Eliminating acetate formation improves citramalate production by metabolically engineered Escherichia coli

Microbial Cell Factories

Research

Xianghao Wu¹  Naga Sirisha Parimi¹  Mark A. Eiteman¹  Ian A. Durie¹  Afaq M. M. Niyas¹ 

[1] School of Chemical, Materials and Biomedical Engineering, Driftmier Engineering Center, University of Georgia, 30602, Athens, GA, USA;

关键词: Acetyl-CoA;    Pyruvate;    Citramalate;    Escherichia coli;    Acetate;    Glucose;    Fed-batch;   

DOI  :  10.1186/s12934-017-0729-2

 received in 2017-02-21, accepted in 2017-06-19,  发布年份 2017

来源: Springer

【 摘 要 】

BackgroundCitramalate, a chemical precursor to the industrially important methacrylic acid (MAA), can be synthesized using Escherichia coli overexpressing citramalate synthase (cimA gene). Deletion of gltA encoding citrate synthase and leuC encoding 3-isopropylmalate dehydratase were critical to achieving high citramalate yields. Acetate is an undesirable by-product potentially formed from pyruvate and acetyl-CoA, the precursors of citramalate during aerobic growth of E. coli. This study investigated strategies to minimize acetate and maximize citramalate production in E. coli mutants expressing the cimA gene.ResultsKey knockouts that minimized acetate formation included acetate kinase (ackA), phosphotransacetylase (pta), and in particular pyruvate oxidase (poxB). Deletion of glucose 6-phosphate dehydrogenase (zwf) and ATP synthase (atpFH) aimed at improving glycolytic flux negatively impacted cell growth and citramalate accumulation in shake flasks. In a repetitive fed-batch process, E. coli gltA leuC ackA-pta poxB overexpressing cimA generated 54.1 g/L citramalate with a yield of 0.64 g/g glucose (78% of theoretical maximum yield), and only 1.4 g/L acetate in 87 h.ConclusionsThis study identified the gene deletions critical to reducing acetate accumulation during aerobic growth and citramalate production in metabolically engineered E. coli strains. The citramalate yield and final titer relative to acetate at the end of the fed-batch process are the highest reported to date (a mass ratio of citramalate to acetate of nearly 40) without being detrimental to citramalate productivity, significantly improving a potential process for the production of this five-carbon chemical.

【 授权许可】

CC BY   
© The Author(s) 2017