Skip to main content
SpringerLink
Log in

Molecular Tools and Protocols for Engineering the Acid-Tolerant Yeast Zygosaccharomyces bailii as a Potential Cell Factory

  • Protocol
  • First Online:
Yeast Metabolic Engineering

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1152))

Abstract

Microorganisms offer a tremendous potential as cell factories, and they are indeed used by humans for centuries for biotransformations. Among them, yeasts combine the advantage of unicellular state with a eukaryotic organization, and, in the era of biorefineries, their biodiversity can offer solutions to specific process constraints. Zygosaccharomyces bailii, an ascomycetales budding yeast, is widely known for its peculiar tolerance to various stresses, among which are organic acids. Despite the possibility to apply with this yeast some of the molecular tools and protocols routinely used to manipulate Saccharomyces cerevisiae, adjustments and optimizations are necessary. Here, we describe in detail protocols for transformation, for target gene disruption or gene integration, and for designing episomal expression plasmids helpful for developing and further studying the yeast Z. bailii.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution
Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 84.99
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 119.00
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 109.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Erickson B, Nelson WP (2012) Perspective on opportunities in industrial biotechnology in renewable chemicals. Biotechnol J 7:176–185

    Article  CAS  Google Scholar 

  2. Otero JM, Nielsen J (2010) Industrial systems biology. Biotechnol Bioeng 105:439–460

    Article  CAS  Google Scholar 

  3. Porro D, Gasser B, Fossati T, Maurer M, Branduardi P, Sauer M, Mattanovich D (2011) Production of recombinant proteins and metabolites in yeasts: when are these systems better than bacterial production systems? Appl Microbiol Biotechnol 89:939–948

    Article  CAS  Google Scholar 

  4. Sauer M, Porro D, Mattanovich D, Branduardi P (2010) 16 years research on lactic acid production with yeast – ready for the market? Biotechnol Genet Eng Rev 27:229–256

    Article  CAS  Google Scholar 

  5. Mattanovich D, Branduardi P, Dato L, Gasser B, Sauer M, Porro D (2012) Recombinant protein production in yeasts. Methods Mol Biol 824:329–358

    Article  CAS  Google Scholar 

  6. Johnson EA (2013) Biotechnology of non-Saccharomyces yeasts-the ascomycetes. Appl Microbiol Biotechnol 97(2):503–517

    Google Scholar 

  7. Thomas DS, Davenport RR (1985) Zygosaccharomyces bailii – a profile of characteristics and spoilage activities. Food Microbiol 2:157–169

    Article  Google Scholar 

  8. Cole MB, Keenan MHJ (1987) Effects of weak acids and external ph on the intracellular pH of Zygosaccharomyces bailii, and its implications in weak-acid resistance. Yeast 3:23–32

    Article  CAS  Google Scholar 

  9. Makdesi AK, Beuchat LR (1996) Evaluation of media for enumerating heat-stressed, benzoate-resistant Zygosaccharomyces bailii. Int J Food Microbiol 33:169–181

    Article  CAS  Google Scholar 

  10. Sousa MJ, Miranda L, CorteReal M, Leao C (1996) Transport of acetic acid in Zygosaccharomyces bailii: effects of ethanol and their implications on the resistance of the yeast to acidic environments. Appl Environ Microbiol 62:3152–3157

    CAS  Google Scholar 

  11. Sousa MJ, Rodrigues F, Corte-Real M, Leao C (1998) Mechanisms underlying the transport and intracellular metabolism of acetic acid in the presence of glucose in the yeast Zygosaccharomyces bailii. Microbiology 144:665–670

    Article  CAS  Google Scholar 

  12. Dang TD, De Maeseneire SL, Zhang BY, De Vos WH, Rajkovic A, Vermeulen A, Van Impe JF, Devlieghere F (2012) Monitoring the intracellular pH of Zygosaccharomyces bailii by green fluorescent protein. Int J Food Microbiol 156:290–295

    Article  CAS  Google Scholar 

  13. Guerreiro JF, Mira NP, Sá-Correia I (2012) Adaptive response to acetic acid in the highly resistant yeast species Zygosaccharomyces bailii revealed by quantitative proteomics. Proteomics 12:2303–2318

    Article  CAS  Google Scholar 

  14. Mira NP, Teixeira MC, Sá-Correia I (2010) Adaptive response and tolerance to weak acids in Saccharomyces cerevisiae: a genome-wide view. OMICS 14:525–540

    Article  CAS  Google Scholar 

  15. Liu ZL (2011) Molecular mechanisms of yeast tolerance and in situ detoxification of lignocellulose hydrolysates. Appl Microbiol Biotechnol 90:809–825

    Article  CAS  Google Scholar 

  16. Sauer M, Porro D, Mattanovich D, Branduardi P (2008) Microbial production of organic acids: expanding the markets. Trends Biotechnol 26:100–108

    Article  CAS  Google Scholar 

  17. Branduardi P, Valli M, Brambilla L, Sauer M, Alberghina L, Porro D (2004) The yeast Zygosaccharomyces bailii: a new host for heterologous protein production, secretion and for metabolic engineering applications. FEMS Yeast Res 4:493–504

    Article  CAS  Google Scholar 

  18. Branduardi P, Fossati T, Sauer M, Pagani R, Mattanovich D, Porro D (2007) Biosynthesis of vitamin C by yeast leads to increased stress resistance. PLoS One 2:e1092

    Article  Google Scholar 

  19. Vigentini I, Brambilla L, Branduardi P, Merico A, Porro D, Compagno C (2005) Heterologous protein production in Zygosaccharomyces bailii: physiological effects and fermentative strategies. FEMS Yeast Res 5:647–652

    Article  CAS  Google Scholar 

  20. Camattari A, Bianchi MM, Branduardi P, Porro D, Brambilla L (2007) Induction by hypoxia of heterologous-protein production with the KlPDC1 promoter in yeasts. Appl Environ Microbiol 73:922–929

    Article  CAS  Google Scholar 

  21. Passolunghi S, Riboldi L, Dato L, Porro D, Branduardi P (2010) Cloning of the Zygosaccharomyces bailii GAS1 homologue and effect of cell wall engineering on protein secretory phenotype. Microb Cell Fact 9:7

    Article  Google Scholar 

  22. Dato L, Branduardi P, Passolunghi S, Cattaneo D, Riboldi L, Frascotti G, Valli M, Porro D (2010) Advances in molecular tools for the use of Zygosaccharomyces bailii as host for biotechnological productions and construction of the first auxotrophic mutant. FEMS Yeast Res 10:894–908

    Article  CAS  Google Scholar 

  23. Sauer M, Branduardi P, Valli M, Porro D (2004) Production of L-ascorbic acid by metabolically engineered Saccharomyces cerevisiae and Zygosaccharomyces bailii. Appl Environ Microbiol 70:6086–6091

    Article  CAS  Google Scholar 

  24. Mann C, Davis RW (1986) Structure and sequence of the centromeric DNA of chromosome 4 in Saccharomyces cerevisiae. Mol Cell Biol 6:241–245

    CAS  Google Scholar 

  25. Araki H, Oshima Y (1989) An autonomously replicating sequence of pSRI plasmid is effective in two yeast species, Zygosaccharomyces rouxii and Saccharomyces cerevisiae. J Mol Biol 207:757–769

    Article  CAS  Google Scholar 

  26. Gritz L, Davies J (1983) Plasmid-encoded hygromycin B resistance: the sequence of hygromycin B phosphotransferase gene and its expression in Escherichia coli and Saccharomyces cerevisiae. Gene 25:179–188

    Article  CAS  Google Scholar 

  27. Goldstein AL, McCusker JH (1999) Three new dominant drug resistance cassette for gene disruption in Saccharomyces cerevisiae. Yeast 15:13

    Google Scholar 

  28. Krügel H, Fiedler G, Smith C, Baumberg S (1993) Sequence and transcriptional analysis of the nourseothricin acetyltransferase-encoding gene nat1 from Streptomyces noursei. Gene 127:127–131

    Article  Google Scholar 

  29. Kikuchi Y (1983) Yeast plasmid requires a cis-acting locus and two plasmid proteins for its stable maintenance. Cell 34:7

    Article  Google Scholar 

  30. Toh-e A, Araki H, Utatsu I, Oshima Y (1984) Plasmids resembling 2-micrometers DNA in the osmotolerant yeasts Saccharomyces bailii and Saccharomyces bisporus. J Gen Microbiol 130:8

    Google Scholar 

  31. Utatsu I, Sakamoto S, Imura T, Tohe A (1987) Yeast plasmids resembling 2-μm DNA – regional similarities and diversities at the molecular level. J Bacteriol 169:5537–5545

    CAS  Google Scholar 

  32. Merico A, Rodrigues F, Côrte-Real M, Porro D, Ranzi B, Compagno C (2001) Isolation and sequence analysis of the gene encoding triose phosphate isomerase from Zygosaccharomyces bailii. Yeast 18:775–780

    Article  CAS  Google Scholar 

  33. Gonzalez C, Perdomo G, Tejera P, Brito N, Siverio JM (1999) One-step, PCR-mediated, gene disruption in the yeast Hansenula polymorpha. Yeast 15:1323–1329

    Article  CAS  Google Scholar 

  34. Kelly R, Miller SM, Kurtz MB, Kirsch DR (1987) Directed mutagenesis in Candida albicans – one-step gene disruption to isolate ura3 mutants. Mol Cell Biol 7:199–207

