Biotechnology Letters

, Volume 38, Issue 12, pp 2145–2151 | Cite as

Surface-engineered Saccharomyces cerevisiae displaying α-acetolactate decarboxylase from Acetobacter aceti ssp xylinum

  • Rudolf Cejnar
  • Kateřina Hložková
  • Pavel Kotrba
  • Pavel Dostálek
Original Research Paper



To convert α-acetolactate into acetoin by an α-acetolactate decarboxylase (ALDC) to prevent its conversion into diacetyl that gives beer an unfavourable buttery flavour.


We constructed a whole Saccharomyces cerevisiae cell catalyst with a truncated active ALDC from Acetobacter aceti ssp xylinum attached to the cell wall using the C-terminal anchoring domain of α-agglutinin. ALDC variants in which 43 and 69 N-terminal residues were absent performed equally well and had significantly decreased amounts of diacetyl during fermentation. With these cells, the highest concentrations of diacetyl observed during fermentation were 30 % less than those in wort fermented with control yeasts displaying only the anchoring domain and, unlike the control, virtually no diacetyl was present in wort after 7 days of fermentation.


Since modification of yeasts with ALDC variants did not affect their fermentation performance, the display of α-acetolactate decarboxylase activity is an effective approach to decrease the formation of diacetyl during beer fermentation.


Acetobacter aceti Acetoin α-Acetolactate α-Acetolactate decarboxylase Beer Diacetyl Saccharomyces cerevsiae 



This work was supported by the project TE02000177 “Centre for Innovative Use and Strengthening of Competitiveness of Czech Brewery Raw Materials and Products” of the Technology Agency of the Czech Republic.

Supporting information

Supplementary Table 1—Fermentation performance of S. cerevisiae W303-1A strains transformed with indicated plasmids.

Supplementary Fig. 1—Surface display of ALDC130 and ALDC208 fusions to V5-AGα1Cp expressed in S. cerevisiae W303-1A.

Supplementary material

10529_2016_2205_MOESM1_ESM.docx (203 kb)
Supplementary material 1 (DOCX 203 kb)


  1. ASBC Methods of Analysis, online. Beer method 25. Diacetyl. Approved (2011) rev. (2014). American Society of Brewing Chemists, St. Paul doi: 10.1094/ASBCMOA-Beer-25
  2. ASBC Methods of Analysis, online. Beer method 4. Alcohol. Approved (1958), rev. (2004). American Society of Brewing Chemists, St. Paul doi: 10.1094/ASBCMOA-Beer-4
  3. ASBC Methods of Analysis, online. Beer method 5. Real Extract. Approved (1958), rev. (1982). American Society of Brewing Chemists, St. Paul doi: 10.1094/ASBCMOA-Beer-5
  4. Blomqvist K, Suihko ML, Knowles J, Penttila M (1991) Chromosomal integration and expression of two bacterial α-acetolactate decarboxylase genes in Brewers’ yeast. Appl Environ Microbiol 57:2796–2803PubMedPubMedCentralGoogle Scholar
  5. Ghaemmaghami S et al (2003) Global analysis of protein expression in yeast. Nature 425:737–741CrossRefPubMedGoogle Scholar
  6. Gibson B, Krogerus K, Ekberg J, Monroux A (2015) Variation in α-acetolactate production within the hybrid lager yeast group Saccharomyces pastorianus and affirmation of the central role of the ILV6 gene. Yeast 32:301–316PubMedGoogle Scholar
  7. Gietz RD, Schiestl RH, Willems AR, Woods RA (1995) Studies of the transformation of intact yeast cells by the LiAc/SS-DNA/PEG procedure. Yeast 11:355–360CrossRefPubMedGoogle Scholar
  8. Godtfredsen SE, Ottesen M (1982) Maturation of beer with α-acetolactate decarboxylase. Carlsberg Res Commun 47:93–102CrossRefGoogle Scholar
  9. Godtfredsen SE, Lorck H, Sigsgaard P (1983) On the occurrence of α-acetolactate decarboxylases among microorganisms. Carlsberg Res Commun 48:239–247CrossRefGoogle Scholar
  10. Godtfredsen SE, Rasmussen AM, Ottesen M, Mathiasen T, Ahrenst-Larsen B (1984) Application of the acetolactate decarboxylase from Lactobacillus casei for accelerated maturation of beer. Carlsberg Res Commun 49:69–74CrossRefGoogle Scholar
  11. Goelling D, Stahl U (1988) Cloning and expression of an alpha-acetolactate decarboxylase gene from Streptococcus lactis subsp diacetylactis in Escherichia coli. Appl Environ Microbiol 54:1889–1891PubMedPubMedCentralGoogle Scholar
  12. Inoue T (2008) Diacetyl in fermented foods and beverages. American Society of Brewing Chemists, St. Paul, p 139Google Scholar
  13. Inoue T, Yamamoto Y (1970) Diacetyl and beer fermentation. Proc Am Soc Brew Chem 28:198–208Google Scholar
  14. Kotrba P, Ruml T (2010) Surface display of metal fixation motifs of bacterial P1-type ATPases specifically promotes biosorption of Pb2+ by Saccharomyces cerevisiae. Appl Environ Microbiol 76:2615–2622CrossRefPubMedPubMedCentralGoogle Scholar
  15. Krogerus K, Gibson BR (2013) 125th anniversary review: diacetyl and its control during brewery fermentation. J Inst Brew 119:86–97Google Scholar
  16. Mumberg D, Mueller R, Funk M (1995) Yeast vectors for the controlled expression of heterologous proteins in different genetic backgrounds. Gene 156:119–122CrossRefPubMedGoogle Scholar
  17. Murai T et al (1997) Construction of a starch-utilizing yeast by cell surface engineering. Appl Environ Microbiol 63:1362–1366PubMedPubMedCentralGoogle Scholar
  18. Petruzzi L, Corbo MR, Sinigaglia M, Bevilacqua A (2016) Brewer’s yeast in controlled and uncontrolled fermentations, with a focus on novel, nonconventional, and superior strains. Food Rev Int 32:341–363CrossRefGoogle Scholar
  19. Ruehle G et al (2014) Headspace gas chromatography/electron capture detector analysis of total vicinal diketones in beer. J Am Soc Brew Chem 72:305–306Google Scholar
  20. Sone H, Fujii T, Kondo K, Shimizu F (1988) Nucleotide sequence and expression of the enterobacter aerogenes α-acetolactate decarboxylase gene in brewers’ yeast. Appl Environ Microbiol 54:38–42PubMedPubMedCentralGoogle Scholar
  21. Tanaka T, Kondo A (2015) Cell surface engineering of industrial microorganisms for biorefining applications. Biotechnol Adv 33:1403–1411CrossRefPubMedGoogle Scholar
  22. Tanaka T, Yamada R, Ogino C, Kondo A (2012) Recent developments in yeast cell surface display toward extended applications in biotechnology. Appl Microbiol Biotechnol 95:577–591CrossRefPubMedGoogle Scholar
  23. Vinopal S, Ruml T, Kotrba P (2007) Biosorption of Cd2+ and Zn2+ by cell surface-engineered Saccharomyces cerevisiae. Int Biodeterior Biodegrad 60:96–102CrossRefGoogle Scholar
  24. Vogel J, Wackerbauer K, Stahl U (1995) Improvement of beer brewing by using genetically modified yeast. ACS Symp Ser 605:160–170CrossRefGoogle Scholar
  25. Yamano S, Kondo K, Tanaka J, Inoue T (1994a) Construction of a brewer’s yeast having α-acetolactate decarboxylase gene from Acetobacter aceti ssp xylinum integrated in the genome. J Biotechnol 32:173–178CrossRefPubMedGoogle Scholar
  26. Yamano S, Tanaka J, Inoue T (1994b) Cloning and expression of the gene encoding α-acetolactate decarboxylase from Acetobacter aceti ssp xylinum in brewer’s yeast. J Biotechnol 32:165–171CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media Dordrecht 2016

Authors and Affiliations

  1. 1.Department of Biotechnology, Faculty of Food and Biochemical TechnologyUniversity of Chemistry and Technology, PraguePrague 6-DejviceCzech Republic
  2. 2.Department of Biochemistry and Microbiology, Faculty of Food and Biochemical TechnologyUniversity of Chemistry and Technology, PraguePrague 6-DejviceCzech Republic

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