Skip to main content
SpringerLink
Log in

Engineering pathways for malate degradation in Saccharomyces cerevisiae

  • Research Article
  • Published:

From Nature Biotechnology

View current issue Submit your manuscript

Abstract

Deacidification of grape musts is crucial for the production of well-balanced wines, especially in colder regions of the world. The major acids in wine are tartaric and malic acid. Saccharomyces cerevisiae cannot degrade malic acid efficiently due to the lack of a malate transporter and the low substrate affinity of its malic enzyme. We have introduced efficient pathways for malate degradation in S. cerevisiae by cloning and expressing the Schizosaccharomyces pombe malate permease (mae1) gene with either the S. pombe malic enzyme (mae2) or Lactococcus lactis malolactic (mleS) gene in this yeast. Under aerobic conditions, the recombinant strain expressing the mae1 and mae2 genes efficiently degraded 8 g/L of malate in a glycerol-ethanol medium within 7 days. The recombinant malolactic strain of S. cerevisiae (mael and mleS genes) fermented 4.5 g/L of malate in a synthetic grape must within 4 days.

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

Similar content being viewed by others

References

  1. Beelman, R.B. and Gallander, J.F. 1979. Wine deacidification. Adv. Food Res. 25: 1–53.

    Article  CAS  Google Scholar 

  2. van Vuuren, H.J.J. and Dicks, L.M.T. Leuconostoc oenos: A review. Am. J. Enol. Vitic. 44: 99–112.

  3. Davies, C.R., Wibowo, W., Eschenbruch, R., Lee, T.H., and Fleet, G.H. 1985. Practical implications of malolactic fermentation: A review. Am. J. Enol. Vitic. 36: 290–301.

    Google Scholar 

  4. Henick-Kling, T. 1995. Control of malolactic fermentation in wine: energetics, flavor modification and methods of starter culture preparation. J. Appl. Bacteriol. (Symp. Suppl.) 79: 29S–37S.

    Google Scholar 

  5. Gallander, J.F. 1977.Deacidification of Eastern table wines with Schizosaccharomyces pombe . Am. J. Enol. Vitic. 28: 65–68.

    CAS  Google Scholar 

  6. Radler, F. 1993. Yeast metabolism of organic acids pp. 165–182 in Wine Microbiology and Biotechnology. Fleet (ed.), Harwood Academic Publishers, Switzerland.

    Google Scholar 

  7. Gao, C. and Fleet, G.H. 1995. Degradation of malic and tartaric acids by high density cell suspensions of wine yeasts. Food Microbiol. 12: 65–71.

    Article  CAS  Google Scholar 

  8. Mayer, K. and Temperli, A. 1963. The metabolism of L-malate and other compounds by Schizosaccharomyces pombe . Arch. Mikrobiol. 46: 321–328.

    Article  CAS  Google Scholar 

  9. Fuck, E., Stark, G., and Radler, F. 1973. Äpfelsäurestoff-wechsel bei SaccharomycesII. Anreicherung und Eigenschaften eines Malatenzymes. Arch. Mikrobiol. 89: 223–231.

    Article  CAS  PubMed  Google Scholar 

  10. Temperli, A., Kunsch, V., Mayer, K., and Busch, I. 1965. Reinigung und Eigenschaften der Malatdehydrogenase (decarboxylierent) aus Hefe. Biochem. Biophys. Acta. 110: 630–632.

    CAS  PubMed  Google Scholar 

  11. Ansanay, V., Dequin, S., Camarasa, C., Schaeffer, V., Grivet, J., Blondin, B., Salmon, J., and Barre, P. 1996. Malolactic fermentation by engineered Saccharomyces cerevisiaeas compared with engineered Schizosaccharomyces pombe . Yeast 12: 215–225.

    Article  CAS  PubMed  Google Scholar 

  12. Ansanay, V., Dequin, S., Blondin, B., and Barre, P., 1993. loning, sequence and expression of the gene encoding the malolactic enzyme from Lactococcus lactis . FEBS Lett. 332: 74–80.

    Article  CAS  PubMed  Google Scholar 

  13. Denayrolles, M., Aigle, M., and Lonvaud-Funel, A. 1995. Functional expression in Saccharomyces cerevisiaeof the Lactococcus lactis mleSgene encoding the malolactic enzyme. FEMS Microbiol. Lett. 125: 3744.

    Article  Google Scholar 

  14. Williams, S.A., Hodges, R.A., Strike, T.L., Snow, R., and Kunkee, R.E. 1984. Cloning the gene for the malolactic fermentation of wine from Lactobacillus delbrueckiiin Escherichia coliand yeasts. Appl. Environ. Microbiol. 47: 288–293.

    CAS  PubMed  PubMed Central  Google Scholar 

  15. Grobler, J., Bauer, F., Subden, R.E., and van Vuuren, H.J.J. . 1995. The maelgene of Schizosaccharomyces pombeencodes a permease for malate and other C4 dicarboxylic acids. Veasf 11: 1485–1491.

    CAS  Google Scholar 

  16. Viljoen, M., Subden, R.E., Krizus, A., and van Vuuren, H.J.J. . 1994. Molecular analysis of the malic enzyme gene (mae2)of Schizosaccharomyces pombe . Yeast 10: 613–624.

