Skip to main content
Log in

Reduced production of diacetyl by overexpressing BDH2 gene and ILV5 gene in yeast of the lager brewers with one ILV2 allelic gene deleted

  • Fermentation, Cell Culture and Bioengineering - Original Paper
  • Published:
Journal of Industrial Microbiology & Biotechnology

Abstract

Diacetyl causes an unwanted buttery off-flavor in lager beer. The production of diacetyl is reduced by modifying the metabolic pathway of yeast in the beer fermentation process. In this study, BDH2 and ILV5 genes, coding diacetyl reductase and acetohydroxy acid reductoisomerase, respectively, were expressed using a PGK1 promoter in Saccharomyces cerevisiae, which deleted one ILV2 allelic gene. Diacetyl contents and fermentation performances were examined and compared. Results showed that the diacetyl content in beer was remarkably reduced by 16.52% in QI2-KP (one ILV2 allelic gene deleted), 55.65% in QI2-B2Y (overexpressed BDH2 gene and one ILV2 allelic gene deleted), and 69.13% in QI2-I5Y (overexpressed ILV5 gene and one ILV2 allelic gene deleted) compared with the host strain S2. The fermentation ability of mutant strains was similar to that of S2. Results of the present study can lead to further advances in this technology and its broad application in scientific investigations and industrial beer production.

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.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

Similar content being viewed by others

References

  1. Bartowsky EJ, Henschke PA (2004) The ‘buttery’ attribute of wine-diacetyl-desirability, spoilage and beyond. Int J Food Microbiol 96(3):235–252. doi:10.1016/j.ijfoodmicro.2004.05.013

    Article  CAS  PubMed  Google Scholar 

  2. Dillemans M, Goossens E, Goffin O, Masschelein CA (1987) The amplification effect of the ILV5 gene on the production of vicinal diketones in Saccharomyces cerevisiae. J Am Soc Brew Chem 45:81–84

    CAS  Google Scholar 

  3. Duong CT, Strack L, Futschik M, Katou Y, Nakao Y, Fujimura T, Shirahige K, Kodama Y, Nevoigt E (2011) Identification of Sc-type ILV6 as a target to reduce diacetyl formation in lager brewers’ yeast. Metab Eng 13(6):638–647. doi:10.1016/j.ymben.2011.07.005

    Article  CAS  PubMed  Google Scholar 

  4. Gietz RD, Woods RA (2002) Transformation of yeast by lithium acetate/single-stranded carrier DNA/polyethylene glycol method. Methods Enzymol 350:87–96. doi:10.1016/S0076-6879(02)50957-5

    Article  CAS  PubMed  Google Scholar 

  5. Gjermansen C, Nilsson-Tillgren T, Petersen JG, Kielland-Brandt MC, Sigsgaard P, Holmberg S (1988) Towards diacetyl-less brewers’ yeast. Influence of ilv2 and ilv5 mutations. J Basic Microbiol 28(3):175–183. doi:10.1002/jobm.3620280304

    Article  CAS  PubMed  Google Scholar 

  6. Gueldener U, Heinisch J, Koehler GJ, Voss D, Hegemann JH (2002) A second set of loxP marker cassettes for Cre-mediated multiple gene knockouts in budding yeast. Nucleic Acid Res 30(6):e23

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Haukeli A, Lie S (1978) Conversion of a-acetolactate and removal of diacetyl: a kinetic study. J Inst Brew 84:85–89. doi:10.1002/j.2050-0416.1978.tb03843.x

    Article  CAS  Google Scholar 

  8. Kele P, Li XH, Duerkop A (2013) New luminescent ruthenium probes for detection of diacety. Microchem J 108:156–160. doi:10.1016/j.microc.2012.10.012

    Article  CAS  Google Scholar 

  9. Lilly M, Lambrechts MG, Pretorius IS (2000) Effect of increased yeast alcohol acetyltransferase activity on flavor profiles of wine and distillates. Appl Environ Microbiol 66(2):744–753

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Omura F (2008) Targeting of mitochondrial Saccharomyces cerevisiae Ilv5p to the cytosol and its effect on vicinal diketone formation in brewing. Appl Microbiol Biotechnol 78:503–513. doi:10.1007/s00253-007-1333-x

    Article  CAS  PubMed  Google Scholar 

  11. Qin Y, Park HD (2012) Overexpressed acetohydroxyacid reductoisomerase (ILV5) gene in saccharomyces cerevisiae reduces diacetyl contents in Korean Campbell early and Muscat Bailey A grape wines. Korean Soc Appl Biol Chem 55:799–801. doi:10.1007/s13765-012-1157-y

    Article  CAS  Google Scholar 

  12. Rodrigues JA, Barros AA, Machado Cruz JM, Ferreira AA (1997) Determination of diacetyl in beer using differential-pulse polarography. J Inst Brew 103:311–314. doi:10.1002/j.2050-0416.1997.tb00962.x

    Article  CAS  Google Scholar 

  13. Saerens SM, Duong CT, Nevoigt E (2010) Genetic improvement of brewer’s yeast: current state, perspectives and limits. Appl Microbiol Biotechnol 86:1195–1212. doi:10.1007/s00253-010-2486-6

    Article  CAS  PubMed  Google Scholar 

  14. Saison D, De Schutter DP, Uyttenhove B, Delvaux F, Delvaux FR (2009) Contribution of staling compounds to the aged flavour of lager beer by studying their flavour thresholds. Food Chem 114(4):1206–1215. doi:10.1016/j.foodchem.2008.10.078

    Article  CAS  Google Scholar 

  15. Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning: a laboratory manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor

    Google Scholar 

  16. Sambrook J, Russell DW (2001) Molecular cloning: a laboratory manual, vol 2. CSHL Press, New York

    Google Scholar 

  17. Shi TT, Guo XU, Li P, Zhou Z, Xiao DG (2015) Diacetyl content reduction in industrial brewer’s yeast through ILV2 disruption and BDH1 expression. Eur Food Res Technol. doi:10.1007/s00217-015-2598-4 (unpublished results)

    Google Scholar 

  18. Villa KF, Lee S, Goossens E, Debourg A, Masschelein C (1995) Control of vicinal diketone production by brewer’s yeast. I. Effects of ILV5 and ILV3 gene amplification on vicinal diketone production and ILV enzyme activity. J Am Soc Brew Chem 53:49–53

    CAS  Google Scholar 

  19. Yamauchi Y, Okamoto T, Murayama H, Kajino K, Nagara A, Noguchi K (1995) Rapid maturation of beer using an immobilized yeast bioreactor. 2. Balance of total diacetyl reduction and regeneration. Biotechnology 38:109–116. doi:10.1016/0168-1656(94)00114-R

    CAS  Google Scholar 

  20. Zhang Y, Wang ZY, He XP, Liu N, Zhang BR (2008) New industrial brewing yeast strains with ILV2 disruption and LSD1 expression. Int J Food Microbiol 123(1–2):18–24. doi:10.1016/j.ijfoodmicro.2007.11.07T0

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

The current study was financially supported by the National Natural Science Foundation of China (31671834), the National Natural Science Foundation of China (31471724), and the Project supported by the National Natural Science Foundation of China (31271916).

Author information

Authors and Affiliations

Authors

Corresponding authors

Correspondence to Xue-Wu Guo or Dong-Guang Xiao.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Shi, TT., Li, P., Chen, SJ. et al. Reduced production of diacetyl by overexpressing BDH2 gene and ILV5 gene in yeast of the lager brewers with one ILV2 allelic gene deleted. J Ind Microbiol Biotechnol 44, 397–405 (2017). https://doi.org/10.1007/s10295-017-1903-6

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10295-017-1903-6

Keywords

Navigation