Skip to main content
SpringerLink
Log in
Book cover

Recombinant Protein Production in Yeast pp 181–190Cite as

Optimising Pichia pastoris Induction

  • Protocol
  • First Online:

Part of the book series: Methods in Molecular Biology ((MIMB,volume 866))

Abstract

A common method for inducing the production of recombinant proteins in Pichia pastoris is through the use of methanol. However, the by-products of methanol metabolism are toxic to yeast cells and therefore its addition to recombinant cultures must be controlled and monitored throughout the process in order to maximise recombinant protein yields. Described here are online and off-line methods to monitor and control methanol addition to bench-top-scale bioreactors.

This is a preview of subscription content, 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   89.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD   119.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD   169.99
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

Learn about institutional subscriptions

Springer Nature is developing a new tool to find and evaluate Protocols. Learn more

References

  1. Ogata K, Nishikaw H, Ohsugi M (1969) A yeast capable of utilizing methanol. Agr Biol Chem 33:1519

    Article  CAS  Google Scholar 

  2. Wegner GH (1990) Emerging applications of the methylotrophic yeasts. FEMS Microbiol Rev 87:279–283

    Article  Google Scholar 

  3. Cereghino JL, Cregg JM (2000) Heterologous protein expression in the methylotrophic yeast Pichia pastoris. FEMS Microbiol Rev 24:45–66

    Article  PubMed  CAS  Google Scholar 

  4. Cos O, Ramon R, Montesinos JL, Valero F (2006) Operational strategies, monitoring and control of heterologous protein production in the methylotrophic yeast Pichia pastoris under different promoters: a review. Microb Cell Fact 5:17

    Article  PubMed  Google Scholar 

  5. Faber KN, Harder W, Ab G, Veenhuis M (1995) Review – methylotrophic yeasts as factories for the production of foreign proteins. Yeast 11:1331–1344

    Article  PubMed  CAS  Google Scholar 

  6. Inan M, Meagher MM (2001) Non-repressing carbon sources for alcohol oxidase (AOX1) promoter of Pichia pastoris. J Biosci Bioeng 92:585–589

    PubMed  CAS  Google Scholar 

  7. Minning S, Serrano A, Ferrer P, Sola C, Schmid RD, Valero F (2001) Optimization of the high-level production of Rhizopus oryzae lipase in Pichia pastoris. J Biotechnol 86:59–70

    Article  PubMed  CAS  Google Scholar 

  8. Guarna MM, Lesnicki GJ, Tam BM, Robinson J, Radziminski CZ, Hasenwinkle D, Boraston A, Jervis E, MacGillivray RTA, Turner RFB, Kilburn DG (1997) On-line monitoring and control of methanol concentration in shake-flask cultures of Pichia pastoris. Biotechnol Bioeng 56:279–286

    Article  PubMed  CAS  Google Scholar 

  9. Thorpe ED, d’Aanjou MC, Daugulis AJ (1999) Sorbitol as a non-repressing carbon source for fed-batch fermentation of recombinant Pichia pastoris. Biotechnol Lett 21:669–672

    Article  CAS  Google Scholar 

  10. d’Anjou MC, Daugulis AJ (2000) Mixed-feed exponential feeding for fed-batch culture of recombinant methylotrophic yeast. Biotechnol Lett 22:341–346

    Article  Google Scholar 

  11. Jungo C, Schenk J, Pasquier M, Marison IW, von Stockar U (2007) A quantitative analysis of the benefits of mixed feeds of sorbitol and methanol for the production of recombinant avidin with Pichia pastoris. J Biotechnol 131:57–66

    Article  PubMed  CAS  Google Scholar 

  12. Ramon R, Ferrer P, Valero F (2007) Sorbitol co-feeding reduces metabolic burden caused by the overexpression of a Rhizopus oryzae lipase in Pichia pastorism. J Biotechnol 130:39–46

    Article  PubMed  CAS  Google Scholar 

  13. Holmes WJ, Darby RAJ, Wilks MDB, Smith R, Bill RM (2009) Developing a scalable model of recombinant protein yield from Pichia pastoris: the influence of culture conditions, biomass and induction regime. Microb Cell Fact 8:35

    Article  PubMed  Google Scholar 

  14. Waterham HR, Digan ME, Koutz PJ, Lair SV, Cregg JM (1997) Isolation of the Pichia pastoris glyceraldehyde-3-phosphate dehydrogenase gene and regulation and use of its promoter. Gene 186:37–44

    Article  PubMed  CAS  Google Scholar 

  15. Kim SJ, Lee JA, Kim YH, Song BK (2009) Optimization of the functional expression of Coprinus cinereus peroxidase in Pichia pastoris by varying the host and promoter. J Microbiol Biotechnol 19:966–971

    Article  PubMed  CAS  Google Scholar 

  16. Resina D, Cos O, Ferrer P, Valero F (2005) Developing high cell density fed-batch cultivation strategies for heterologous protein production in Pichia pastoris using the nitrogen source-regulated FLD1 promoter. Biotechnol Bioeng 91:760–767

    Article  PubMed  CAS  Google Scholar 

  17. Pichia fermentation process guidelines. http://tools.invitrogen.com/content/sfs/manuals/pichiaferm_prot.pdf

  18. Nyblom M, Öberg F, Petersson-Lindkvist K, Findlay H, Wikström J, Karlsson A, Hansson O, Booth PJ, Bill RM, Neutze R, Hedfalk K (2007) Exceptional overproduction of a functional human membrane protein. Prottein Expr Purif 56:110–120

    Article  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. School of Life & Health Sciences and Aston Research Centre for Healthy Ageing, Aston University, Birmingham, UK

    Zharain Bawa

  2. Nuffi eld Department of Clinical Laboratory Sciences, John Radcliffe Hospital, University of Oxford, Oxford, UK

    Richard A. J. Darby

Authors
  1. Zharain Bawa
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Richard A. J. Darby
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Zharain Bawa .

Editor information

Editors and Affiliations

  1. School of Life and Health Sciences, Aston University, Aston Triangle, Birmingham, B4 7ET, United Kingdom

    Roslyn M. Bill

Rights and permissions

Reprints and permissions

Copyright information

© 2012 Springer Science+business Media, LLC

About this protocol

Cite this protocol

Bawa, Z., Darby, R.A.J. (2012). Optimising Pichia pastoris Induction. In: Bill, R. (eds) Recombinant Protein Production in Yeast. Methods in Molecular Biology, vol 866. Humana Press. https://doi.org/10.1007/978-1-61779-770-5_15

Download citation

  • DOI: https://doi.org/10.1007/978-1-61779-770-5_15

  • Published:

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-61779-769-9

  • Online ISBN: 978-1-61779-770-5

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation