Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Research Paper
  • Published:

Xylitol Production by Recombinant Saccharomyces Cerevisiae

Abstract

We obtained efficient conversion of xylose to xylitol by transforming Saccharomyces cerevisiae with the gene encoding the xylose reductase (XR) of Pichia stipitis CBS 6054. Comparison of the chromosomal and cDNA copies of the XYL1 gene showed that the genomic XYL1 contains no introns, and an XR monomer of 318 amino acids (35,985 D) is encoded by an open reading frame of 954 bp. The amino acid sequence of the P. stipitis XR is similar to several aldose reductases, suggesting that P. stipitis XR is part of the aldoketo reductase superfamily. S. cerevisiae transformed with the XYL1 gene gave over 95% conversion of xylose into xylitol, a yield not obtainable with natural xylose utilizing yeasts.

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

Access options

Buy this article

Prices may be subject to local taxes which are calculated during checkout

Similar content being viewed by others

References

  1. Hyvönen, L. and Koivistoinen, P. 1983. Food technological evaluation of xylitol. Adv. in Food Research 28: 373–403.

    Article  Google Scholar 

  2. Melaja, A. and Hämäläinen, L. 1977. Process for making xylitol. US Patent 4,008,285.

    Google Scholar 

  3. Jeffries, T.W. 1990. Fermentation of D-xylose and cellobiose. p. 349–394 In: Yeast: Biotechnology and Biocatalysis. Verachtert, H. and De Mot, R. (Eds.). Marcel Dekker, Inc., NY.

    Google Scholar 

  4. Verduyn, C., Frank, J., Jzn Van Dijken, J.P. and Scheffers, W.A. 1985. Multiple forms of xylose reductase in Pachysolen tannophilus. FEMS Microbiol. Lett. 30: 313–317.

    Article  CAS  Google Scholar 

  5. Bruinenberg, P.M., de Bot, P.M., van Dijken, J.P. and Scheffers, W.A. 1984. NADH-linked aldose reductase: The key to anaerobic alcoholic fermentation of xylose by yeasts. Appl. Microbiol. Biotechnol. 19: 256–260.

    Article  CAS  Google Scholar 

  6. Ojamo, H., Ylinen, L. and Linko, M. 1987. Mikrobiologinen valmistusmenetelmä. Finnish patent 76377.

  7. Meyrial, V., Delgenes, J.P., Moletta, R., Navarro, J.M. and Inra, I.A.A. 1991. Xylitol production from D-xylose by Candida guillermondii—Fermentation behaviour. Biotechnol. Lett. 13: 281–286.

    Article  CAS  Google Scholar 

  8. Verduyn, C., Van Kleff, R., Schreuder, H., Van Dijken, J.P. and Scheffers, W.A. 1985. Properties of the NAD(P)H-dependent xylose reductase from the xylose fermenting yeast Pichia stipitis. Biochem. J. 226: 669–677.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Rizzi, M., Erlemann, P., Bui-Thanh, N.-A. and Dellweg, H. 1988. Xylose fermentation by yeasts 4. Purification and kinetic studies of xylose reductase from Pichia stipitis. Appl. Microbiol. Biotechnol. 29: 148–154.

    Article  CAS  Google Scholar 

  10. Ditzelmüller, G., Kubicek, C.P., Wöhrer, W. and Röhr, M. 1984. Xylose metabolism in Pachysolen tannophilus: Purification and properties of xylose reductase. Can. J. Microbiol. 30: 1330–1336.

    Article  Google Scholar 

  11. Sharp, P., Cowe, E., Higgins, D.G., Shields, D.C., Wolfe, K.H. and Wright, F. 1988. Codon usage patterns in Escherichia coli, Bacillus subtilis, Schizosaccharomyes pombe, Drosophila melanogaster and Homo sapiens; a review of the considerable within-species diversity. Nuc. Acids Res. 16: 8207–8211.

    Article  CAS  Google Scholar 

  12. Bohren, K.M., Bullock, B., Wermuth, B. and Gabbay, K.M. 1989. The aldo-keto reductase superfamily cDNAs and deduced amino acid sequences of human aldehyde and aldose reductases. J. Biol. Chem. 264: 9547–9551.

    CAS  PubMed  Google Scholar 

  13. Carper, D., Nishimura, C., Shinohara, T., Dietzchold, B., Wistow, G., Kador, P. and Kinoshita, I.H. 1987. Aldose reductase and rho-crystallin belong to the same protein superfamily as aldehyde reductase. FEES Lett. 220: 209–213.

    Article  CAS  Google Scholar 

  14. Petrash, J.M. and Favello, A.D. 1989. Isolation and characterization of cDNA clones encoding aldose reductase. Curr. Eye Res. 8: 1021–1027.

    Article  CAS  PubMed  Google Scholar 

  15. Watanabe, K., Fuji, Y., Nakayama, K., Okhuba, H., Kuramitsu, S., Kagamiyama, H., Nakanishi, S. and Hayashi, O. 1988. Structural similarity of bovine lung prostaglandin F synthase to lens epsilon-crystallin of the European common frog. Proc. Natl. Acad. Sci. USA 85: 11–15.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Winters, C.J., Molowa, D.T. and Guzelian, P.S. 1990. Isolation and characterization of cloned cDNAs encoding human liver chlordecone reductase. Biochemistry 29: 1080–1087.

