Skip to main content
SpringerLink
Log in

Enhanced freeze tolerance of baker’s yeast by overexpressed trehalose-6-phosphate synthase gene (TPS1) and deleted trehalase genes in frozen dough

  • Genetics and Molecular Biology of Industrial Organisms
  • Published:
Journal of Industrial Microbiology & Biotechnology

Abstract

Several recombinant strains with overexpressed trehalose-6-phosphate synthase gene (TPS1) and/or deleted trehalase genes were obtained to elucidate the relationships between TPS1, trehalase genes, content of intracellular trehalose and freeze tolerance of baker’s yeast, as well as improve the fermentation properties of lean dough after freezing. In this study, strain TL301TPS1 overexpressing TPS1 showed 62.92 % higher trehalose-6-phosphate synthase (Tps1) activity and enhanced the content of intracellular trehalose than the parental strain. Deleting ATH1 exerted a significant effect on trehalase activities and the degradation amount of intracellular trehalose during the first 30 min of prefermentation. This finding indicates that acid trehalase (Ath1) plays a role in intracellular trehalose degradation. NTH2 encodes a functional neutral trehalase (Nth2) that was significantly involved in intracellular trehalose degradation in the absence of the NTH1 and/or ATH1 gene. The survival ratio, freeze-tolerance ratio and relative fermentation ability of strain TL301TPS1 were approximately twice as high as those of the parental strain (BY6-9α). The increase in freeze tolerance of strain TL301TPS1 was accompanied by relatively low trehalase activity, high Tps1 activity and high residual content of intracellular trehalose. Our results suggest that overexpressing TPS1 and deleting trehalase genes are sufficient to improve the freeze tolerance of baker’s yeast in frozen dough. The present study provides guidance for the commercial baking industry as well as the research on the intracellular trehalose mobilization and freeze tolerance of baker’s yeast.

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

Fig. 1
Fig. 2
Fig. 3

Similar content being viewed by others

References

  1. Alizadeh P, Klionsky DJ (1996) Purification and biochemical characterization of the ATH1 gene product, vacuolar acid trehalase, from Saccharomyces cerevisiae. FEBS Lett 391(3):273–278

    Article  CAS  PubMed  Google Scholar 

  2. Almeida M, Pais C (1996) Leavening ability and freeze tolerance of yeasts isolated from traditional corn and rye bread doughs. Appl Environ Microbiol 62(12):4401–4404

    CAS  PubMed Central  PubMed  Google Scholar 

  3. Basu A, Bhattacharyya S, Chaudhuri P, Sengupta S, Ghosh AK (2006) Extracellular trehalose utilization by Saccharomyces cerevisiae. Biochim Biophys Acta 1760(2):134–140

    Article  CAS  PubMed  Google Scholar 

  4. Destruelle M, Holzer H, Klionsky DJ (1995) Isolation and characterization of a novel yeast gene, ATH1, that is required for vacuolar acid trehalase activity. Yeast 11(11):1015–1025

    Article  CAS  PubMed  Google Scholar 

  5. Garre E, Matallana E (2009) The three trehalases Nth1p, Nth2p and Ath1p participate in the mobilization of intracellular trehalose required for recovery from saline stress in Saccharomyces cerevisiae. Microbiology 155(9):3092–3099

    Article  CAS  PubMed  Google Scholar 

  6. Garre E, Pérez-Torrado R, Gimeno-Alcañiz JV, Matallana E (2009) Acid trehalase is involved in intracellular trehalose mobilization during postdiauxic growth and severe saline stress in Saccharomyces cerevisiae. FEMS Yeast Res 9(1):52–62

    Article  CAS  PubMed  Google Scholar 

  7. Ge XY, Xu Y, Chen X (2013) Improve carbon metabolic flux in Saccharomyces cerevisiae at high temperature by overexpressed TSL1 gene. J Ind Microbiol Biotechnol 40(3–4):345–352. doi:10.1007/s10295-013-1233-2

    Article  CAS  PubMed  Google Scholar 

  8. Harris SD, Cotter DA (1988) Transport of yeast vacuolar trehalase to the vacuole. Can J Microbiol 34(7):835–838

    Article  CAS  PubMed  Google Scholar 

  9. Hottiger T, Schmutz P, Wiemken A (1987) Heat-induced accumulation and futile cycling of trehalose in Saccharomyces cerevisiae. J Bacteriol 169(12):5518–5522

    CAS  PubMed Central  PubMed  Google Scholar 

  10. Hsu K, Hoseney R, Seib P (1979) Frozen dough. I. Factors affecting stability of yeasted doughs [quality, fermentation, freeze damage]. Cereal Chem 56:419–424

    Google Scholar 

  11. Huang J, Reggiori F, Klionsky DJ (2007) The transmembrane domain of acid trehalase mediates ubiquitin-independent multivesicular body pathway sorting. Mol Biol Cell 18(7):2511–2524

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  12. Izawa S, Sato M, Yokoigawa K, Inoue Y (2004) Intracellular glycerol influences resistance to freeze stress in Saccharomyces cerevisiae: analysis of a quadruple mutant in glycerol dehydrogenase genes and glycerol-enriched cells. Appl Microbiol Biotechnol 66(1):108–114

    Article  CAS  PubMed  Google Scholar 

  13. Jules M, Beltran G, François J, Parrou JL (2008) New insights into trehalose metabolism by Saccharomyces cerevisiae: NTH2 encodes a functional cytosolic trehalase, and deletion of TPS1 reveals Ath1p-dependent trehalose mobilization. Appl Environ Microbiol 74(3):605–614

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  14. Jules M, Guillou V, François J, Parrou J-L (2004) Two distinct pathways for trehalose assimilation in the yeast Saccharomyces cerevisiae. Appl Environ Microbiol 70(5):2771–2778. doi:10.1128/aem.70.5.2771-2778.2004

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  15. Kim J, Alizadeh P, Harding T, Hefner-Gravink A, Klionsky DJ (1996) Disruption of the yeast ATH1 gene confers better survival after dehydration, freezing, and ethanol shock: potential commercial applications. Appl Environ Microbiol 62(5):1563–1569

    CAS  PubMed Central  PubMed  Google Scholar 

  16. Lewis J, Learmonth R, Watson K (1993) Role of growth phase and ethanol in freeze-thaw stress resistance of Saccharomyces cerevisiae. Appl Environ Microbiol 59(4):1065–1071

    CAS  PubMed Central  PubMed  Google Scholar 

  17. Lv Y, Xiao D, He D, Guo X (2008) Construction and stress tolerance of trehalase mutant in Saccharomyces cerevisiae. Acta microbiologica Sinica 48(10):1301–1307 (Wei Sheng Wu Xue Bao)

    PubMed  Google Scholar 

  18. Murakami Y, Yokoigawa K, Kawai F, Kawai H (1996) Lipid composition of commercial bakers’ yeasts having different freeze-tolerance in frozen dough. Biosci Biotechnol Biochem 60:1874–1878

    Article  CAS  PubMed  Google Scholar 

  19. Nwaka S, Kopp M, Holzer H (1995) Expression and function of the trehalase genes NTH1 and YBR0106 in Saccharomyces cerevisiae. J Biol Chem 270(17):10193–10198

    Article  CAS  PubMed  Google Scholar 

  20. Nwaka S, Mechler B, Destruelle M, Holzer H (1995) Phenotypic features of trehalase mutants in Saccharomyces cerevisiae. FEBS Lett 360(3):286–290

    Article  CAS  PubMed  Google Scholar 

  21. Nwaka S, Mechler B, Holzer H (1996) Deletion of the ATH1 gene in Saccharomyces cerevisiae prevents growth on trehalose. FEBS Lett 386(2):235–238

    Article  CAS  PubMed  Google Scholar 

  22. Parrou JL, Jules M, Beltran G, François J (2005) Acid trehalase in yeasts and filamentous fungi: localization, regulation and physiological function. FEMS Yeast Res 5(6–7):503–511

    Article  CAS  PubMed  Google Scholar 

  23. Rossouw D, Heyns EH, Setati ME, Bosch S, Bauer FF (2013) Adjustment of trehalose metabolism in wine Saccharomyces cerevisiae strains to modify ethanol yields. Appl Environ Microbiol 79(17):5197–5207. doi:10.1128/AEM.00964-13

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  24. Sasano Y, Haitani Y, Hashida K, Ohtsu I, Shima J, Takagi H (2012) Overexpression of the transcription activator Msn2 enhances the fermentation ability of industrial baker’s yeast in frozen dough. Biosci Biotechnol Biochem 76(3):624–662. doi:10.1271/bbb.110959

    Article  CAS  PubMed  Google Scholar 

  25. Sebollela A, Louzada PR, Sola-Penna M, Sarone-Williams V, Coelho-Sampaio T, Ferreira ST (2004) Inhibition of yeast glutathione reductase by trehalose: possible implications in yeast survival and recovery from stress. Int J Biochem Cell Biol 36(5):900–908

    Article  CAS  PubMed  Google Scholar 

  26. Shima J, Hino A, Yamada-Iyo C, Suzuki Y, Nakajima R, Watanabe H, Mori K, Takano H (1999) Stress tolerance in doughs of Saccharomyces cerevisiae trehalase mutants derived from commercial baker’s yeast. Appl Environ Microbiol 65(7):2841–2846

