gldh-bacteria/PEI-MWNTs/GCE). The MWNTs could not only significantly lower the oxidation overpotential towards NAPDH, which was the product of NADP(+) involving in the oxidation of glutamate by Gldh, but also enhanced the current response. Under the optimized experimental conditions, the current-time curve of the Nafion/Gldh-bacteria/PEI-MWNTs/GCE was performed at +0.52 V (vs. SCE) by amperometry varying glutamate concentration. The current response was linear with glutamate concentration in two ranges (10 μM-1 mM and 2-10 mM). The low limit of detection was estimated to be 2 μM glutamate (S/N=3). Moreover, the proposed biosensor is stable, specific, reproducible and simple, which can be applied to real samples detection.

KEYWORDS:

Amperometric detection; Bacterial surface display; Electrochemical microbial biosensor; Glutamate dehydrogenase displayed bacteria; l-Glutamate biosensor

PMID:
26073813
DOI:
10.1016/j.aca.2015.05.012
[Indexed for MEDLINE]

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