13C Metabolic Flux Analysis of Escherichia coli Engineered for Gamma-Aminobutyrate Production

Dae Kyun Im, Jaeseung Hong, Boncheol Gu, Changmin Sung, Min Kyu Oh

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Abstract

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.

Original languageEnglish
JournalBiotechnology Journal
DOIs
Publication statusAccepted/In press - 2020 Jan 1

Keywords

  • C metabolic flux analysis
  • metabolic engineering
  • tricarboxylic acid cycle
  • γ-aminobutyrate

ASJC Scopus subject areas

  • Applied Microbiology and Biotechnology
  • Molecular Medicine

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