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Pichia surface display: display of proteins on the surface of glycoengineered Pichia pastoris strains

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Abstract

Expression of proteins on the surface of yeasts has a wide range of applications in biotechnology, such as directed evolution of proteins for increased affinity and thermal stability, screening of antibody libraries, epitope mapping, and use as whole-cell biocatalysts. However, hyperglycosylation can interfere with overall protein accessibility on the surface. Therefore, the less elaborate hyperglycosylation in wild type Pichia pastoris and the availability of glycoengineered strains make this yeast an excellent alternative for surface display of glycoproteins. Here, we report the implementation of the well-established a-agglutinin-based yeast surface display technology in P. pastoris. Four heterologous proteins were expressed on the surface of a wild type and a glycoengineered strain. Surface display levels were monitored by Western blot, immunofluorescence microscopy, and FACS analysis. The availability of glycoengineered strains makes P. pastoris an excellent alternative for surface display of glycoproteins and paves the way for new applications.

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Abbreviations

ER:

Endoplasmic reticulum

PSD:

Pichia surface display

YSD:

Yeast surface display

hEPO:

Human erythropoietin

hGal-1:

Human galectin-1

mIFN-β:

Mouse interferon beta

mIFN-γ:

Mouse interferon gamma

FACS:

Fluorescence activated cell sorting

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Acknowledgements

Pieter Jacobs is supported by a PhD grant from the Institute for the Promotion of Innovation through Science and Technology in Flanders (IWT-Vlaanderen). This research was funded by Research Corporation Technologies (Tucson, AZ). The continuous support and encouragement of Dr. Bennet Cohen is very much appreciated. Eef Parthoens and Wies Deckers are acknowledged for their help with the immunofluorescence microscopy. We thank Dr. Amin Bredan for editing the manuscript.

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Authors and Affiliations

  1. Department for Molecular Biomedical Research, Unit for Fundamental and Applied Molecular Biology, VIB, Technologiepark 927, 9052, Ghent, Belgium

    Pieter P. Jacobs, Stefan Ryckaert, Steven Geysens, Kristof De Vusser & Roland Contreras

  2. Department of Molecular Biology, Ghent University, Technologiepark 927, 9052, Ghent, Belgium

    Pieter P. Jacobs, Stefan Ryckaert, Steven Geysens, Kristof De Vusser & Roland Contreras

  3. Department for Molecular Biomedical Research, Unit for Molecular Glycobiology, VIB, Technologiepark 927, 9052, Ghent, Belgium

    Nico Callewaert

  4. Laboratory for Protein Biochemistry and Biomolecular Engineering, Department of Biochemistry, Physiology and Microbiology, Ghent University, K.L.-Ledeganckstraat 35, 9000, Ghent, Belgium

    Nico Callewaert

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  1. Pieter P. Jacobs
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  2. Stefan Ryckaert
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  3. Steven Geysens
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  4. Kristof De Vusser
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  5. Nico Callewaert
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  6. Roland Contreras
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Correspondence to Nico Callewaert.

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Jacobs, P.P., Ryckaert, S., Geysens, S. et al. Pichia surface display: display of proteins on the surface of glycoengineered Pichia pastoris strains. Biotechnol Lett 30, 2173–2181 (2008). https://doi.org/10.1007/s10529-008-9807-1

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  • DOI: https://doi.org/10.1007/s10529-008-9807-1

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