Skip to main content

Redirecting the lipid metabolism of the yeast Starmerella bombicola from glycolipid to fatty acid production

Abstract

Free fatty acids are basic oleochemicals implemented in a range of applications including surfactants, lubricants, paints, plastics, and cosmetics. Microbial fatty acid biosynthesis has gained much attention as it provides a sustainable alternative for petrol- and plant oil-derived chemicals. The yeast Starmerella bombicola is a microbial cell factory that naturally employs its powerful lipid metabolism for the production of the biodetergents sophorolipids (> 300 g/L). However, in this study we exploit the lipidic potential of S. bombicola and convert it from the glycolipid production platform into a free fatty acid cell factory. We used several metabolic engineering strategies to promote extracellular fatty acid accumulation which include blocking competing pathways (sophorolipid biosynthesis and β-oxidation) and preventing free fatty acid activation. The best producing mutant (Δcyp52m1Δfaa1Δmfe2) secreted 0.933 g/L (± 0.04) free fatty acids with a majority of C18:1 (43.8%) followed by C18:0 and C16:0 (40.0 and 13.2%, respectively). Interestingly, deletion of SbFaa1 in a strain still producing sophorolipids also resulted in 25% increased de novo sophorolipid synthesis (P = 0.0089) and when oil was supplemented to the same strain, a 50% increase in sophorolipid production was observed compared to the wild type (P = 0.03). We believe that our work is pivotal for the further development and exploration of S. bombicola as a platform for synthesis of environmentally friendly oleochemicals.

This is a preview of subscription content, access via your institution.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

References

  1. 1.

    Adrio JL (2017) Oleaginous yeasts: promising platforms for the production of oleochemicals and biofuels. Biotechnol Bioeng 114(9):1915–1920

    CAS  Article  Google Scholar 

  2. 2.

    Baccile N, Babonneau F, Banat IM, Ciesielska K, Cuvier A, Devreese B et al (2017) Development of a cradle-to-grave approach for acetylated acidic sophorolipid biosurfactants. ACS Sustain Chem Eng 5(1):1186–1198

    CAS  Article  Google Scholar 

  3. 3.

    Brakemeier A, Wullbrandt D, Lang S (1998) Candida bombicola: production of novel alkyl glycosides based on glucose/2-dodecanol. Appl Microbiol Biotechnol 50(2):161–166

    CAS  Article  Google Scholar 

  4. 4.

    Daniel HJ, Reuss M, Syldatk C (1998) Production of sophorolipids in high concentration from deproteinized whey and rapeseed oil in a two stage fed batch process using Candida bombicola ATCC 22214 and Cryptococcus curvatus ATCC 20509. Biotechnol Lett 20(12):1153–1156

    CAS  Article  Google Scholar 

  5. 5.

    Desbois AP, Smith VJ (2010) Antibacterial free fatty acids: activities, mechanisms of action and biotechnological potential. Appl Microbiol Biotechnol 85(6):1629–1642

    CAS  Article  Google Scholar 

  6. 6.

    De Weirdt R, Coenen E, Vlaeminck B, Fievez V, Van den Abbeele P, Van de Wiele T (2013) A simulated mucus layer protects Lactobacillus reuteri from the inhibitory effects of linoleic acid. Benef Microbes 4(4):299–312

    Article  Google Scholar 

  7. 7.

    Dulermo R, Gamboa-Meléndez H, Ledesma-Amaro R, Thevenieau F (1851) Nicaud JM (2015) Unraveling fatty acid transport and activation mechanisms in Yarrowia lipolytica. Biochim Biophys Acta 9:1202–1217

    Google Scholar 

  8. 8.

    Fernandez-Moya R, Da Silva NA (2017) Engineering Saccharomyces cerevisiae for high-level synthesis of fatty acids and derived products. FEMS Yeast Res 17(7):1–15

    Article  Google Scholar 

  9. 9.

    Fillet S, Gibert J, Suárez B, Lara A, Ronchel C, Adrio JL (2015) Fatty alcohols production by oleaginous yeast. J Ind Microbiol Biotechnol 42(11):1463–1472

    CAS  Article  Google Scholar 

  10. 10.

    Gao R, Falkeborg M, Xu X, Guo Z (2013) Production of sophorolipids with enhanced volumetric productivity by means of high cell density fermentation. Appl Microbiol Biotechnol 97:1103–1111

    CAS  Article  Google Scholar 

  11. 11.

    Jezierska S, Claus S, Van Bogaert I (2018) Yeast glycolipid biosurfactants. FEBS Lett 592:1312–1329

    CAS  Article  Google Scholar 

  12. 12.

    Johnson DR, Knoll LJ, Levin DE, Gordon JI (1994) Saccharomyces cerevisiae contains four fatty acid activation (FAA) genes: an assessment of their role in regulating protein N-myristoylation and cellular lipid metabolism. J Cell Biol 127(3):751–762

    CAS  Article  Google Scholar 

  13. 13.

    Johnson DR, Knoll LJ, Rowley N, Gordon JI (1994) Genetic analysis of the role of Saccharomyces cerevisiae Acyl-CoA synthetase genes in regulating protein N-myristoylation. J Biol Chem 269(27):18037–18046

    CAS  PubMed  Google Scholar 

  14. 14.

    Lang S, Brakemeier A, Heckmann R, Spöckner S, Rau U (2000) Production of native and modified sophorose lipids. Chim Oggi 18(10):76–79

    CAS  Google Scholar 

  15. 15.

    Ledesma-Amaro R (2015) Microbial oils: a customizable feedstock through metabolic engineering. Eur J Lipid Sci Technol 117(7):141–144

    CAS  Article  Google Scholar 

  16. 16.

    Ledesma-Amaro R, Dulermo R, Niehus X, Nicaud JM (2016) Combining metabolic engineering and process optimization to improve production and secretion of fatty acids. Metab Eng 38:38–46

    CAS  Article  Google Scholar 

  17. 17.

    Li X, Guo D, Cheng Y, Zhu F, Deng Z, Liu T (2014) Overproduction of fatty acids in engineered Saccharomyces cerevisiae. Biotechnol Bioeng 111(9):1841–1852

    CAS  Article  Google Scholar 

  18. 18.

    Pfleger BF, Gossing M, Nielsen J (2015) Metabolic engineering strategies for microbial synthesis of oleochemicals. Metab Eng 29:1–11

    CAS  Article  Google Scholar 

  19. 19.

    Ratledge C (2002) Regulation of lipid accumulation in oleaginous microorganisms. Biochem Soc Trans 30(6):47–50

    Article  Google Scholar 

  20. 20.

    Roelants S, Ciesielska K, De Maeseneire SL, Moens H, Everaert B, Verweire S et al (2016) Towards the industrialization of new biosurfactants: biotechnological opportunities for the lactone esterase gene from Starmerella bombicola. Biotechnol Bioeng 113(3):550–559

    CAS  Article  Google Scholar 

  21. 21.

    Runguphan W, Keasling JD (2014) Metabolic engineering of Saccharomyces cerevisiae for production of fatty acid-derived biofuels and chemicals. Metab Eng 21:103–113

    CAS  Article  Google Scholar 

  22. 22.

    Saerens KMJ, Saey L, Soetaert W (2011) One-step production of unacetylated sophorolipids by an acetyltransferase negative Candida bombicola. Biotechnol Bioeng 108(12):2923–2931

    CAS  Article  Google Scholar 

  23. 23.

    Scharnewski M, Pongdontri P, Mora G, Hoppert M, Fulda M (2008) Mutants of Saccharomyces cerevisiae deficient in acyl-CoA synthetases secrete fatty acids due to interrupted fatty acid recycling. FEBS J 275(11):2765–2778

    CAS  Article  Google Scholar 

  24. 24.

    Trotter PJ (2001) The genetics of fatty acid metabolism in Saccharomyces cerevisiae. Annu Rev Nutr 21:97–119

    CAS  Article  Google Scholar 

  25. 25.

