Applied Microbiology and Biotechnology

, Volume 102, Issue 10, pp 4511–4522 | Cite as

Metabolic engineering of Clostridium tyrobutyricum for enhanced butyric acid production with high butyrate/acetate ratio

  • Yukai Suo
  • Mengmeng Ren
  • Xitong Yang
  • Zhengping Liao
  • Hongxin Fu
  • Jufang Wang
Bioenergy and biofuels


Butyric acid fermentation by Clostridium couples with the synthesis of acetic acid. But the presence of acetic acid reduces butyric acid yield and increases separation and purification costs of butyric acid. Hence, enhancing the butyrate/acetate ratio is important for economical butyric acid production. This study indicated that enhancing the acetyl-CoA to butyrate flux by overexpression of both the butyryl-CoA/acetate CoA transferase (cat1) and crotonase (crt) genes in C. tyrobutyricum could significantly reduce acetic acid concentration. Fed-batch fermentation of ATCC 25755/cat1 + crt resulted in increased butyrate/acetate ratio of 15.76 g/g, which was 2.24-fold higher than that of the wild-type strain. Furthermore, in order to simultaneously increase the butyrate/acetate ratio, butyric acid concentration and productivity, the recombinant strain ATCC 25755/ppcc (co-expression of 6-phosphofructokinase (pfkA) gene, pyruvate kinase (pykA) gene, cat1, and crt) was constructed. Consequently, ATCC 25755/ppcc produced more butyric acid (46.8 vs. 35.0 g/L) with a higher productivity (0.83 vs. 0.49 g/L·h) and butyrate/acetate ratio (13.22 vs. 7.22 g/g) as compared with the wild-type strain in batch fermentation using high glucose concentration (120 g/L). This study demonstrates that enhancing the acetyl-CoA to butyrate flux is an effective way to reduce acetic acid production and increase butyrate/acetate ratio.


Metabolic engineering Butyric acid Clostridium tyrobutyricum Butyrate/acetate ratio Butyryl-CoA/acetate CoA transferase Crotonase 



This work was funded by the National Natural Science Foundation of China (21676098), the State Key Laboratory of Pulp and Paper Engineering (2017C03), the China Postdoctoral Science Foundation Funded Project (2017M612667), and the Fundamental Research Funds for the Central Universities (2017BQ084).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no competing interests.

Ethical approval

This article does not contain any studies with human participants or animals performed by any of the authors.


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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  1. 1.School of Biology and Biological EngineeringSouth China University of TechnologyGuangzhouChina
  2. 2.State Key Laboratory of Pulp and Paper EngineeringSouth China University of TechnologyGuangzhouChina

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