    Article  CAS  PubMed  Google Scholar 

  17. Laing, E. and Pretorius, I.S. 1992. Synthesis and secretion of an Erwinia chrysanthemipectate lyase in Saccharomyces cerevisiaeregulated by different combinations of bacterial and yeast promoter and signal sequences. Gene 121: 35–45.

    Article  CAS  PubMed  Google Scholar 

  18. Maconi, E., Manachini, R., Aragozzini, F., Gennari, C., and Ricca, G. 1984. A study on the malo-alcoholic fermentation pathway in Schizosaccharomyces pombe . Biochem. J. 217: 585–588.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Murai, T., Tokushige, M., Nagai, J., and Katsuki, H. 1972. Studies on the regulatory functions of malic enzymes. J. Biochem. 71: 1015–1028.

    Article  CAS  PubMed  Google Scholar 

  20. Artus, N.N. and Edwards, G.E. 1985. NAD-malic enzyme from plants. FEBS Lett 182: 225–233.

    Article  CAS  Google Scholar 

  21. Yanisch-Perron, C., Vieira, J., and Messing, J. 1985. Improved M13 cloning vectors and host strains: nucleotide sequence of the M13mp18 and pUC19 vectors. Gene 33: 103–119.

    Article  CAS  PubMed  Google Scholar 

  22. Sambrook, J., Fritsh, E.F., and Maniatis, T. 1989. Molecular Cloning: A Laboratory Manual (2nd ed.). Cold Spring Harbor Laboratory Press, New York.

    Google Scholar 

  23. Sikorski, R.S. and Hieter, P. 1989. A system of shuttle vectors and yeast host strains designed for efficient manipulation of DMA in Saccharomyces cerevisiae . Genetics 122: 19–27.

    CAS  PubMed  PubMed Central  Google Scholar 

  24. Ausubel, F.M., Brent, R., Kingston, R.E., Moore, D.D., Seidman, J.G., Smith, J.A., and Struhl, K. (eds). 1995. Current protocols in molecular biology. John Wiley & Sons, New York.

    Google Scholar 

  25. Gietz, R.D. and Sugino, A., 1988. yeast-Escherichia colishuttle vectors constructed with in vitromutagenized yeast genes lacking six-base pair restriction sites. Gene 74: 527–534.

    Article  CAS  PubMed  Google Scholar 

  26. Kunkee, R.E. 1968. Simplified chromatographic procedure for detection of malolactic fermentation. Wines and Vines 49: 23–24.

    Google Scholar 

  27. Smith, D.B. and Johnson, K.S. 1988. Single step purification of polypeptides expressed in Escherichia colias fusions with glutathione S-transferase. Gene 67: 31–40.

    Article  CAS  PubMed  Google Scholar 

  28. Chirio, M.-C., Brethes, D., Napias, C., Grandier-Vazille, X., Rakotomanana, F., and Chevallier, J. 1990. Photoaffinity labelling of the purine-cytosine permease of Saccharomyces cerevisiae . Eur. J. Biochem. 194: 293–299.

    Article  CAS  PubMed  Google Scholar 

  29. Crous, J.M., Pretorius, I.S., and Van Zyl, W.H. 1995. Cloning and expression of an Aspergillus kawachiiendo-1,4-β-xylanase gene in Saccharomyces cerevisiae . Curr. Genet. 28: 467–473.

    Article  CAS  PubMed  Google Scholar 

  30. Martineau, B., Henick-Kling, T., and Acree, T. 1995. Reassessment of the influence of malolactic fermentation on the concentration of diacetyl in wines. Am. J. Enol. Vitic. 46: 385–388.

    CAS  Google Scholar 

Download references

Author information

Author notes

  1. Hendrik J.J. van Vuuren: Corresponding author (e-mail: hjjvv@maties.sun.ac.za).

Authors and Affiliations

  1. Department of Microbiology and Institute for Biotechnology, University of Stellenbosch, Stellenbosch, 7600, South Africa

    Heinrich Volschenk, Marinda Viljoen, Jandré Grobler, Barbara Petzold, Florian Bauer & Hendrik J.J. van Vuuren

  2. Department of Molecular Biology and Genetics, University of Guelph, Guelph, Ontario, NIG2WI, Canada

    Ron E. Subden

  3. Whitehead Institute for Biomedical Research, 9 Cambridge Center, Cambridge, MA, 02142, USA

    Richard A. Young

  4. Faculté d'Oenologie, Université de Bordeaux II, 351 cours de la Libération, 33405, Talence cedex, France

    Aline Lonvaud & Muriel Denayrolles

Authors
  1. Heinrich Volschenk
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Marinda Viljoen
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Jandré Grobler
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Barbara Petzold
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Florian Bauer
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Ron E. Subden
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Richard A. Young
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Aline Lonvaud
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Muriel Denayrolles
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Hendrik J.J. van Vuuren
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Volschenk, H., Viljoen, M., Grobler, J. et al. Engineering pathways for malate degradation in Saccharomyces cerevisiae. Nat Biotechnol 15, 253–257 (1997). https://doi.org/10.1038/nbt0397-253

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nbt0397-253

  • Springer Nature America, Inc.

This article is cited by

Search

Navigation