Escherichia coli is engineered for γ‐aminobutyrate (GABA) production in glucose minimal medium. For this, overexpression of mutant glutamate decarboxylase (GadB) and mutant glutamate/GABA antiporter (GadC), as well as deletion of GABA transaminase (GabT), are accomplished. In addition, the carbon flux to the tricarboxylic acid cycle is engineered by the overexpression of gltA , ppc , or both. The overexpression of citrate synthase (CS), encoded by gltA , increases GABA productivity, as expected. Meanwhile, the overexpression of phosphoenolpyruvate carboxylase (PPC) causes a decrease in the rate of glucose uptake, resulting in a decrease in GABA production. The phenotypes of the strains are characterized by ¹³C metabolic flux analysis (¹³C MFA). The results reveal that CS overexpression increases glycolysis and anaplerotic reaction rates, as well as the citrate synthesis rate, while PPC overexpression causes little changes in metabolic fluxes, but reduces glucose uptake rate. The engineered strain produces 1.2 g L⁻¹ of GABA from glucose. Thus, by using ¹³C MFA, important information is obtained for designing metabolically engineered strains for efficient GABA production.

中文翻译：

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为生产γ-氨基丁酸而设计的大肠杆菌的13 C代谢通量分析。

大肠杆菌经过工程设计，可在葡萄糖基本培养基中生产γ-氨基丁酸酯（GABA）。为此，实现了突变型谷氨酸脱羧酶（GadB）和突变型谷氨酸/ GABA反转运蛋白（GadC）的过表达，以及GABA转氨酶（GabT）的缺失。此外，通向gltA，ppc或两者的过表达可以控制三羧酸循环的碳通量。如预期的那样，由gltA编码的柠檬酸合酶（CS）的过表达提高了GABA的生产率。同时，磷酸烯醇丙酮酸羧化酶（PPC）的过表达引起葡萄糖摄取速率的降低，从而导致GABA产生的降低。菌株的表型特征在于¹³C代谢通量分析（¹³ C MFA）。结果表明，CS的过表达增加了糖酵解和动脉粥样硬化反应的速率，以及柠檬酸盐的合成速率，而PPC的过表达引起的代谢通量变化很小，但降低了葡萄糖的摄取速率。该工程菌株从葡萄糖产生1.2g L ^-1的GABA。因此，通过使用¹³ C MFA，可以获得用于设计代谢工程菌株以有效产生GABA的重要信息。