TY - JOUR
T1 - A multi-level study of recombinant Pichia pastoris in different oxygen conditions
AU - Baumann, Kristin
AU - Carnicer, Marc
AU - Dragosits, Martin
AU - Graf, Alexandra B.
AU - Stadlmann, Johannes
AU - Jouhten, Paula
AU - Maaheimo, Hannu
AU - Gasser, Brigitte
AU - Albiol, Joan
AU - Mattanovich, Diethard
AU - Ferrer, Pau
N1 - Funding Information:
This work has been supported by the European Science Foundation (ESF, program EuroSCOPE), through the Complementary Actions Plan (Project BIO2005-23733-E) of the Spanish Ministry of Science and Education, the Integrated Action HU2005-0001 and project CTQ2007-60347/PPQ of the Spanish Ministry of Science and Education, the Austrian Science Fund (FWF), project no. I37-B03, and the Austrian Exchange Service. The Ministry of Innovation and Universities of the Generalitat de Catalunya gave support through contract grant 2005-SGR-00698 and 2009SGR-281, Xarxa de Referència en Biotecnologia and doctoral fellowship for K.B. We would like to thank Klaus Fortschegger for his support with real-time PCR analysis and Philipp Ternes for helpful comments on the sphingolipid metabolism.
PY - 2010/10/22
Y1 - 2010/10/22
N2 - Background: Yeasts are attractive expression platforms for many recombinant proteins, and there is evidence for an important interrelation between the protein secretion machinery and environmental stresses. While adaptive responses to such stresses are extensively studied in Saccharomyces cerevisiae, little is known about their impact on the physiology of Pichia pastoris. We have recently reported a beneficial effect of hypoxia on recombinant Fab secretion in P. pastoris chemostat cultivations. As a consequence, a systems biology approach was used to comprehensively identify cellular adaptations to low oxygen availability and the additional burden of protein production. Gene expression profiling was combined with proteomic analyses and the 13C isotope labelling based experimental determination of metabolic fluxes in the central carbon metabolism.Results: The physiological adaptation of P. pastoris to hypoxia showed distinct traits in relation to the model yeast S. cerevisiae. There was a positive correlation between the transcriptomic, proteomic and metabolic fluxes adaptation of P. pastoris core metabolism to hypoxia, yielding clear evidence of a strong transcriptional regulation component of key pathways such as glycolysis, pentose phosphate pathway and TCA cycle. In addition, the adaptation to reduced oxygen revealed important changes in lipid metabolism, stress responses, as well as protein folding and trafficking.Conclusions: This systems level study helped to understand the physiological adaptations of cellular mechanisms to low oxygen availability in a recombinant P. pastoris strain. Remarkably, the integration of data from three different levels allowed for the identification of differences in the regulation of the core metabolism between P. pastoris and S. cerevisiae. Detailed comparative analysis of the transcriptomic data also led to new insights into the gene expression profiles of several cellular processes that are not only susceptible to low oxygen concentrations, but might also contribute to enhanced protein secretion.
AB - Background: Yeasts are attractive expression platforms for many recombinant proteins, and there is evidence for an important interrelation between the protein secretion machinery and environmental stresses. While adaptive responses to such stresses are extensively studied in Saccharomyces cerevisiae, little is known about their impact on the physiology of Pichia pastoris. We have recently reported a beneficial effect of hypoxia on recombinant Fab secretion in P. pastoris chemostat cultivations. As a consequence, a systems biology approach was used to comprehensively identify cellular adaptations to low oxygen availability and the additional burden of protein production. Gene expression profiling was combined with proteomic analyses and the 13C isotope labelling based experimental determination of metabolic fluxes in the central carbon metabolism.Results: The physiological adaptation of P. pastoris to hypoxia showed distinct traits in relation to the model yeast S. cerevisiae. There was a positive correlation between the transcriptomic, proteomic and metabolic fluxes adaptation of P. pastoris core metabolism to hypoxia, yielding clear evidence of a strong transcriptional regulation component of key pathways such as glycolysis, pentose phosphate pathway and TCA cycle. In addition, the adaptation to reduced oxygen revealed important changes in lipid metabolism, stress responses, as well as protein folding and trafficking.Conclusions: This systems level study helped to understand the physiological adaptations of cellular mechanisms to low oxygen availability in a recombinant P. pastoris strain. Remarkably, the integration of data from three different levels allowed for the identification of differences in the regulation of the core metabolism between P. pastoris and S. cerevisiae. Detailed comparative analysis of the transcriptomic data also led to new insights into the gene expression profiles of several cellular processes that are not only susceptible to low oxygen concentrations, but might also contribute to enhanced protein secretion.
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U2 - 10.1186/1752-0509-4-141
DO - 10.1186/1752-0509-4-141
M3 - Article
C2 - 20969759
AN - SCOPUS:77958044182
SN - 1752-0509
VL - 4
JO - BMC Systems Biology
JF - BMC Systems Biology
M1 - 141
ER -