    Article  CAS  PubMed  Google Scholar 

  17. Ho, N.W.Y., Lin, F.P., Huang, S., Andrews, P.C. and Tsao, G.T. 1990. Purification, characterization, and amino acid terminal sequence of xylose reductase from Candida shehatae. Enzyme Microb. Technol. 12: 33–39.

    Article  CAS  PubMed  Google Scholar 

  18. Kurtzman, C.P. 1990. Candida shehatae—genetic diversity and phylo-genetic relationships with other xylose-fermenting yeasts. Antonie van Leeuwenhoek 57: 215–222.

    Article  CAS  PubMed  Google Scholar 

  19. Prior, B.A., Kilian, S.G. and du Perez, J.C. 1989. Fermentation of D-xylose by the yeasts Candida shehatae and Pichia stipitis. Process. Biochem. 24: 21–32.

    CAS  Google Scholar 

  20. Kötter, P., Amore, R., Hollenberg, C.P. and Ciriacy, M. 1990. Isolation and characterization of the Pichia stipitis xylitol dehydroge-nase gene, XYL2, and construction of a xylose-utilizing Saccharomyces cerevisiae transformant. Curr. Genet. 18: 493–500.

    Article  PubMed  Google Scholar 

  21. Van Zyl, C., Prior, O.L., Kilian, S.G. and Kock, J.L.F. 1989. D-Xylose utilization by Saccharomyces cerevisiae. J. Gen. Microbiol. 135: 2791–2798.

    CAS  PubMed  Google Scholar 

  22. Barbosa, M.F.S., de Medeiros, M.B., de Manchila, I.M., Schneider, H. and Lee, H. 1988. Screening for yeasts for production of xylitol from D-xylose and some factors which affect xylitol yield in Candida guilliermondii. J. Ind. Microbiol. 3: 241–251.

    Article  CAS  Google Scholar 

  23. Mellor, J., Dobson, M.J., Roberts, N.A., Tuite, M.F., Emtage, J.S., White, S., Lowe, P.A., Patel, T., Kingsman, A.J. and Kingsman, S.M. 1983. Efficient synthesis of enzymatically active calf chymosin in Saccharomyces cerevisiae. Gene 24: 1–14.

    Article  CAS  PubMed  Google Scholar 

  24. Bradford, M.M. 1976. A rapid and sensitive method for the quantification of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72: 248–254.

    Article  CAS  PubMed  Google Scholar 

  25. Smiley, K.L. and Bolen, P.L. 1982. Demonstration of D-xylose reductase and D-xylitol dehydrogenase in Pachysolen tannophilus. Biotechnol. Lett. 4: 607–610.

    Article  CAS  Google Scholar 

  26. Laemmli, U.K. 1970. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227: 680–685.

    Article  CAS  PubMed  Google Scholar 

  27. Shaw, C.R. and Prasad, R. 1970. Starch electrophoresis of enzymes—a compilation of recipes. Biochem. Genet. 4: 297–320.

    Article  CAS  PubMed  Google Scholar 

  28. Chirgwin, J.M., Przybyla, A.E., MacDonald, R.J. and Rutter, W.J. 1979. Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease. Biochemistry 18: 5294–5299.

    Article  CAS  PubMed  Google Scholar 

  29. Young, R.A. and Davis, R.W. 1983. Yeast RNA polymerase II genes: isolation with antibody probes. Science 222: 778–782.

    Article  CAS  PubMed  Google Scholar 

  30. Güssow, D. and Clackson, T. 1989. Direct clone characterization from plaques and colonies by the polymerase chain reaction. Nuc. Acid Res. 17: 4000.

    Article  Google Scholar 

  31. Cryer, D.R., Eccleshall, R. and Marmur, J. 1975. Isolation of yeast DNA. Methods Cell Biol. 12: 39–44.

    Article  CAS  PubMed  Google Scholar 

  32. Zagursky, R.I., Berman, M.L., Baumeister, K. and Lomax, N. 1986. Rapid and easy sequencing of large linear double stranded DNA and supercoiled plasmid DNA. Gene Anal. Techn. 2: 89–94.

    Article  Google Scholar 

  33. Devereux, J., Haberli, P. and Smithies, O. 1984. A comprehensive set of sequence analysis programs for the VAX.Nuc. Acids Res. 12: 387–395.

    Article  CAS  Google Scholar 

  34. Ito, H., Fukuda, M., Murata, K. and Kimura, A. 1983. Transformation of intact yeast cells treated with alkali cations. J. Bact. 153: 163–168.

    CAS  PubMed  PubMed Central  Google Scholar 

  35. Sherman, F., Fink, G. and Hicks, I.B. 1983 Methods in Yeast Genetics. A Laboratory Manual. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY.

    Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Rights and permissions

Reprints and permissions

About this article

Cite this article

Hallborn, J., Walfridsson, M., Airaksinen, U. et al. Xylitol Production by Recombinant Saccharomyces Cerevisiae. Nat Biotechnol 9, 1090–1095 (1991). https://doi.org/10.1038/nbt1191-1090

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nbt1191-1090

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing