Metabolic evolution and a comparative omics analysis of Corynebacterium glutamicum for putrescine production

  • Zhen Li
  • Yu-Ping Shen
  • Xuan-Long Jiang
  • Li-Shen Feng
  • Jian-Zhong Liu
Metabolic Engineering and Synthetic Biology - Original Paper
First Online:
Received:
Accepted:

Abstract

Putrescine is widely used in the industrial production of bioplastics, pharmaceuticals, agrochemicals, and surfactants. Because the highest titer of putrescine is much lower than that of its precursor l-ornithine reported in microorganisms to date, further work is needed to increase putrescine production in Corynebacterium glutamicum. We first compared 7 ornithine decarboxylase genes and found that the Enterobacter cloacae ornithine decarboxylase gene speC1 was most suitable for putrescine production in C. glutamicum. Increasing NADPH availability and blocking putrescine oxidation and acetylation were chosen as targets for metabolic engineering. The putrescine producer C. glutamicum PUT4 was first constructed by deleting puo, butA and snaA genes, and replacing the fabG gene with E. cloacae speC1. After adaptive evolution with C. glutamicum PUT4, the evolved strain C. glutamicum PUT-ALE, which produced an 96% higher amount of putrescine compared to the parent strain, was obtained. The whole genome resequencing indicates that the SNPs located in the odhA coding region may be associated with putrescine production. The comparative proteomic analysis reveals that the pentose phosphate and anaplerotic pathway, the glyoxylate cycle, and the ornithine biosynthetic pathway were upregulated in the evolved strain C. glutamicum PUT-ALE. The aspartate family, aromatic, and branched chain amino acid and fatty acid biosynthetic pathways were also observed to be downregulated in C. glutamicum PUT-ALE. Reducing OdhA activity by replacing the odhA native start codon GTG with TTG and overexpression of cgmA or pyc458 further improved putrescine production. Repressing the carB, ilvH, ilvB and aroE expression via CRISPRi also increased putrescine production by 5, 9, 16 and 19%, respectively.

Keywords

Putrescine Corynebacterium glutamicum Metabolic evolution Whole genome resequencing Proteome CRISPRi 
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Notes

Acknowledgements

This work was funded by the National Natural Science Foundation of China (Grant no. 21276289), the Natural Science Foundation of Guangdong Province (no. 2015A030311036), the Project of the Scientific and Technical Program of Guangdong Province (no. 2015A010107004) and the Project of the Scientific and Technical Program of Guangzhou (no. 201607010028).

Compliance with ethical standards

Conflict of interest

The authors declare no conflict of interests.

Supplementary material

10295_2018_2003_MOESM1_ESM.docx (178 kb)
Supplementary material 1 (DOCX 178 kb)
10295_2018_2003_MOESM2_ESM.xls (246 kb)
Supplementary material 2 (XLS 246 kb)

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Copyright information

© Society for Industrial Microbiology and Biotechnology 2018

Authors and Affiliations

  • Zhen Li
    • 1
  • Yu-Ping Shen
    • 1
  • Xuan-Long Jiang
    • 1
  • Li-Shen Feng
    • 1
  • Jian-Zhong Liu
    • 1
    • 2
  1. 1.Institute of Synthetic Biology, Biomedical Center, Guangdong Province Key Laboratory of Improved Variety Reproduction in Aquatic Economic Animals and South China Sea Bio-Resource Exploitation and Utilization Collaborative Innovation Center, School of Life SciencesSun Yat-sen UniversityGuangzhouPeople’s Republic of China
  2. 2.Biotechnology Research Centre, School of Life ScienceSun Yat-Sen UniversityGuangzhouPeople’s Republic of China

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