Abstract: We have developed a gamma-aminobutyric acid (GABA) production technique using his-tag mediated immobilization of Escherichia coli-derived glutamate decarboxylase (GAD), an enzyme that catalyzes the conversion of glutamate to GABA. The GAD was obtained at 1.43 g/L from GAD-overexpressed E. coli fermentation and consisted of 59.7% monomer, 29.2% dimer and 2.3% tetramer with a 97.6% soluble form of the total GAD. The harvested GAD was immobilized to metal affinity gel with an immobilization yield of 92%. Based on an investigation of specific enzyme activity and reaction characteristics, glutamic acid (GA) was chosen over monosodium glutamate (MSG) as a substrate for immobilized GAD, resulting in conversion of 2.17 M GABA in a 1 L reactor within 100 min. The immobilized enzymes retained 58.1% of their initial activities after ten consecutive uses. By using cation exchange chromatography followed by enzymatic conversion, GABA was separated from the residual substrate and leached GAD. As a consequence, the glutamic acid was mostly removed with no detectable GAD, while 91.2% of GABA was yielded in the purification step.
Keywords: gamma aminobutyric acid; glutamate decarboxylase; glutamic acid; immobilization; cation exchange chromatography
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Lee, S.; Ahn, J.; Kim, Y.-G.; Jung, J.-K.; Lee, H.; Lee, E.G. Gamma-Aminobutyric Acid Production Using Immobilized Glutamate Decarboxylase Followed by Downstream Processing with Cation Exchange Chromatography. Int. J. Mol. Sci. 2013, 14, 1728-1739.
Lee S, Ahn J, Kim Y-G, Jung J-K, Lee H, Lee EG. Gamma-Aminobutyric Acid Production Using Immobilized Glutamate Decarboxylase Followed by Downstream Processing with Cation Exchange Chromatography. International Journal of Molecular Sciences. 2013; 14(1):1728-1739.
Lee, Seungwoon; Ahn, Jungoh; Kim, Yeon-Gu; Jung, Joon-Ki; Lee, Hongweon; Lee, Eun G. 2013. "Gamma-Aminobutyric Acid Production Using Immobilized Glutamate Decarboxylase Followed by Downstream Processing with Cation Exchange Chromatography." Int. J. Mol. Sci. 14, no. 1: 1728-1739.