Skip to main content
Log in

Characterization of expression of a plant oleate desaturase in yeast

  • Published:
Journal of the American Oil Chemists' Society

Abstract

We have expressed the oleate desaturase gene of Arabidopsis thaliana in Saccharomyces cerevisiae by means of a galactose-inducible promoter. Transfer of yeast cells to galactose-containing media and growth at 30°C resulted in the production of active oleate desaturase, as shown by the appearance of the diunsaturated octadecadienoic acid (18:2) and hexadecadienoic acid (16:2). The highest level of these two fatty acids was 46.2%. In induced cultures, 18:2 appeared before 16:2, and we show that both of these fatty acids were present first on phosphatidylcholine and then appeared in other phospholipids and a neutral lipid fraction. We raised antibodies against the C-terminal 100 amino acids of oleate desaturase and demonstrated that this protein was in the microsomal fraction of induced cells.

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

Access this article

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

Instant access to the full article PDF.

Similar content being viewed by others

References

  1. Murphy, D.J., Biogenesis, Function and Biotechnology of Plant Storage Lipids, Prog. Lipid Res. 33:71–85 (1994).

    Article  CAS  Google Scholar 

  2. Ohlrogge, J., and J. Browse, Lipid Biosynthesis, Plant Cell 7:957–970 (1995).

    Article  CAS  Google Scholar 

  3. Somerville, C., and J. Browse, Dissecting Desaturation—Plants Prove Advantageous, Trends in Cell Biol. 6:148–153 (1995).

    Article  Google Scholar 

  4. Lindqvist, Y., W. Huang, G. Scneider, and J. Shanklin, Crystal Structure of Delta-9 Stearoyl-Acyl Carrier Protein Desaturase from Castor Seed and Its Relationship to Other Di-Iron Proteins, EMBO J. 16:4081–4092 (1995).

    Google Scholar 

  5. Stobart, A.K., and S. Stymne, The Regulation of the Fatty Acid Composition of the Triacylglycerols in Microsomal Preparations from Avocado Mesocarp and the Developing Cotyledons of Safflower, Planta 163:119–125 (1985).

    Article  CAS  Google Scholar 

  6. Smith, M.A., A.R. Cross, O.T.G. Jones, W.T. Griffiths, S. Stymne, and K. Stobart, Electron-Transport Components of the 1-Acyl-2-Oleoyl sn-Glycero-3-Phosphocholine Delta-12 Desaturase in Microsomal Preparations from Developing Safflower (Carthamus tinctorius) Cotyledons, Biochem. J. 272:23–29 (1990).

    CAS  Google Scholar 

  7. Kearns, E.V., S. Hugly, and C.R. Somerville, The Role of Cytochrome b5 in Delta-12 Desaturation of Oleic Acid by Microsomes of Safflower (Carthamus tinctorius), Arch. Biochem. Biophys. 284:431–436 (1991).

    Article  CAS  Google Scholar 

  8. Okuley, J., J. Lightner, K. Feldmann, N. Yadav, E. Lark, and J. Browse, Arabidopsis FAD2 Gene Encodes the Enzyme That is Essential for Polyunsaturated Lipid Synthesis, Plant Cell 6:147–158 (1994).

    Article  CAS  Google Scholar 

  9. Shanklin, J., E. Whittle, and B.G. Fox, Eight Histidine Residues are Catalytically Essential in a Membrane-Associated Iron Enzyme, Stearoyl-CoA Desaturase and are Conserved in Alkane Hydroxylase and Xylene Monooxygenase, Biochemistry 33:12787–12794 (1994).

    Article  CAS  Google Scholar 

  10. Avelange-Macherel, M.-H., D. Macherel, H. Wada, and N. Murata, Site Directed Mutagenesis of Histidine Residues in the Delta-12 Acyl Lipid Desaturase of Synechocystis, FEBS Lett. 361:111–114 (1995).

    Article  CAS  Google Scholar 

  11. Bossie, M.A., and C.E. Martin, Nutritional Regulation of Yeast Delta-9 Fatty Acid Desaturase Activity, J. Bacteriol. 171:6409–6413 (1989).

    CAS  Google Scholar 

  12. Covello, P.S., and D.W. Reed, Functional Expression of the Extraplastidial Arabidopsis thaliana Oleate Desaturase Gene (Fad2) in Saccharomyces cerevisiae, Plant Physiol. 111:223–226 (1996).

    Article  CAS  Google Scholar 

  13. Ito, H., Y. Fukuda, K. Murata, and A. Kimura, Transformation of Intact Yeast Cells Treated with Alkali Cations, J. Bacteriol. 153:163–168 (1983).

    CAS  Google Scholar 

  14. Harlow, E., and D. Lane, Antibodies: A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, 1988.

    Google Scholar 

  15. Ausubel, F.M., R. Brent, R.E. Kingston, D.D. Moore, J.G. Seidman, J.A. Smith, K. Struhl, L.M. Albright, D.M. Coen, and A. Varki, Current Protocols in Molecular Biology Unit 13.13, John Wiley & Sons, New York, 1993.

    Google Scholar 

  16. Stukey, J.E., V.M. McDonough, and C.E. Martin, Isolation and Characterization of Olel, a Gene Affecting Fatty Acid Desaturation from Saccharomyces cerevisiae. J. Biol. Chem. 264:16537–16544 (1989).

    CAS  Google Scholar 

  17. McDonough, V.M., J.E. Stukey, and C.E. Martin, Specificity of Unsaturated Fatty Acid Regulated Expression of the Saccharomyces cerevisiae Olel Gene, J. Biol. Chem. 267:5931–5936 (1992).

    CAS  Google Scholar 

  18. Wagner, S., and F. Paltauf, Generation of Glycerophospholipid Molecular Species in the Yeast Saccharmyces cerevisiae Fatty Acid Pattern of Phospholipid Classes and Selective Acyl Turnover at sn-1 and sn-2 Positions, Yeast 10:1429–1437 (1994).

    Article  CAS  Google Scholar 

  19. Sperling, P., M. Linscheid, S. Stöcker, H.-P. Mühlbach, and E. Heinz, In Vivo Desaturation of cis Delta-9 Monounsaturated to cis Delta-9,12 Diunsaturated Alkenylether Glycerolipids, J. Biol. Chem. 268:26935–26940 (1993).

    CAS  Google Scholar 

  20. Gennity, J.M., and P.K. Stumpf, Studies of the Delta-12 Desaturase of Carthamus-tinctorius, Arch. Biochem. Biophys. 239:444–454 (1985).

    Article  CAS  Google Scholar 

  21. Lightner, J.E., and J.J. Okuley, Genes for Microsomal Delta-12 Fatty Acid Desaturases and Related Enzymes from Plants, Patent WO 94/11516 (1994).

  22. Piffanelli, P., Molecular and Biochemical Studies of Stearoyl-ACP Desaturase in Gametophytic and Sporophytic Generations of Plants, Ph.D. University of East Anglia, United Kingdom, 1997.

    Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Matthew J. Hills.

About this article

Cite this article

Brown, A.P., Dann, R., Bowra, S. et al. Characterization of expression of a plant oleate desaturase in yeast. J Amer Oil Chem Soc 75, 77–79 (1998). https://doi.org/10.1007/s11746-998-0014-y

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11746-998-0014-y

Key words

Navigation