Salt‐free production of γ‐aminobutyric acid from glutamate using glutamate decarboxylase separated from <i>Escherichia coli</i>

Dinh, Thu Huong; Ho, Ngoc Anh Thu; Kang, Taek Jin; McDonald, Karen A.; Won, Keehoon

doi:10.1002/jctb.4251

Cited by 14 publications

(10 citation statements)

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“…Purified GAD and natural and recombinant microorganisms expressing GAD have been employed for the conversion of glutamate monosodium salt (MSG) or glutamic acid to GABA. Recent studies have suggested that GABA can be efficiently produced from L-glutamate by some lactic acid bacteria (LAB) [ 7 , 8 ] and recombinant Escherichia coli [ 5 , 9 , 10 ]. Basically, it is needed to develop additional enzymatic and whole cell mediated conversion of MSG into GABA when fermentation-derived MSG is used for the production of GABA.…”

Section: Introductionmentioning

confidence: 99%

Enhanced production of gamma-aminobutyrate (GABA) in recombinant Corynebacterium glutamicum by expressing glutamate decarboxylase active in expanded pH range

et al. 2015

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BackgroundGamma-aminobutylate (GABA) is an important chemical in pharmacetucal field and chemical industry. GABA has mostly been produced in lactic acid bacteria by adding L-glutamate to the culture medium since L-glutamate can be converted into GABA by inherent L-glutamate decarboxylase. Recently, GABA has gained much attention for the application as a major building block for the synthesis of 2-pyrrolidone and biodegradable polyamide nylon 4, which opens its application area in the industrial biotechnology. Therefore, Corynebacterium glutamicum, the major L-glutamate producing microorganism, has been engineered to achieve direct fermentative production of GABA from glucose, but their productivity was rather low.ResultsRecombinant C. glutamicum strains were developed for enhanced production of GABA from glucose by expressing Escherichia coli glutamate decarboxylase (GAD) mutant, which is active in expanded pH range. Synthetic PH36, PI16, and PL26 promoters, which have different promoter strengths in C. glutamicum, were examined for the expression of E. coli GAD mutant. C. glutamicum expressing E. coli GAD mutant under the strong PH36 promoter could produce GABA to the concentration of 5.89 ± 0.35 g/L in GP1 medium at pH 7.0, which is 17-fold higher than that obtained by C. glutamicum expressing wild-type E. coli GAD in the same condition (0.34 ± 0.26 g/L). Fed-bath culture of C. glutamicum expressing E. coli GAD mutant in GP1 medium containing 50 μg/L of biotin at pH 6, culture condition of which was optimized in flask cultures, resulted in the highest GABA concentration of 38.6 ± 0.85 g/L with the productivity of 0.536 g/L/h.ConclusionRecombinant C. glutamicum strains developed in this study should be useful for the direct fermentative production of GABA from glucose, which allows us to achieve enhanced production of GABA suitable for its application area in the industrial biotechnology.

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Section: Introductionmentioning

confidence: 99%

Enhanced production of gamma-aminobutyrate (GABA) in recombinant Corynebacterium glutamicum by expressing glutamate decarboxylase active in expanded pH range

et al. 2015

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“…Since the crystal structures of E. coli GADs revealed the structural basis for its optimal activity at acidic pH, several mutants with high activity toward more alkaline pH values have been constructed to improve GABA production (Choi et al 2015 ; Shi et al 2014 ; Thu Ho et al 2013 ). On the other hand, hydrochloric acid, sodium acetate buffer and acidic cation-exchange resins were used to maintain the acidic condition during reaction from MSG into GABA, and the conversion efficiency were remarkably improved (Dinh et al 2014 ; Park et al 2013 ; Plokhov et al 2000 ). However, these methods were also unsatisfactory for the following separation and purification of GABA due to the introduction of the high amount of salts or resins.…”

Section: Introductionmentioning

confidence: 99%

Whole-cell conversion of l-glutamic acid into gamma-aminobutyric acid by metabolically engineered Escherichia coli

