Gene Mining, Expression and Identification of Novel Glutamate Decarboxylases Based on Genome Mining

doi:10.7506/spkx1002-6630-20200102-012

FOOD SCIENCE ›› 2021, Vol. 42 ›› Issue (10): 79-85.doi: 10.7506/spkx1002-6630-20200102-012

• Bioengineering • Previous Articles Next Articles

Gene Mining, Expression and Identification of Novel Glutamate Decarboxylases Based on Genome Mining

LI Xiang, XIE Yuli, JIA Yuanyuan, ZHANG Shan, WANG Hongyan, LIU Xianli, LUO Xiangyan, TANG Cunduo

(1. Henan Provincial Engineering Laboratory of Insect Bio-reactor, Nanyang Normal University, Nanyang 473061, China; 2. State Key Laboratory of Automotive Biofuel Technology, Nanyang 473061, China)

Online:2021-05-25 Published:2021-06-02

Abstract

Abstract: In this study, three hypothesized glutamate decarboxylases (GAD) genes from Lactococcus lactis, Lactobacillus senmaizukei and Enterococcus sulfureus, respectively named as LlGAD, LsGAD and EsGAD, were excavated using genome mining technology with the high-activity GAD gene (LbGAD) from Lactobacillus brevis as the probe. By means of pET28a plasmid, these four genes were expressed in E. coli BL21. The expressed products of LsGAD and LlGAD had a good solubility, exhibiting GAD activity of 34.17 and 38.91 U/mL, respectively in the fermentation broths. The specific activity, temperature characteristics, pH characteristics and Kcat/Km value of LsGAD were significantly superior to those of the other GAD enzymes. In addition, the biosynthesis of γ-aminobutyric acid (GABA) from L-glutamic acid by whole-cell catalysis was studied. When L-glutamic acid at 6 g/L was transformed for 24 h, the maximum yield of GABA of 58% was obtained. In conclusion, this study has realized a leap from genomic data to real enzymes by obtaining GAD with excellent performance as well as the biosynthesis of GABA, which will lay a solid foundation for the low-cost and large-scale biosynthesis of GABA.

Key words: glutamate decarboxylase; genome mining; expression; identification; biocatalysis

CLC Number:

TS202.1

LI Xiang, XIE Yuli, JIA Yuanyuan, ZHANG Shan, WANG Hongyan, LIU Xianli, LUO Xiangyan, TANG Cunduo. Gene Mining, Expression and Identification of Novel Glutamate Decarboxylases Based on Genome Mining[J]. FOOD SCIENCE, 2021, 42(10): 79-85.

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Gene Mining, Expression and Identification of Novel Glutamate Decarboxylases Based on Genome Mining

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