    CAS  PubMed Central  PubMed  Google Scholar 

  27. Shima J, Sakata-Tsuda Y, Suzuki Y, Nakajima R, Watanabe H, Kawamoto S, Takano H (2003) Disruption of the CAR1 gene encoding arginase enhances freeze tolerance of the commercial baker’s yeast Saccharomyces cerevisiae. Appl Environ Microbiol 69(1):715–718

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  28. Shima J, Takagi H (2009) Stress-tolerance of baker’s-yeast (Saccharomyces cerevisiae) cells: stress-protective molecules and genes involved in stress tolerance. Biotechnol Appl Biochem 53(3):155–164

    Article  CAS  PubMed  Google Scholar 

  29. Singer MA, Lindquist S (1998) Multiple effects of trehalose on protein folding in vitro and in vivo. Mol Cell 1(5):639–648

    Article  CAS  PubMed  Google Scholar 

  30. Stewart PR (1982) Trehalose extraction and determination. In: Prescott DM (ed) Methods in cell biology, vol 12. Academic Press, London, pp 111–147

    Google Scholar 

  31. Sun X, Zhang C, Dong J, Wu M, Zhang Y, Xiao D (2012) Enhanced leavening properties of baker’s yeast overexpressing MAL62 with deletion of MIG1 in lean dough. J Ind Microbiol Biotechnol 39(10):1533–1539

    Article  PubMed  Google Scholar 

  32. Terao Y, Nakamori S, Takagi H (2003) Gene dosage effect of l-proline biosynthetic enzymes on l-proline accumulation and freeze tolerance in Saccharomyces cerevisiae. Appl Environ Microbiol 69(11):6527–6532

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  33. Teunissen A, Dumortier F, Gorwa M-F, Bauer J, Tanghe A, Loïez A, Smet P, Van Dijck P, Thevelein JM (2002) Isolation and characterization of a freeze-tolerant diploid derivative of an industrial baker’s yeast strain and its use in frozen doughs. Appl Environ Microbiol 68(10):4780–4787

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  34. Trevelyan WE, Harrison JS (1956) Studies on yeast metabolism. 5. The trehalose content of baker’s yeast during anaerobic fermentation. Biochem J 62(2):177–183

    CAS  PubMed Central  PubMed  Google Scholar 

  35. Wolfe KH, Lohan AJ (1994) II. Yeast sequencing reports. Sequence around the centromere of Saccharomyces cerevisiae chromosome II: similarity of CEN2 to CEN4. Yeast 10(S1994A):S41–S46

    Article  PubMed  Google Scholar 

  36. Yokoigawa K, Sato M, Soda K (2006) Simple improvement in freeze-tolerance of bakers’ yeast with poly-gamma-glutamate. J Biosci Bioeng 102(3):215–219. doi:10.1263/jbb.102.215

    Article  CAS  PubMed  Google Scholar 

  37. Zhang F, Wang ZP, Chi Z, Raoufi Z, Abdollahi S, Chi ZM (2013) The changes in Tps1 activity, trehalose content and expression of TPS1 gene in the psychrotolerant yeast Guehomyces pullulans 17-1 grown at different temperatures. Extremophiles 17(2):241–249. doi:10.1007/s00792-013-0511-2

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

This work was financially supported by the National Natural Science Foundation of China (31171730), National High Technology Research and Development Program of China (863 Program) (Grant No. 2013AA102106), The Cheung Kong Scholars and Innovative Research Team Program in University of Ministry of Education, China (Grant No. IRT1166) and the Youth Foundation of Application Base and Frontier Technology Project of Tianjin, China (12JCQNJC06500).

Author information

Authors and Affiliations

  1. Tianjin Industrial Microbiology Key Laboratory, College of Biotechnology, Tianjin University of Science and Technology, Tianjin, 300457, People’s Republic of China

    Haigang Tan, Jian Dong, Guanglu Wang, Haiyan Xu, Cuiying Zhang & Dongguang Xiao

  2. Key Laboratory of Industrial Fermentation Microbiology, Ministry of Education, Tianjin, People’s Republic of China

    Haigang Tan, Jian Dong, Guanglu Wang, Haiyan Xu, Cuiying Zhang & Dongguang Xiao

  3. College of Food Science and Engineering, Qingdao Agricultural University, Qingdao, 266109, People’s Republic of China

    Haigang Tan

Authors
  1. Haigang Tan
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Jian Dong
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Guanglu Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Haiyan Xu
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Cuiying Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Dongguang Xiao
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Dongguang Xiao.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Tan, H., Dong, J., Wang, G. et al. Enhanced freeze tolerance of baker’s yeast by overexpressed trehalose-6-phosphate synthase gene (TPS1) and deleted trehalase genes in frozen dough. J Ind Microbiol Biotechnol 41, 1275–1285 (2014). https://doi.org/10.1007/s10295-014-1467-7

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10295-014-1467-7

Keywords

Search

Navigation