Applied Biochemistry and Biotechnology

, Volume 184, Issue 4, pp 1308–1318 | Cite as

Efficient Malic Acid Production in Escherichia coli Using a Synthetic Scaffold Protein Complex

  • Sivachandiran Somasundaram
  • Gyeong Tae Eom
  • Soon Ho Hong
Article
  • 150 Downloads

Abstract

Recently, malic acid has gained attention due to its potential application in food, pharmaceutical, and medical industries. In this study, the synthetic scaffold complex strategy was employed between the two key enzymes pyruvate kinase (PykF) and malic enzyme (SfcA); SH3 ligand was attached to PykF, and the SH3 domain was attached to the C-terminus of ScfA. Synthetic scaffold systems can organize enzymes spatially and temporally to increase the local concentration of intermediates. In a flask culture, the recombinant strain harboring scaffold complex produced a maximum concentration of 5.72 g/L malic acid from 10 g/L glucose. The malic acid production was significantly increased 2.1-fold from the initial culture period. Finally, malic acid production was elevated to 30.2 g in a 5 L bioreactor from recombinant strain XL-1 blue.

Keywords

Malic acid Co-localization Scaffold complex Malic enzyme Pathway flux 

Notes

Acknowledgements

This work was supported by a grant from the Next-Generation BioGreen 21 Program (SSAC, grant number: PJ01111601), Rural Development Administration, Republic of Korea.

Compliance with Ethical Standards

Conflicts of Interests

The authors declare that they have no conflicts of interest.

References

  1. 1.
    Bressler, E., Pines, O., Goldberg, I., & Braun, S. (2002). Conversion of fumaric acid to L-malic by sol-gel immobilized Saccharomyces cerevisiae in a supported liquid membrane bioreactor. Biotechnology Progress, 18, 445–450.CrossRefGoogle Scholar
  2. 2.
    Chi, Z., Wang, Z. P., Wang, G. Y., Khan, I., & Chi, Z. M. (2016). Microbial biosynthesis and secretion of l-malic acid and its applications. Critical Reviews in Biotechnology, 36(1), 99–107.CrossRefGoogle Scholar
  3. 3.
    Dong, X., Chen, X., Yuanyuan, Q., Wang, Y., Wang, L., Qiao, W., & Liu, L. (2016). Metabolic engineering of Escherichia coli W3110 to produce L-malic acid. Biotechnology and Bioengineering, 114, 656–664.CrossRefGoogle Scholar
  4. 4.
    Duan, X., Chen, J., & Wu, J. (2013). Optimization of pullulanase production in Escherichia coli by regulation of process conditions and supplement with natural osmolytes. Bioresource Technology, 146, 379–385.CrossRefGoogle Scholar
  5. 5.
    Dueber, J. E., Wu, G. C., Malmirchegini, G. R., Moon, T. S., Petzold, C. J., Ullal, A. V., Prather, K. L. J., & Keasling, J. D. (2009). Synthetic protein scaffolds provide modular control over metabolic flux. Nature Biotechnology, 27, 753–759.CrossRefGoogle Scholar
  6. 6.
    Moon, S. Y., Hong, S. H., Kim, T. Y., & Lee, S. Y. (2008). Metabolic engineering of Escherichia coli for the production of malic acid. Biochemical Engineering Journal, 40, 312–320.CrossRefGoogle Scholar
  7. 7.
    Moon, T. S., Dueber, J. E., Shiue, E., & Prather, K. L. (2010). Use of modular, synthetic scaffolds for improved production of glucaric acid in engineered E. coli. Metabolic Engineering, 27, 298–305.CrossRefGoogle Scholar
  8. 8.
    Neubauer, A., Soini, J., Bollok, M., Zenker, M., Sandqvist, J., Myllyharju, J., & Neubauer, P. (2007). Fermentation process for tetrameric human collagen prolyl 4-hydroxylase in Escherichia coli: Improvement by gene optimisation of the PDI/β subunit and repeated addition of the inducer anhydrotetracycline. Journal of Biotechnology, 128(2), 308–321.CrossRefGoogle Scholar
  9. 9.
    Pham, V. D., Lee, S. H., Park, S. J., & Hong, S. H. (2015). Production of gamma-aminobutyric acid from glucose by introduction of synthetic scaffolds between isocitrate dehydrogenase, glutamatesynthase and glutamate decarboxylase in recombinant Escherichia coli. Journal of Biotechnology, 207, 52–57.CrossRefGoogle Scholar
  10. 10.
    Roa Engel, C. A., Straathof, A. J., Zijlmans, T. W., Van Gulik, W. M., & Van der Wielen, L. A. (2008). Fumaric acid production by fermentation. Applied Microbiology Biotechnology, 78, 379–389.CrossRefGoogle Scholar
  11. 11.
    Rosenberg, M., Mikova, H., & Kristofikova, L. (1999). Formation of L-malic acid by yeasts of the genus dipodascus. Letters in Applied Microbiology, 29, 221–223.CrossRefGoogle Scholar
  12. 12.
    Sambrook, J., & Russell, D. W. (2001). Molecular cloning: a laboratory manual, 3rd edn. New York: Cold Spring Harbor Laboratory Press.Google Scholar
  13. 13.
    Somasundaram, S., Tran, T. K. N., Ravikumar, S., & Hong, S. H. (2017). Introduction of synthetic protein complex between Pyrococcus horikoshii glutamate decarboxylase and Escherichia coli GABA transporter for the improved production of GABA. Biochemical Engineering Journal, 120, 1–6.CrossRefGoogle Scholar
  14. 14.
    Stols, L., & Donnelly, M. I. (1997). Production of succinic acid through overexpression of NAD1-dependent malic enzyme in an Escherichia coli mutant. Applied and Environmental Microbiology, 63, 2695–2701.Google Scholar
  15. 15.
    Wang, Y., & Yu, O. (2012). Synthetic scaffolds increased resveratrol biosynthesis in engineered yeast cells. Journal of Biotechnology, 157, 258–260.CrossRefGoogle Scholar
  16. 16.
    Werpy, T., & Petersen, G. (Eds.). (2004). Top value added chemicals from biomass. Volume I: results of screening for potential candidates from sugars and synthesis gas. Washington: US department of Energy, produced by Pacific Northwest National Laboratory (PNNL) and National Renewable Energy Laboratory (NREL).Google Scholar
  17. 17.
    Ye, X., Honda, K., Morimoto, Y., Okano, K., & Ohtake, H. (2013). Direct conversion of glucose to malic acid by synthetic metabolic engineering. Journal of Biotechnology, 164, 34–40.CrossRefGoogle Scholar
  18. 18.
    Zelle, R. M., De Hulster, E., Van Winden, W. A., De Waard, P., Dijkema, C., Winkler, A. A., Geertman, J. M., Van Dijken, J. P., Pronk, J. T., & Van Maris, A. J. (2008). Malic acid production by Saccharomyces cerevisiae: engineering of pyruvate carboxylation, oxaloacetate reduction, and malic acid export. Applied and Environmental Microbiology, 74(9), 2766–2777.CrossRefGoogle Scholar
  19. 19.
    Zhang, X., Wang, X., Shanmugam, K. T., & Ingram, L. O. (2011). L-malic acid production by metabolically engineered Escherichia coli. Applied and Environmental Microbiology, 77, 427–434.CrossRefGoogle Scholar
  20. 20.
    Zhao, A., Hu, X., Li, Y., Chen, C., & Wang, X. (2016). Extracellular expression of glutamate decarboxylase B in Escherichia coli to improve gamma-aminobutyric acid production. Applied and Industrial Microbiology and Biotechnology Express, 6, 55.Google Scholar

Copyright information

© Springer Science+Business Media, LLC 2017

Authors and Affiliations

  • Sivachandiran Somasundaram
    • 1
  • Gyeong Tae Eom
    • 2
    • 3
  • Soon Ho Hong
    • 1
  1. 1.Department of Chemical EngineeringUniversity of UlsanUlsanRepublic of Korea
  2. 2.Research Center for Bio-based ChemistryKorea Research Institute of Chemical Technology (KRICT)UlsanRepublic of Korea
  3. 3.Department of Green Chemistry and Environmental BiotechnologyKorea University of Science and Technology (UST)DaejeonRepublic of Korea

Personalised recommendations