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Synthetic cell–cell communication in a three-species consortium for one-step vitamin C fermentation

  • En-Xu Wang
  • Yu Liu
  • Qian Ma
  • Xiu-Tao Dong
  • Ming-Zhu DingEmail author
  • Ying-Jin Yuan
Original Research Paper
  • 39 Downloads

Abstract

Objectives

A three-species consortium for one-step fermentation of 2-keto-l-gulonic acid (2-KGA) was constructed to better strengthen the cell–cell communication. And the programmed cell death module based on the LuxI/LuxR quorum-sensing (QS) system was established in Gluconobacter oxydans to reduce the competition that between G. oxydans and Ketogulonicigenium vulgare.

Results

By constructing and optimizing the core region of the promoter, which directly regulated the expression of lethal ccdB genes in QS system, IR3C achieved the best lethal effect. The consortium of IR3C- K. vulgareBacillus megaterium (abbreviated as 3C) achieved the highest 2-KGA titer (68.80 ± 4.18 g/l), and the molar conversion rate was 80.7% within 36 h in 5 l fermenter. Metabolomic analysis on intracellular small molecules of consortia 3C and 1C showed that most amino acids (such as glycine, leucine, methionine and proline) and TCA cycle intermediates (such as succinic acid, fumaric acid and malic acid) were significantly affected. These results further validated that the programmed cell death module based on the LuxI/LuxR QS system in G. oxydans could also faciliate better growth and higher production of consortium 3C for one-step fermentation.

Conclusions

We successfully constructed a novel three-species consortia for one-step vitamin C fermentation by strengthening the cell–cell communication. This will be very useful for probing the rational design principles of more complex multi-microbial consortia.

Keywords

One-step fermentation Programmed cell death Quorum-sensing 2-keto-l-gulonic acid 

Notes

Acknowledgements

This work was funded by the National Key Research and Development Program of China (Grant Nos. 2018YFA0902103, 2018YFA0902200), National Natural Science Foundation of China (Grant Nos. 21676190, 21621004), Innovative Talents and Platform Program of Tianjin (Grant No. 16PTGCCX00140).

Supporting information

Supplementary Table 1— Primers for the synthesis of IR1C

Supplementary Table 2— Primers for qPCR reaction

Supplementary Fig. 1— Flow chart of the construction of the gene circuit

Supplementary Fig. 2—Construction of a synthetic microbial consortium of G. oxydans- K. vulgare- Bacillus megaterium for cell-cell communication in one-step vitamin C precursor fermentation

Supplementary Fig. 3—Relationship between OD600 and dry cell weight (DCW) of G. oxydans

Additional file 1—IR1C is split into full sequence of 4 Building Blocks

Supplementary material

10529_2019_2705_MOESM1_ESM.docx (340 kb)
Supplementary material 1 (DOCX 339 kb)

