2. Academic Validation
  3. Metabolic engineering of Escherichia coli for producing adipic acid through the reverse adipate-degradation pathway

Metabolic engineering of Escherichia coli for producing adipic acid through the reverse adipate-degradation pathway

  • Metab Eng. 2018 May:47:254-262. doi: 10.1016/j.ymben.2018.04.002.
Mei Zhao 1,  Dixuan Huang 1,  Xiaojuan Zhang 2,  Mattheos A G Koffas 3,  Jingwen Zhou 4,  Yu Deng 5
Affiliations

Affiliations

  • 1 National Engineering Laboratory for Cereal Fermentation Technology (NELCF), Jiangnan University, 1800 Lihu Road, Wuxi, Jiangsu 214122, China; School of Biotechnology, Jiangnan University, 1800 Lihu Rd, Wuxi, Jiangsu 214122, China.
  • 2 National Engineering Laboratory for Cereal Fermentation Technology (NELCF), Jiangnan University, 1800 Lihu Road, Wuxi, Jiangsu 214122, China.
  • 3 Department of Chemical and Biological Engineering, Rensselaer Polytechnic Institute, Troy, NY 12180, USA; Department of Biological Sciences, Rensselaer Polytechnic Institute, Troy NY 12180, USA.
  • 4 National Engineering Laboratory for Cereal Fermentation Technology (NELCF), Jiangnan University, 1800 Lihu Road, Wuxi, Jiangsu 214122, China; School of Biotechnology, Jiangnan University, 1800 Lihu Rd, Wuxi, Jiangsu 214122, China. Electronic address: zhoujw1982@jiangnan.edu.cn.
  • 5 National Engineering Laboratory for Cereal Fermentation Technology (NELCF), Jiangnan University, 1800 Lihu Road, Wuxi, Jiangsu 214122, China; School of Biotechnology, Jiangnan University, 1800 Lihu Rd, Wuxi, Jiangsu 214122, China. Electronic address: dengyu@jiangnan.edu.cn.
PMID: 29625225 DOI: 10.1016/j.ymben.2018.04.002
Abstract

Adipic acid is an important dicarboxylic acid mainly used for the production of nylon 6-6 fibers and resins. Previous studies focused on the biological production of adipic acid directly from different substrates, resulting in low yields and titers. In this study, a five-step reverse adipate-degradation pathway (RADP) identified in Thermobifida fusca has been reconstructed in Escherichia coli BL21 (DE3). The resulting strain (Mad136) produced 0.3 g L-1 adipic acid with a 11.1% theoretical yield in shaken flasks, and we confirmed that the step catalyzed by 5-Carboxy-2-pentenoyl-CoA reductase (Tfu_1647) as the rate-limiting step of the RADP. Overexpression of Tfu_1647 by pTrc99A carried by strain Mad146 produced with a 49.5% theoretical yield in shaken flasks. We further eliminated pathways for major metabolites competing for carbon flux by CRISPR/Cas9 and deleted the succinate-CoA Ligase gene to promote accumulation of succinyl-CoA, which is the precursor for adipic acid synthesis. The final engineered strain Mad123146, which could achieve 93.1% of the theoretical yield in the shaken flask, was able to produce 68.0 g L-1 adipic acid by fed-batch fermentation. To the best of our knowledge, these results constitute the highest adipic acid titer reported in E. coli.

Keywords

Adipic acid; E. coli; High titer; Metabolic engineering; The reverse adipate degradation pathway.

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