    Tulloch AP, Spencer JFT, Deinema MH (1968) A new hydroxy fatty acid sophoroside from Candida bogoriensis. Can J Chem 46(3):345–348

    CAS  Article  Google Scholar 

  26. 26.

    Van Bogaert I, Develter D, Soetaert W, Vandamme EJ (2008) Cerulenin inhibits de novo sophorolipid synthesis of Candida bombicola. Biotechnol Lett 30(10):1829–1832

    Article  Google Scholar 

  27. 27.

    Van Bogaert I, Holvoet K, Roelants SLKW, Li B, Lin YC, Van de Peer Y et al (2013) The biosynthetic gene cluster for sophorolipids: a biotechnological interesting biosurfactant produced by Starmerella bombicola. Mol Microbiol 88(3):501–509

    Article  Google Scholar 

  28. 28.

    Van Bogaert I, De Maeseneire S, Develter D, Soetaert W, Vandamme EJ (2008) Cloning and characterisation of the glyceraldehyde 3-phosphate dehydrogenase gene of Candida bombicola and use of its promoter. J Ind Microbiol Biotechnol 35(10):1085–1092

    Article  Google Scholar 

  29. 29.

    Van Bogaert I, Sabirova J, Develter D, Soetaert W, Vandamme EJ (2009) Knocking out the MFE-2 gene of Candida bombicola leads to improved medium-chain sophorolipid production. FEMS Yeast Res 9(4):610–617

    Article  Google Scholar 

  30. 30.

    Van Bogaert I, Saerens K, De Muynck C, Develter D, Soetaert W, Vandamme EJ (2007) Microbial production and application of sophorolipids. Appl Microbiol Biotechnol 76(1):23–34

    Article  Google Scholar 

  31. 31.

    Wang J, Zhang B, Chen S (2011) Oleaginous yeast Yarrowia lipolytica mutants with a disrupted fatty acyl-CoA synthetase gene accumulate saturated fatty acid. Process Biochem 46(7):1436–1441

    CAS  Article  Google Scholar 

  32. 32.

    Yang L, Li Y, Zhang X, Liu T, Chen J, Wei L et al (2019) Metabolic profiling and flux distributions reveal a key role of acetyl-CoA in sophorolipid synthesis by Candida bombicola. Biochem Eng J 145:74–82

    CAS  Article  Google Scholar 

  33. 33.

    Zhang S, Ito M, Skerker JM, Arkin AP, Rao CV (2016) Metabolic engineering of the oleaginous yeast Rhodosporidium toruloides IFO0880 for lipid overproduction during high-density fermentation. Appl Microbiol Biotechnol 100(21):9393–9405

    CAS  Article  Google Scholar 

  34. 34.

    Zhou YJ, Buijs NA, Zhu Z, Qin J, Siewers V, Nielsen J (2016) Production of fatty acid-derived oleochemicals and biofuels by synthetic yeast cell factories. Nat Commun 7(May):11709

    CAS  Article  Google Scholar 

Download references

Acknowledgements

This work was funded by the Strategic Basic Research from the Research Foundation Flanders (FWO), (Sylwia Jezierska PhD Grant Number 151610). We would like to thank the Centre for Advanced Light Microscopy at Ghent University (Belgium) for the use of the fluorescence microscope during BodiPy staining experiments. We would also like to thank the Laboratory for Animal Nutrition and Animal Product Quality (Lanupro) at Ghent University for FID-MS analysis. The authors would like to acknowledge Dries Duchi for the excellent technical support during HPLC and UPLC analysis. The authors have no conflict of interest to declare.

Author information

Affiliations

Authors

Corresponding author

Correspondence to Inge Van Bogaert.

Additional information

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary material 1 (DOCX 15 kb)

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Jezierska, S., Claus, S., Ledesma-Amaro, R. et al. Redirecting the lipid metabolism of the yeast Starmerella bombicola from glycolipid to fatty acid production. J Ind Microbiol Biotechnol 46, 1697–1706 (2019). https://doi.org/10.1007/s10295-019-02234-x

Download citation

Keywords

  • Starmerella bombicola
  • Yeast
  • Free fatty acid
  • Lipid metabolism
  • Sophorolipid