title: Amperometric L-glutamate biosensor based on bacterial cell-surface displayed glutamate dehydrogenase.
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.
authors.affiliation: 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.
authors.forename: Bo
authors.initials: B
authors.lastname: Liang
authors.affiliation: 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.
authors.forename: Shu
authors.initials: S
authors.lastname: Zhang
authors.affiliation: 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.
authors.forename: Qiaolin
authors.initials: Q
authors.lastname: Lang
authors.affiliation: Key Laboratory of Marine Chemistry Theory and Technology of Ministry of Education, Ocean University of China, 238 Songling Road, Qingdao 266100, China.
authors.forename: Jianxia
authors.initials: J
authors.lastname: Song
authors.affiliation: Key Laboratory of Marine Chemistry Theory and Technology of Ministry of Education, Ocean University of China, 238 Songling Road, Qingdao 266100, China.
authors.forename: Lihui
authors.initials: L
authors.lastname: Han
authors.affiliation: 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.
authors.forename: Aihua
authors.initials: A
authors.lastname: Liu
mesh_terms: D001425:Bacterial Outer Membrane Proteins; D015374:Biosensing Techniques / Q000295:instrumentation / Q000379:methods*; D002138:Calibration; D002462:Cell Membrane / Q000378:metabolism; D055664:Electrochemical Techniques / Q000295:instrumentation*; D004566:Electrodes; D004795:Enzyme Stability; D004867:Equipment Design; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000378:metabolism; D005969:Glutamate Dehydrogenase / Q000235:genetics / Q000378:metabolism*; D018698:Glutamic Acid / Q000032:analysis*; D057230:Limit of Detection; D037742:Nanotubes, Carbon / Q000737:chemistry*; D010084:Oxidation-Reduction; D011094:Polyethyleneimine / Q000737:chemistry; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D001425:Bacterial Outer Membrane Proteins; D029968:Escherichia coli Proteins; D037742:Nanotubes, Carbon; D011994:Recombinant Proteins; C063113:ice nucleation protein; D018698:Glutamic Acid; D011094:Polyethyleneimine; D005969:Glutamate Dehydrogenase
keywords: Amperometric detection; Bacterial surface display; Electrochemical microbial biosensor; Glutamate dehydrogenase displayed bacteria; l-Glutamate biosensor