    CAS  Google Scholar 

  35. Guangtao Z, Hartl L, Schuster A, Polak S, Schmoll M, Wang T, Seidl V, Seiboth B (2009) Gene targeting in a nonhomologous end joining deficient Hypocrea jecorina. J Biotechnol 139:146–151

    Google Scholar 

  36. Mollapour M, Piper PW (2001) Targeted gene deletion in Zygosaccharomyces bailii. Yeast 18:173–186

    Article  CAS  Google Scholar 

  37. Rodrigues F, Ludovico P, Sousa MJ, Steensma HY, Corte-Real M, Leao C (2003) The spoilage yeast Zygosaccharomyces bailii forms mitotic spores: a screening method for haploidization. Appl Environ Microbiol 69:649–653

    Article  CAS  Google Scholar 

  38. Dato L, Sauer M, Passolunghi S, Porro D, Branduardi P (2008) Investigating the multibudded and binucleate phenotype of the yeast Zygosaccharomyces bailii growing on minimal medium. FEMS Yeast Res 8:906–915

    Article  CAS  Google Scholar 

  39. Rodrigues F, Zeeman AM, Alves C, Sousa MJ, Steensma HY, Corte-Real M, Leao C (2001) Construction of a genomic library of the food spoilage yeast Zygosaccharomyces bailii and isolation of the beta-isopropylmalate dehydrogenase gene (ZbLEU2). FEMS Yeast Res 1:67–71

    CAS  Google Scholar 

  40. Wach A, Brachat A, Pöhlmann R, Philippsen P (1994) New heterologous modules for classical or PCR-based gene disruptions in Saccharomyces cerevisiae. Yeast 10:1793–1808

    Article  CAS  Google Scholar 

  41. Popolo L, Vai M (1999) The Gas1 glycoprotein, a putative wall polymer cross-linker. Biochim Biophys Acta 1426:385–400

    Article  CAS  Google Scholar 

  42. Goldstein A, McCusker J (1999) Three new dominant drug resistance cassettes for gene disruption in Saccharomyces cerevisiae. Yeast 15:1541–1553

    Article  CAS  Google Scholar 

  43. Gietz RD, Sugino A (1988) New yeast-Escherichia coli shuttle vectors constructed with in vitro mutagenized yeast genes lacking 6-base pair restriction sites. Gene 74:527–534

    Google Scholar 

  44. Ogawa Y, Tatsumi H, Murakami S, Ishida Y, Murakami K, Masaki A, Kawabe H, Arimura H, Nakano E, Motai H (1990) Secretion of Aspergillus oryzae alkaline protease in an osmophilic yeast, Zygosaccharomyces rouxii. Agric Biol Chem 54:2521–2529

    Article  CAS  Google Scholar 

  45. Branduardi P (2002) Molecular cloning and sequence analysis of the Zygosaccharomyces bailii HIS3 gene encoding the imidazole glycerol phosphate dehydratase. Yeast 19:1165–1170

    Article  CAS  Google Scholar 

  46. Hoffman CS, Winston F (1987) A ten-minute DNA preparation from yeast efficiently releases autonomous plasmids for transformation of Escherichia coli. Gene 57:267–272

    Article  CAS  Google Scholar 

  47. Venturini M, Morrione A, Pisarra P, Martegani E, Vanoni M (1997) In Saccharomyces cerevisiae a short amino acid sequence facilitates excretion in the growth medium of periplasmic proteins. Mol Microbiol 23:997–1007

    Article  CAS  Google Scholar 

Download references

Acknowledgements

These works were partially supported by FAR (Fondo di Ateneo per la Ricerca) of the University of Milano-Bicocca to PB and DP.

Author information

Authors and Affiliations

  1. Department of Biotechnology and Biosciences, University of Milano-Bicocca, Piazza della Scienza, 2 – 20126, Milan, Italy

    Paola Branduardi, Laura Dato & Danilo Porro

Authors
  1. Paola Branduardi
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Laura Dato
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Danilo Porro
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Paola Branduardi .

Editor information

Editors and Affiliations

  1. Industrial Biotechnology, Chalmers University of Technology, Gothenburg, Sweden

    Valeria Mapelli

Rights and permissions

Reprints and permissions

Copyright information

© 2014 Springer Science+Business Media, LLC

About this protocol

Cite this protocol

Branduardi, P., Dato, L., Porro, D. (2014). Molecular Tools and Protocols for Engineering the Acid-Tolerant Yeast Zygosaccharomyces bailii as a Potential Cell Factory. In: Mapelli, V. (eds) Yeast Metabolic Engineering. Methods in Molecular Biology, vol 1152. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-0563-8_4

Download citation

  • DOI: https://doi.org/10.1007/978-1-4939-0563-8_4

  • Published:

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-0562-1

  • Online ISBN: 978-1-4939-0563-8

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation