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Selection of the Optimal Yeast Host for the Synthesis of Recombinant Enzymes

  • Felix Bischoff
  • Martin Giersberg
  • Falko Matthes
  • Tobias Schwalenberg
  • Sebastian Worch
  • Gotthard KunzeEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1923)

Abstract

Yeasts, like Arxula adeninivorans, Hansenula polymorpha, Pichia pastoris, Debaryomyces hansenii, Debaryomyces polymorphus, Schwanniomyces occidentalis, Yarrowia lipolytica, and Saccharomyces cerevisiae are frequently used producers of recombinant enzymes, particularly when posttranslational modifications are mandatory to obtain full functionality. The wide-range transformation/expression platform presented in this chapter can be used to select the optimal yeast host for high-level synthesis of the desired enzyme with favorable biochemical properties. This platform is composed of a selection marker and up to four expression modules in a linearized cassette. Here we describe the protocols for the assembly as well as the transformation of yeast strains with the respective cassettes, screening of transformants, the isolation and biochemical characterization of the enzymes, and finally a simple fermentation strategy to achieve maximal yields of the chosen recombinant enzyme.

Key words

Wide-range expression module Gibson assembly Yeast transformation Transformant screening Recombinant enzymes Fermentation strategy 

Notes

Acknowledgment

The work was supported by grants from the German Federal Ministry of Economics and Technology (Bundesministerium für Wirtschaft und Technologie, BMWi) within its program “Zentrales Innovationsprogramm Mittelstand, ZIM” (Grant Numbers KF2131620MD2, KF2131628SA4).

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Felix Bischoff
    • 1
  • Martin Giersberg
    • 1
  • Falko Matthes
    • 1
  • Tobias Schwalenberg
    • 1
  • Sebastian Worch
    • 1
  • Gotthard Kunze
    • 1
    Email author
  1. 1.Leibniz Institute of Plant Genetics and Crop Plant Research (IPK)GaterslebenGermany

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