Yang

Rao

et al. 2016

SpringerPlus

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A simple and high efficient way for the synthesis of gamma-aminobutyric acid (GABA) was developed by using engineered Escherichia coli as whole-cell biocatalyst from l-glutamic acid (l-Glu). Codon optimization of Lactococcus lactis GadB showed the best performance on GABA production when middle copy-number plasmid was used as expression vector in E. coli BW25113. The highest production of GABA reached 308.96 g L−1 with 99.9 mol% conversion within 12 h, when E. coli ΔgabAB (pRB-lgadB) concentrated to an OD600 of 15 in 3 M l-Glu at 45 °C. Furthermore, the strain could be reused at least three cycles in 2 M crude l-Glu with an average productivity of 40.94 g L−1 h−1. The total GABA yield reached 614.15 g L−1 with a molar yield over 99 %, which represented the highest GABA production ever reported. The whole-cell bioconversion system allowed us to achieve a promising cost-effective resource for GABA in industrial application.Electronic supplementary materialThe online version of this article (doi:10.1186/s40064-016-2217-2) contains supplementary material, which is available to authorized users.

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“…Comparison of LlGAD with other GAD proteins showed LlGAD contained a highly conserved motif (HVDAAFGG) that belongs to the PLP-dependent decarboxylase superfamily [15]. The domain includes a conserved lysine residue essential for PLP binding discovered in LlGAD (K 277 ), and this lysine residue was located with the adjacent histidine residue [16]. In addition, the active site residues involved in promoting decarboxylation were also present in LlGAD (T 213 and D 244 ) [15] (Figure 1).…”

Section: Resultsmentioning

confidence: 99%

“…The optimum induction conditions for the expression of GAD in E. coli were investigated by single-factor experimental design, i.e. the recombinant cells induced by different concentrations of IPTG (0.4, 0.7, 1.0, 1.3 and 1.6 mmol L ¡1 ) added to the exponentially growing cells, different induction times (10,12,16, 18 and 20 h) and different induction temperatures (26,28,30, 32 and 34 C). The single-factor experiments were conducted in triplicate for each group and mean values with standard deviations ( §SD) were calculated for GABA production and cell density.…”

Section: Optimization Of Induction Conditionsmentioning

confidence: 99%

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Efficient gamma-aminobutyric acid bioconversion by engineered Escherichia coli

Wei

Ren

et al. 2018

Biotechnology & Biotechnological Equipment

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Gamma-aminobutyric acid (GABA) can be converted into 2-pyrrolidone, an intermediate in the synthesis of nylon 4 and agrochemicals, which has great potential for application in the chemical industry. The main aim of this work was to construct a simple and efficient process for industrial production of GABA from L-glutamic acid (L-Glu) by whole-cell bioconversion. The gene encoding glutamate decarboxylase (GAD) from Lactococcus lactis FJNUGA01 was overexpressed using the most available vector in Escherichia coli, pET-28a. GABA production was enhanced by optimizing the concentration of the catalyst isopropyl-b-D-thiogalactoside (IPTG), induction temperature and induction time. After optimization, GABA production reached 98.4 g L ¡1 with 96 mol% conversion within 6 h was achieved by whole-cell conversion using the cells induced with 1 mmol L ¡1 IPTG for 12 h at 30 C. Furthermore, when the initial concentration of L-Glu was adjusted, 204.12 g L ¡1 GABA with 34 g L ¡1 h ¡1 productivity was reached using 2 mol L ¡1 L-Glu as the substrate. During the 6 h whole-cell bioconversion process, almost all L-Glu was converted to GABA with a conversion efficiency of »99%. The high conversion efficiency and high productivity made the GABA purification occur downstream, and the whole process is more simple and economical for industrial application.

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Salt‐free production of γ‐aminobutyric acid from glutamate using glutamate decarboxylase separated from Escherichia coli

Cited by 14 publications

References 20 publications

Enhanced production of gamma-aminobutyrate (GABA) in recombinant Corynebacterium glutamicum by expressing glutamate decarboxylase active in expanded pH range

Enhanced production of gamma-aminobutyrate (GABA) in recombinant Corynebacterium glutamicum by expressing glutamate decarboxylase active in expanded pH range

Whole-cell conversion of l-glutamic acid into gamma-aminobutyric acid by metabolically engineered Escherichia coli

Efficient gamma-aminobutyric acid bioconversion by engineered Escherichia coli

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