High level production of γ-aminobutyric acid in engineered Escherichia coli by refactoring the glutamate decarboxylase

Lyu, Jia-Fei; Lyu, Changjiang; Cao, Jia-Ren; Mei, Jiaqi; Hu, Sheng; Wang, Zhao; Xu, Tianyi; Wang, Yuting; Wang, Danli; Huang, Jun; Mei, Lehe

doi:10.1016/j.procbio.2022.04.036

Cited by 5 publications

(3 citation statements)

References 25 publications

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“…Preparation of pure enzyme solution as described above, set up 1 mL of mixed reaction system including 15 g/L MSG, 0.1 mM PLP, 0.4 U purified enzyme and 50 mM Na 2 HPO 4 -citric acid buffer, pH 5, 50 • C (unless otherwise stated), and the yield of GABA was measured by TLC after 2 h of reaction. The effect of reaction conditions on the conversion of GABA was determined by adjusting the amount of enzyme added (0.05-0.8 U), reaction temperature (30-80 • C), pH (3)(4)(5)(6)(7)(8), MSG concentration (5-25 g/L) and reaction time (1-6 h).…”

Section: Examination Of the Ability Of Lcgad10s Catalyzed Synthesis O...mentioning

confidence: 99%

“…The effect of pH (3)(4)(5)(6)(7)(8) on the yield of GABA was investigated. As shown in Figure 4c, the enhancement of GABA concentration with increasing initial pH from 4 to 5, where the maximum GABA production was obtained, followed by a reduction of GABA production when the initial pH exceeded 7.…”

Section: Examination Of the Ability Of Lcgad10s Catalyzed Synthesis O...mentioning

confidence: 99%

“…Glutamate decarboxylase (GAD; EC 4.1.1.15) is a key enzyme that catalyzes the irreversible α-decarboxylation of L-glutamate (Glu) to GABA [7]. The enzymatic reaction is a more promising approach for the synthesis of GABA through GADs [4].…”

Section: Introductionmentioning

confidence: 99%

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Expression and Transformation Characteristics of a Novel Glutamic Acid Decarboxylase LcGAD10s and Its Application on Sufu Processing

Chen,

Wang,

Song

et al. 2023

Foods

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Gamma-aminobutyric acid (GABA) is an important non-proteinogenic amino acid and a potent bioactive compound with many anti-hypertensive and anti-depressant activities. The bioconversion of GABA by glutamic acid decarboxylase (GAD) has been eagerly studied. Herein, novel pyridoxal-5-phosphate monohydrates (PLP)-dependent GAD, which is not quite similar to reporting, was cloned from Latilactobacillus curvatus and efficiently expressed in E. coli. The conveniently purified GAD (designated LcGAD10s) appeared as a single protein on SDS-PAGE with a molecular mass of 52.0 kDa. LcGAD10s exhibited a specific activity of 303.7 U/mg after purification by Ni–IDA affinity chromatography, with optimal activity at 55 °C and pH 5. LcGAD10s displayed excellent temperature (50 °C) and pH (4–8) stability which relative activity above 80% and 70%, respectively. The enzymatic activity was, respectively, increased and depressed by 130%, and 24% in the presence of Mn+ and Cu2+. Enzyme activity over 90% can be achieved by adding at least 25 mM of PLP. LcGAD10s was able to efficiently transform 15 g/L GABA with a single-factor optimized reaction of pH (5), temperature (50 °C), time (2 h), LcGAD10s dosage (0.4 U) and monosodium glutamate level (5 g/L). Additionally, LcGAD10s can be applied to a tofu fermentation system to achieve GABA conversion and achieved 14.9 mg/g of GABA conversion when added at 2 U/mL, which is higher than most of the commercial sufu and previous application reports, increasing its functional substances.

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Section: Examination Of the Ability Of Lcgad10s Catalyzed Synthesis O...mentioning

confidence: 99%

Section: Examination Of the Ability Of Lcgad10s Catalyzed Synthesis O...mentioning

confidence: 99%

See 1 more Smart Citation

Expression and Transformation Characteristics of a Novel Glutamic Acid Decarboxylase LcGAD10s and Its Application on Sufu Processing

Chen,

Wang,

Song

et al. 2023

Foods

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High-level production of γ-aminobutyric acid via efficient co-expression of the key genes of glutamate decarboxylase system in Escherichia coli

Yao¹,

Lyu²,

Wang³

et al. 2023

Engineering Microbiology

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Directed evolution of glutamate decarboxylase B for enhancing its enzyme activity towards nearly neutral pHs based on error-prone PCR

Guan,

Fu,

Wang

et al. 2024

International Journal of Biological Macromolecules

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High level production of γ-aminobutyric acid in engineered Escherichia coli by refactoring the glutamate decarboxylase

Cited by 5 publications

References 25 publications

Expression and Transformation Characteristics of a Novel Glutamic Acid Decarboxylase LcGAD10s and Its Application on Sufu Processing

Expression and Transformation Characteristics of a Novel Glutamic Acid Decarboxylase LcGAD10s and Its Application on Sufu Processing

High-level production of γ-aminobutyric acid via efficient co-expression of the key genes of glutamate decarboxylase system in Escherichia coli

Directed evolution of glutamate decarboxylase B for enhancing its enzyme activity towards nearly neutral pHs based on error-prone PCR

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