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Bacterial cell-surface displaying of thermo-tolerant glutamate dehydrogenase and its application in l-glutamate assay

Paper ID Volume ID Publish Year Pages File Format Full-Text
17052 42636 2015 7 PDF Available
Title
Bacterial cell-surface displaying of thermo-tolerant glutamate dehydrogenase and its application in l-glutamate assay
Abstract

•Gldh expressed efficiently on the surface of Escherichia coli using INP display system.•Gldh strain showed excellent enzyme activity, specificity and stability.•Gldh-displayed whole cell biocatalyst capable of detecting l-glutamate sensitively.

In this paper, glutamate dehydrogenase (Gldh) is reported to efficiently display on Escherichia coli cell surface by using N-terminal region of ice the nucleation protein as an anchoring motif. The presence of Gldh was confirmed by SDS-PAGE and enzyme activity assay. Gldh was detected mainly in the outer membrane fraction, suggesting that the Gldh was displayed on the bacterial cell surface. The optimal temperature and pH for the bacteria cell-surface displayed Gldh (bacteria-Gldh) were 70 °C and 9.0, respectively. Additionally, the fusion protein retained almost 100% of its initial enzymatic activity after 1 month incubation at 4 °C. Transition metal ions could inhibit the enzyme activity to different extents, while common anions had little adverse effect on enzyme activity. Importantly, the displayed Gldh is most specific to l-glutamate reported so far. The bacterial Gldh was enabled to catalyze oxidization of l-glutamate with NADP+ as cofactor, and the resultant NADPH can be detected spectrometrically at 340 nm. The bacterial-Gldh based l-glutamate assay was established, where the absorbance at 340 nm increased linearly with the increasing l-glutamate concentration within the range of 10 − 400 μM. Further, the proposed approach was successfully applied to measure l-glutamate in real samples.

Keywords
Bacterial surface display; l-Glutamate; Thermo-tolerant glutamate dehydrogenase; Enzyme inhibition; l-Glutamate detection
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Publisher
Database: Elsevier - ScienceDirect
Journal: Enzyme and Microbial Technology - Volume 70, March 2015, Pages 72–78
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us