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Purification and characterization of glutamate decarboxylase from Enterococcus raffinosus TCCC11660

  • Biocatalysis - Original Paper
  • Published:
Journal of Industrial Microbiology & Biotechnology

Abstract

Glutamate decarboxylase (GAD) is the sole enzyme that synthesizes γ-aminobutyric acid through the irreversible decarboxylation of l-glutamate. In this study, the purification and characterization of an unreported GAD from a novel strain of Enterococcus raffinosus TCCC11660 were investigated. The native GAD from E. raffinosus TCCC11660 was purified 32.3-fold with a recovery rate of 8.3%, using ultrafiltration and ammonium sulfate precipitation, followed by ion-exchange and size-exclusion chromatography. The apparent molecular weight of purified GAD, as determined by SDS-PAGE and size-exclusion chromatography was 55 and 110 kDa, respectively, suggesting that GAD exists as a dimer of identical subunits in solution. In the best sodium citrate buffer, metal ions of Mo6+ and Mg2+ had positive effects, while Cu2+, Fe2+, Zn2+ and Co2+ showed significant adverse effects on enzyme activity. The optimum pH and temperature of GAD were determined to be 4.6 and 45 °C, while the K m and V max values for the sole l-glutamate substrate were 5.26 and 3.45 μmol L−1 min−1, respectively.

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Acknowledgements

This work was financially supported by the National Basic Research Program (973 Program) of China (2013CB734004), the Natural Science Foundation of China (31370075, 31471725 and 61603273), and the Youth Innovation Fund of Tianjin University of Science and Technology of China (2014CXLG28).

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Authors and Affiliations

  1. Key Laboratory of Industrial Fermentation Microbiology, Ministry of Education, College of Biotechnology, Tianjin University of Science and Technology, Tianjin, 300457, People’s Republic of China

    Chuanyou Chang, Jun Zhang, Shenxi Ma, Depei Wang, Jian Zhang & Qiang Gao

  2. School of Computer Science and Information Engineering, Tianjin University of Science and Technology, Tianjin, 300457, People’s Republic of China

    Lin Wang

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  1. Chuanyou Chang
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  2. Jun Zhang
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  4. Lin Wang
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  6. Jian Zhang
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  7. Qiang Gao
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Correspondence to Qiang Gao.

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Chang, C., Zhang, J., Ma, S. et al. Purification and characterization of glutamate decarboxylase from Enterococcus raffinosus TCCC11660. J Ind Microbiol Biotechnol 44, 817–824 (2017). https://doi.org/10.1007/s10295-017-1906-3

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