References

  1. Bassler BL, Losick R (2006) Bacterially Speaking. Cell 125:237–246CrossRefGoogle Scholar
  2. Brenner K, You L, Arnold FH (2008) Engineering microbial consortia: a new frontier in synthetic biology. Trends Biotechnol 26:483–489CrossRefGoogle Scholar
  3. Chambers ST, Kunin CM (1987) Isolation of glycine betaine and proline betaine from human urine. Assessment of their role as osmoprotective agents for bacteria and the kidney. J Clin Investig 79:731–737CrossRefGoogle Scholar
  4. Daothi MH, Van LM, De EG, Afif H, Buts L, Wyns L, Loris R (2005) Molecular basis of gyrase poisoning by the addiction toxin CcdB. J Mol Biol 348:1091–1102CrossRefGoogle Scholar
  5. Ding MZ, Zhou X, Yuan YJ (2010) Metabolome profiling reveals adaptive evolution of Saccharomyces cerevisiae during repeated vacuum fermentations. Metabolomics 6:42–55CrossRefGoogle Scholar
  6. Hanzelka BL, Greenberg EP (1996) Quorum sensing in Vibrio fischeri: evidence that S-adenosylmethionine is the amino acid substrate for autoinducer synthesis. J Bacteriol 178:5291CrossRefGoogle Scholar
  7. Jia N, Ding MZ, Du J, Pan CH, Tian G, Lang JD, Fang JH, Gao F, Yuan YJ (2016) Insights into mutualism mechanism and versatile metabolism of Ketogulonicigenium vulgare Hbe602 based on comparative genomics and metabolomics studies. Sci Rep 6:23068CrossRefGoogle Scholar
  8. Jones JA, Wang X (2017) Use of bacterial co-cultures for the efficient production of chemicals. Curr Opin Biotechnol 53:33–38CrossRefGoogle Scholar
  9. Kong W, Meldgin DR, Collins JJ, Lu T (2018) Designing microbial consortia with defined social interactions. Nat Chem Biol 14:821–829CrossRefGoogle Scholar
  10. Li MJ, Wang JS, Geng YP, Li YK, Wang Q, Liang QF, Qi QS (2012) A strategy of gene overexpression based on tandem repetitive promoters in Escherichia coli. Microb Cell Fact 11:19CrossRefGoogle Scholar
  11. Mee MT, Collins JJ, Church GM, Wang HH (2014) Syntrophic exchange in synthetic microbial communities. Proc Natl Acad Sci 111:E2149–E2156CrossRefGoogle Scholar
  12. Pandhal J, Noirel J (2014) Synthetic microbial ecosystems for biotechnology. Biotechnol Lett 36:1141–1151CrossRefGoogle Scholar
  13. Scott SR, Hasty J (2016) Quorum sensing communication modules for microbial consortia. ACS Synthetic Biology 5:969CrossRefGoogle Scholar
  14. Simic M, De JN, Loris R, Vesnaver G, Lah J (2009) Driving forces of gyrase recognition by the addiction toxin CcdB. J Biol Chem 284:20002–20010CrossRefGoogle Scholar
  15. Song H, Ding MZ, Jia XQ, Ma Q, Yuan YJ (2014) Synthetic microbial consortia: from systematic analysis to construction and applications. Chem Soc Rev 43:6954–6981CrossRefGoogle Scholar
  16. Whitehead NA, Barnard AML, Slater H, Simpson NJL, Salmond GPC (2001) Quorum-sensing in gram-negative bacteria. FEMS Microbiol Rev 25:365–404CrossRefGoogle Scholar
  17. Xia J, Wishart DS (2011) Web-based inference of biological patterns, functions and pathways from metabolomic data using MetaboAnalyst. Nat Protoc 6:743–760CrossRefGoogle Scholar
  18. Yang J, Moyana T, Mackenzie S, Xia Q, Xiang J (1998) One hundred seventy-fold increase in excretion of an FV fragment-tumor necrosis factor alpha fusion protein (sFV/TNF-alpha) from Escherichia coli caused by the synergistic effects of glycine and triton X-100. Appl Environ Microbiol 64:2869–2874Google Scholar
  19. You L, Iii RSC, Weiss R, Arnold FH (2004) Programmed population control by cell-cell communication and regulated killing. Nature 428:868CrossRefGoogle Scholar
  20. Zhu Y, Liu J, Du G, Zhou J, Chen J (2012) Sporulation and spore stability of Bacillus megaterium enhance Ketogulonigenium vulgare propagation and 2-keto-L-gulonic acid biosynthesis. Bioresour Technol 107:399–404CrossRefGoogle Scholar
  21. Zou Y, Hu M, Lv Y, Wang Y, Song H, Yuan YJ (2013) Enhancement of 2-keto-gulonic acid yield by serial subcultivation of co-cultures of Bacillus cereus and Ketogulonicigenium vulgare. Bioresour Technol 132:370–373CrossRefGoogle Scholar

Copyright information

© Springer Nature B.V. 2019

Authors and Affiliations

  • En-Xu Wang
    • 1
    • 2
  • Yu Liu
    • 1
    • 2
  • Qian Ma
    • 1
    • 2
  • Xiu-Tao Dong
    • 1
    • 2
  • Ming-Zhu Ding
    • 1
    • 2
    Email author
  • Ying-Jin Yuan
    • 1
    • 2
  1. 1.Frontier Science Center for Synthetic Biology and Key Laboratory of Systems Bioengineering (Ministry of Education), School of Chemical Engineering and TechnologyTianjin UniversityTianjinPeople’s Republic of China
  2. 2.Collaborative Innovation Center of Chemical Science and Engineering (Tianjin)Tianjin UniversityTianjinPeople’s Republic of China

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