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Improvement of levan production in Bacillus amyloliquefaciens through metabolic optimization of regulatory elements

Applied Microbiology and Biotechnology volume 101, pages 4163–4174 (2017)Cite this article

Abstract

Levan is a functional homopolymer of fructose with considerable applications in food, pharmaceutical and cosmetic industries. To improve the levan production in Bacillus amyloliquefaciens, the regulatory elements of sacB (encoding levansucrase) expression and levansucrase secretion were optimized. Four heterologous promoters were evaluated for sacB expression, and the Pgrac promoter led to the highest level for both sacB transcription and levansucrase enzyme activity. The levan production in the corresponding recombinant strain ΔLP-pHTPgrac reached 30.5 g/L, which was 114% higher than that of the control strain NK-ΔLP. In a further step, eight signal peptides were investigated (with Pgrac as the promoter for sacB expression) for their effects on the levansucrase secretion and levan production. The signal peptide yncM was identified as the optimal one, with a secretion efficiency of approximately 90%, and the levan production in the corresponding recombinant strain ΔLP-Y reached 37.4 g/L, which was 161% higher when compared with the control strains NK-ΔLP. Finally, fed-batch fermentation was carried out in 5-L bioreactors for levan production using the recombinant strain ΔLP-Y. A final levan concentration of 102 g/L was achieved, which is very close to the ever reported highest levan production level from the literature. To our best knowledge, this is the first report of the improvement of levan production through metabolic optimization for sacB expression and levansucrase secretion. The results from this study provided essential insights for systematically metabolic engineering of microbial cell factories for enhanced biochemical production.

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Acknowledgments

This work was supported by the National Key Technology Support Program 2015BAD16B04; the Natural Science Foundation of China Grant Nos. 81671842, 31470213 and 31670093; and the Project of Tianjin, China, i.e., 16JCZDJC37600, 15ZCZDNC00450 and 14ZCZDSF00009; and the program of State Key Laboratory of Medicinal Chemical Biology 201603007.

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Affiliations

  1. Key Laboratory of Molecular Microbiology and Technology for Ministry of Education, Nankai University, Tianjin, 300071, China

    Yanyan Gu, Jiayi Zheng, Jun Feng, Weixia Gao, Yufen Quan, Yulei Dang & Cunjiang Song

  2. Biosystems Engineering Department, Auburn University, Auburn, AL, 36849, USA

    Yanyan Gu, Jun Feng & Yi Wang

  3. State Key Laboratory of Medicinal Chemical Biology, Nankai University, 94 Weijin Road, Tianjin, 300071, China

    Jun Feng & Shufang Wang

  4. Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi’an Jiaotong University, Xi’an, Shaanxi, 710049, China

    Jun Feng

  5. Department of Chemical and Biological Engineering, Iowa State University, Ames, IA, 50011, USA

    Mingfeng Cao

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  1. Yanyan Gu
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  2. Jiayi Zheng
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  3. Jun Feng
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  4. Mingfeng Cao
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  6. Yufen Quan
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  7. Yulei Dang
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  8. Yi Wang
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  9. Shufang Wang
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  10. Cunjiang Song
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Correspondence to Shufang Wang or Cunjiang Song.

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Gu, Y., Zheng, J., Feng, J. et al. Improvement of levan production in Bacillus amyloliquefaciens through metabolic optimization of regulatory elements. Appl Microbiol Biotechnol 101, 4163–4174 (2017). https://doi.org/10.1007/s00253-017-8171-2

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  • DOI: https://doi.org/10.1007/s00253-017-8171-2

Keywords

  • Levan
  • Levansucrase
  • Bacillus amyloliquefaciens
  • Promoter
  • Signal peptide