Engineering a synthetic pathway for maleate in Escherichia coli

Shuhei Noda,Yutaro Mori,Akihiko Kondo,Sachiko Oyama,Tomokazu Shirai

doi:10.1038/s41467-017-01233-9

Shuhei Noda, Yutaro Mori + Show 3 more

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https://doi.org/10.1038/s41467-017-01233-9

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Abstract

Maleate is one of the most important dicarboxylic acids and is used to produce various polymer compounds and pharmaceuticals. Herein, microbial production of maleate is successfully achieved, to our knowledge for the first time, using genetically modified Escherichia coli. A synthetic pathway of maleate is constructed in E. coli by combining the polyketide biosynthesis pathway and benzene ring cleavage pathway. The metabolic engineering approach used to fine-tune the synthetic pathway drastically improves maleate production and demonstrates that one of the rate limiting steps exists in the conversion of chorismate to gentisate. In a batch culture of the optimised transformant, grown in a 1-L jar fermentor, the amount of produced maleate reaches 7.1 g L−1, and the yield is 0.221 mol mol−1. Our results suggest that the construction of synthetic pathways by combining a secondary metabolite pathway and the benzene ring cleavage pathway is a powerful tool for producing various valuable chemicals.

Highlights

Maleate is one of the most important dicarboxylic acids and is used to produce various polymer compounds and pharmaceuticals
To create an E. coli strain capable of producing gentisate from glucose, we used the 3HBAproducing E. coli strain which is the derivative strain of CFT5
Maleate does not appear in metabolic pathways in most microorganisms, whereas succinate, malate and fumarate are usually produced in the tricarboxylic acid (TCA) cycle[14,15,16]

Summary

Introduction

Maleate is one of the most important dicarboxylic acids and is used to produce various polymer compounds and pharmaceuticals. Fumaric acid is a starting unit for polymerisation and esterification reactions in the production of unsaturated polyester resins and food or beverage additives[7, 15] Production of these organic acids is usually achieved in microorganisms via the tricarboxylic acid (TCA) cycle. Maleic acid is formed in the benzene ring cleavage pathway, which contributes to the degradation of environmental pollutants by some microbes[22, 23] (Fig. 1). In this pathway, aromatic chemicals such as naphthalene, cresol and xylenol are converted via several steps to 3-hydroxybenzoate (3HBA) and gentisate (2,5dihydroxybenzoic acid), which are the key intermediates of the pathway. Microorganisms capable of degrading aromatic compounds do not have the ability to endogenously produce gentisate from sugars or other renewable carbon sources via the central metabolic microbial pathways such as the glycolysis pathway or pentose phosphate pathway

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