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Open Access Highly Accessed Research

Genome, secretome and glucose transport highlight unique features of the protein production host Pichia pastoris

Diethard Mattanovich12*, Alexandra Graf12, Johannes Stadlmann3, Martin Dragosits1, Andreas Redl12, Michael Maurer12, Martin Kleinheinz1, Michael Sauer12, Friedrich Altmann3 and Brigitte Gasser1

Author Affiliations

1 Department of Biotechnology, University of Natural Resources and Applied Life Sciences, Vienna, Austria

2 School of Bioengineering, University of Applied Sciences FH-Campus Wien, Vienna, Austria

3 Department of Chemistry, University of Natural Resources and Applied Life Sciences, Vienna, Austria

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Microbial Cell Factories 2009, 8:29  doi:10.1186/1475-2859-8-29

Published: 2 June 2009

Abstract

Background

Pichia pastoris is widely used as a production platform for heterologous proteins and model organism for organelle proliferation. Without a published genome sequence available, strain and process development relied mainly on analogies to other, well studied yeasts like Saccharomyces cerevisiae.

Results

To investigate specific features of growth and protein secretion, we have sequenced the 9.4 Mb genome of the type strain DSMZ 70382 and analyzed the secretome and the sugar transporters. The computationally predicted secretome consists of 88 ORFs. When grown on glucose, only 20 proteins were actually secreted at detectable levels. These data highlight one major feature of P. pastoris, namely the low contamination of heterologous proteins with host cell protein, when applying glucose based expression systems. Putative sugar transporters were identified and compared to those of related yeast species. The genome comprises 2 homologs to S. cerevisiae low affinity transporters and 2 to high affinity transporters of other Crabtree negative yeasts. Contrary to other yeasts, P. pastoris possesses 4 H+/glycerol transporters.

Conclusion

This work highlights significant advantages of using the P. pastoris system with glucose based expression and fermentation strategies. As only few proteins and no proteases are actually secreted on glucose, it becomes evident that cell lysis is the relevant cause of proteolytic degradation of secreted proteins. The endowment with hexose transporters, dominantly of the high affinity type, limits glucose uptake rates and thus overflow metabolism as observed in S. cerevisiae. The presence of 4 genes for glycerol transporters explains the high specific growth rates on this substrate and underlines the suitability of a glycerol/glucose based fermentation strategy. Furthermore, we present an open access web based genome browser http://www.pichiagenome.org webcite.