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Volume 10
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10.3390/catal10020215
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Article

Active Expression of Membrane-Bound L-Amino Acid Deaminase from Proteus mirabilis in Recombinant Escherichia coli by Fusion with Maltose-Binding Protein for Enhanced Catalytic Performance

by
Dan-Ping Zhang
1,
Xiao-Ran Jing
1,
An-Wen Fan
1,
Huan Liu
1,
Yao Nie
1,2,* and
Yan Xu
1,3
1
School of Biotechnology and Key laboratory of Industrial Biotechnology, Ministry of Education, Jiangnan University, Wuxi 214122, China
2
Suqian Industrial Technology Research Institute of Jiangnan University, Suqian 223814, China
3
State Key Laboratory of Food Science and Technology, Jiangnan University, Wuxi 214122, China
*
Author to whom correspondence should be addressed.
Catalysts 2020, 10(2), 215; https://doi.org/10.3390/catal10020215
Submission received: 24 December 2019 / Revised: 29 January 2020 / Accepted: 6 February 2020 / Published: 10 February 2020
(This article belongs to the Special Issue Industrial Biocatalysis: Challenges and Opportunities)
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Abstract

L-amino acid deaminases (LAADs) are membrane flavoenzymes that catalyze the deamination of neutral and aromatic L-amino acids to α-keto acids and ammonia. LAADs can be used to develop many important biotechnological applications. However, the transmembrane α-helix of LAADs restricts its soluble active expression and purification from a heterologous host, such as Escherichia coli. Herein, through fusion with the maltose-binding protein (MBP) tag, the recombinant E. coli BL21 (DE3)/pET-21b-MBP-PmLAAD was constructed and the LAAD from Proteus mirabilis (PmLAAD) was actively expressed as a soluble protein. After purification, the purified MBP-PmLAAD was obtained. Then, the catalytic activity of the MBP-PmLAAD fusion protein was determined and compared with the non-fused PmLAAD. After fusion with the MBP-tag, the catalytic efficiency of the MBP-PmLAAD cell lysate was much higher than that of the membrane-bound PmLAAD whole cells. The soluble MBP-PmLAAD cell lysate catalyzed the conversion of 100 mM L-phenylalanine (L-Phe) to phenylpyruvic acid (PPA) with a 100% yield in 6 h. Therefore, the fusion of the MBP-tag not only improved the soluble expression of the PmLAAD membrane-bound protein, but also increased its catalytic performance.
Keywords: L-amino acid deaminases; membrane-bound protein; fusion protein; soluble expression; maltose-binding protein tag L-amino acid deaminases; membrane-bound protein; fusion protein; soluble expression; maltose-binding protein tag

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MDPI and ACS Style

Zhang, D.-P.; Jing, X.-R.; Fan, A.-W.; Liu, H.; Nie, Y.; Xu, Y. Active Expression of Membrane-Bound L-Amino Acid Deaminase from Proteus mirabilis in Recombinant Escherichia coli by Fusion with Maltose-Binding Protein for Enhanced Catalytic Performance. Catalysts 2020, 10, 215. https://doi.org/10.3390/catal10020215

AMA Style

Zhang D-P, Jing X-R, Fan A-W, Liu H, Nie Y, Xu Y. Active Expression of Membrane-Bound L-Amino Acid Deaminase from Proteus mirabilis in Recombinant Escherichia coli by Fusion with Maltose-Binding Protein for Enhanced Catalytic Performance. Catalysts. 2020; 10(2):215. https://doi.org/10.3390/catal10020215

Chicago/Turabian Style

Zhang, Dan-Ping, Xiao-Ran Jing, An-Wen Fan, Huan Liu, Yao Nie, and Yan Xu. 2020. "Active Expression of Membrane-Bound L-Amino Acid Deaminase from Proteus mirabilis in Recombinant Escherichia coli by Fusion with Maltose-Binding Protein for Enhanced Catalytic Performance" Catalysts 10, no. 2: 215. https://doi.org/10.3390/catal10020215

APA Style

Zhang, D.-P., Jing, X.-R., Fan, A.-W., Liu, H., Nie, Y., & Xu, Y. (2020). Active Expression of Membrane-Bound L-Amino Acid Deaminase from Proteus mirabilis in Recombinant Escherichia coli by Fusion with Maltose-Binding Protein for Enhanced Catalytic Performance. Catalysts, 10(2), 215. https://doi.org/10.3390/catal10020215

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