Skip to main content
SpringerLink
Log in

Synthetic 2,3-Butanediol Pathway Integrated Using Tn7-tool and Powered Via Elimination of Sporulation and Acetate Production in Acetogen Biocatalyst

  • Published:
Applied Biochemistry and Biotechnology Aims and scope Submit manuscript

Abstract

Acetogen Clostridium sp. MT1802 originally producing 336-mM acetate from inorganic carbon of CO2/CO was engineered to eliminate acetate production and sporulation using Cre-lox66/lox71-approach. The recombinant started producing 105-mM formate expressing synthetic formate dehydrogenase integrated in two copies. Formate-producing recombinant was further engineered to express synthetic formate acetyltransferase, acetolactate synthase, acetolactate decarboxylase, and alcohol dehydrogenase integrated in two copies each using Tn7 tool. The resulted recombinant started producing 102-mM 2,3-butanediol (23BD). 23BD production was confirmed in five independent single step fermentation runs 25 days long each in five repeats using syngas blend 60 % CO and 40 % H2 (v/v) (p <0.005). 23BD production was 78 % if only CO2/H2 blend was fed instead of syngas (p <0.005). 23BD from CO2/H2 blend might serve as a commercial route to mitigate global warming in proportion to CO2 fermentation scale worldwide.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8
Fig. 9
Fig. 10
Fig. 11

Similar content being viewed by others

References

  1. Young, K. D. (2006). The selective value of bacterial shape. Microbiology and Molecular Biology Reviews, 70(3), 660–703. doi:10.1128/MMBR.00001-06.

    Article  Google Scholar 

  2. Berzin, V., Kiriukhin, M., & Tyurin, M. (2012). Selective production of acetone during continuous synthesis gas fermentation by engineered biocatalyst Clostridium sp. MAceT113. Letters in Applied Microbiology, 55, 149–154. doi:10.1111/j.1472-765X.2012.03272.x.

    Article  CAS  Google Scholar 

  3. Tyurin, M., & Kiriukhin, M. (2013). Ethanol overproduction during continuous syngas fermentation due to expression of amplified ethanol biosynthesis pathway integrated using Tn7-tool and powered at the expense of eliminated acetic acid production and spore formation. Journal of Applied Microbiology. doi:10.1111/jam.12123.

    Google Scholar 

  4. Demain, A. L. (2009). Biosolutions to the energy problem. Journal of Industrial Microbiology and Biotechnology, 36(3), 319–332.

    Article  CAS  Google Scholar 

  5. Fischer, C. R., Klein-Marcuschamer, D., & Stephanopoulos, G. (2008). Selection and optimization of microbial hosts for biofuels production. Metabolic Engieneering, 10(6), 295–304.

    Article  CAS  Google Scholar 

  6. Köpke, M., Michalcea, C., Liew, F.-M., Tizard, J. H., Ali, M. S., Conolly, J. J., et al. (2011). 2, 3-butanediol production by acetogenic bacteria, an alternative route to chemical synthesis, using industrial waste gas. Applied and Environmental Microbiology, 77(15), 5467–5475.

    Article  Google Scholar 

  7. Abubackar, H. N., Veiga, M. C., & Kennes, C. (2012). Biological conversion of carbon monoxide to ethanol: effect of pH, gas pressure, reducing agent and yeast extract. Bioresource Technology, 114, 518–522.

    Article  CAS  Google Scholar 

  8. Fischer, C. R., Klein-Marcuschamer, D., & Stephanopoulos, G. (2008). Selection and optimization of microbial hosts for biofuels production. Metabolic Engineering, 10(6), 295–304.

    Article  CAS  Google Scholar 

  9. Gaddy, J. L., Arora, D. D., Ko, C.-W., Phillips, J. R., Basu, R., Wikstrom, C. V., Clausen, E. C. (2001). Methods for increasing the production of ethanol from microbial fermentation. US Patent 7285402.

  10. Ma, C., Wang, A., Qin, J., Li, L., Ai, X., Jiang, T., et al. (2009). Enhanced 2,3-butanediol production by Klebsiella pneumoniae SDM. Applied Microbiology and Biotechnology, 82(1), 49–57. doi:10.1007/s00253-008-1732-7.

    Article  CAS  Google Scholar 

  11. Celińska, E., & Grayek, W. (2009). Biotechnological production of 2,3-butanediol—current state and prospects. Biotechnology Advances, 27, 715–725.

    Article  Google Scholar 

  12. Xiu, Z. L., & Zeng, A. P. (2008). Present state and perspectives of downstream processing of biologically produced 1,3-propanediol and 2,3-butanediol. Applied Microbiology and Biotechnology, 78, 917–926.

    Article  CAS  Google Scholar 

  13. Ji, X. J., Huang, H., & Ouyang, P. K. (2011). Microbial 2,3-butanediol production: a state-of-the-art review. Biotechnology Advances, 29, 351–364.

    Article  CAS  Google Scholar 

  14. Oliver, J. W., Machado, I. M., Yoneda, H., & Atsumi, S. (2013). Cyanobacterial conversion of carbon dioxide to 2,3-butanediol. Proceedings of the National Academy of Sciences U S A, 110(4), 1249–1254. doi:10.1073/pnas.1213024110.

