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Functional Expression of the DUR3 Gene in a Wine Yeast Strain to Minimize Ethyl Carbamate in Chardonnay Wine

¹ M.Sc. student, Wine Research Centre, University of British Columbia, Vancouver, V6T 1Z4, Canada; ² Senior research scientist, Phyterra Yeast Inc., PO Box 21147, Charlottetown, C1A 9H6 Canada; and ³ Professor and Eagles Chair, Director of Wine Research Centre, University of British Columbia, Vancouver, V6T 1Z4, Canada.

Acknowledgments: This work was funded by a grant from Natural Sciences and Engineering Research Council of Canada (217271-04) to H.J.J. van Vuuren.

The authors thank Lina Madilao of the UBC Wine Research Centre for GC-MS analysis of wines.

A U.S. provisional patent has been filed (61/071,138) by H.J.J. van Vuuren and J.I. Husnik on the constitutive expression of DUR3 in wine yeast to limit ethyl carbamate production in wine.

^* Corresponding author (email: hjjvv{at}interchange.ubc.ca; fax: 604 822 5143)

During alcoholic fermentation Saccharomyces cerevisiae metabolizes arginine to ornithine and urea. Urea can be metabolized by wine yeasts; however, the presence of good nitrogen sources in grape must leads to transcriptional suppression of genes involved in urea import and metabolism. Urea is subsequently exported out of the cell where it spontaneously reacts with ethanol in wine to form the carcinogen ethyl carbamate (EC). Constitutive expression of DUR1,2 in the wine yeast UC Davis 522 (Montrachet) leads to an 89% reduction in the EC content of Chardonnay wine. To reabsorb urea secreted into fermenting grape must by non-urea-degrading yeast, we constitutively expressed the DUR3 gene under the control of the S. cerevisiae PGK1 promoter and terminator signals and integrated this linear cassette into the TRP1 locus of S. cerevisiae strain 522. The urea-importing strain 522^DUR3 reduced EC by 81% in Chardonnay wine and was shown to be approximately four times as effective as the urea-degrading strain 522^DUR1,2 at reducing EC in Chardonnay wine made from must with high endogenous urea levels.

Key words: wine, ethyl carbamate, DUR3, urea