    Article  CAS  Google Scholar 

  15. Drake, H. L. (1994). Acetogenesis. NY, London: Chapman & Hall.

    Google Scholar 

  16. Berzin, V., Kiriukhin, M., & Tyurin, M. (2012). Cre-lox66/lox71-based elimination of phosphotransacetylase or acetaldehyde dehydrogenase shifted carbon flux in acetogen rendering selective overproduction of ethanol or acetate. Applied Biochemistry and Biotechnology. doi:10.1007/s12010-012-9864-8.

    Google Scholar 

  17. Craig, N. L. (1991). Micro Review. Tn7: a target site-specific transposon. Molecular Microbiology, 5(11), 2569–2573.

    Article  CAS  Google Scholar 

  18. Tanner, R. S., Miller, L. M., & Yang, D. (1993). Clostridium ljungdahlii sp. nov., an acetogenic species in clostridial rRNA homology group I. International Journal of Systemic Bacteriology, 43(2), 232–236.

    Article  CAS  Google Scholar 

  19. Tyurin, M., Lynd, L. R., & Wiegel, J. (2006). Genetic delivery systems for strictly anaerobic thermophiles. In F. A. Rainey & A. Oren (Eds.), Methods in Microbiology (1st ed., Vol. 35, pp. 307–328). Amsterdam: Elsevier, Academic Press.

    Google Scholar 

  20. Berzin, V., Kiriukhin, M., & Tyurin, M. (2013). Selective n-butanol production by Clostridium sp. MTButOH1365 during continuous synthesis gas fermentation due to expression of synthetic thiolase, 3-hydroxy butyryl-CoA dehydrogenase, crotonase, butyryl-CoA dehydrogenase, butyraldehyde dehydrogenase and NAD-dependent butanol dehydrogenase. Applied Biochemistry and Biotechnology. doi:10.1007/s12010-012-0060-7.

    Google Scholar 

  21. Tyurin, MV. (1992). The method and apparatus for electric cell treatment using partial power capacitor discharge via the sample and the power tetrode. Russian Patent 2005776.

  22. Tyurin, M., Desai, S., & Lynd, L. (2004). Electrotransformation of Clostridium thermocellum. Applied and Environmental Microbiology, 70(2), 883–890.

    Article  CAS  Google Scholar 

  23. Tyurin, M. V., Sullivan, C. R., & Lynd, L. R. (2005). Induced oscillations play an important role in high efficiency transformation of thermophilic anaerobes. Applied and Environmental Microbiology, 71, 8069–8076.

    Article  CAS  Google Scholar 

  24. Tyurin, M., Kiriukhin, M., & Berzin, V. (2012). Electrofusion of cells of acetogen Clostridium sp. MT351 with erm(B) or cat in the chromosome. Journal of Biotech Research, 4, 1–12.

    CAS  Google Scholar 

  25. Berzin, V., & Tyurin, M. (2012). Acetogen biocatalyst Clostridium sp. MTEtOH871 engineered with our proprietary electrotransformation technology and equipment: continuous synthesis gas fermentation for selective ethanol production. Journal of Biotech Research, 4, 54–64.

    CAS  Google Scholar 

  26. Leibig, M., Krismer, B., Kolb, M., Friede, A., Götz, F., & Bertram, R. (2008). Marker removal in staphylococci via Cre recombinase and different lox sites. Applied and Environmental Microbiology, 74(5), 1316–1323.

    Article  CAS  Google Scholar 

  27. Shepherd, C. T., Moran, L. A. N., & Scott, M. P. (2009). Determination of transgene copy number by real-time quantitative PCR. Methods in Molecular Biology, 526, 129–134. doi:10.1007/978-1-59745-494-0_11.

    Article  CAS  Google Scholar 

  28. Ross, S. M. (2000). Introduction to probability and statistics for engineers and scientists (2nd ed.). San Diego: Academic.

    Google Scholar 

  29. Volker, M. (2003). Energy conservation in acetogenic bacteria. Applied and Environmental Microbiology, 69(11), 6345–6353. doi:10.1128/AEM.69.11.6345-6353.2003.

    Article  Google Scholar 

  30. Wang, A., Xu, Y., Ma, C., Gao, C., Li, L., Wang, Y., et al. (2012). Efficient 2,3-Butanediol Production from cassava powder by a crop-biomass-utilizer, Enterobacter cloacae subsp. dissolvens SDM. PLoS One, 7(7), e40442. doi:10.1371/journal.pone.0040442.

    Article  CAS  Google Scholar 

Download references

Acknowledgments

The research was supported by the funds of angel friends of MT family. Syngas Biofuels Energy, Inc. and its successors are the sole distributors of the electroporation and electrofusion equipment: www.syngasbiofuelsenergy.com.

Conflict of Interest

The authors declare that they have no conflict of interest.

Author information

Authors and Affiliations

  1. Syngas Biofuels Energy, Inc., P.O. Box 300819, Houston, TX, 77230, USA

    Michael Tyurin

  2. Ajinomoto-Genetika Research Institute, 1st Dorozhny Pr. 1-1, Moscow, Russia, 117545

    Michael Kiriukhin

Authors
  1. Michael Tyurin
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Michael Kiriukhin
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Michael Tyurin.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Tyurin, M., Kiriukhin, M. Synthetic 2,3-Butanediol Pathway Integrated Using Tn7-tool and Powered Via Elimination of Sporulation and Acetate Production in Acetogen Biocatalyst. Appl Biochem Biotechnol 170, 1503–1524 (2013). https://doi.org/10.1007/s12010-013-0285-0

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12010-013-0285-0

Keywords

Search

Navigation