The effect of hypoxia on the lipidome of recombinant Pichia pastoris

Microbial Cell Factories

Research

Karlheinz Grillitsch¹  Günther Daum¹  Sergi Monforte²  Xavier García-Ortega²  Francisco Valero²  Pau Ferrer²  Núria Adelantado³  Pablo Tarazona⁴  Ivo Feussner⁵ 

[1] Austrian Centre of Industrial Biotechnology (ACIB), Graz, Austria;Institute of Biochemistry, Graz University of Technology, Petersgasse 12/II, 8010, Graz, Austria;Department of Chemical, Biological and Environmental Engineering, Escola d’Enginyeria, Universitat Autònoma de Barcelona, Bellaterra, 08193, Barcelona, Catalonia, Spain;Department of Chemical, Biological and Environmental Engineering, Escola d’Enginyeria, Universitat Autònoma de Barcelona, Bellaterra, 08193, Barcelona, Catalonia, Spain;Evonik Nutrition & Care GmbH, Hanau, Germany;Department of Plant Biochemistry, Albrecht-von-Haller-Institute for Plant Sciences, Georg-August-University, Justus-von-Liebig-Weg 11, 37077, Göttingen, Germany;Department of Plant Biochemistry, Albrecht-von-Haller-Institute for Plant Sciences, Georg-August-University, Justus-von-Liebig-Weg 11, 37077, Göttingen, Germany;Department of Plant Biochemistry, Göttingen Center for Molecular Biosciences (GZMB), Georg-August-University, Justus-von-Liebig-Weg 11, 37077, Göttingen, Germany;

关键词: Lipidomics;    Pichia pastoris;    Hypoxia;    Recombinant protein production;    Antibody fragment;    Protein secretion;    Unfolded protein response;   

DOI  :  10.1186/s12934-017-0699-4

 received in 2016-10-25, accepted in 2017-05-10,  发布年份 2017

来源: Springer

【 摘 要 】

BackgroundCultivation of recombinant Pichia pastoris (Komagataella sp.) under hypoxic conditions has a strong positive effect on specific productivity when the glycolytic GAP promoter is used for recombinant protein expression, mainly due to upregulation of glycolytic conditions. In addition, transcriptomic analyses of hypoxic P. pastoris pointed out important regulation of lipid metabolism and unfolded protein response (UPR). Notably, UPR that plays a role in the regulation of lipid metabolism, amino acid metabolism and protein secretion, was found to be upregulated under hypoxia.ResultsTo improve our understanding of the interplay between lipid metabolism, UPR and protein secretion, the lipidome of a P. pastoris strain producing an antibody fragment was studied under hypoxic conditions. Furthermore, lipid composition analyses were combined with previously available transcriptomic datasets to further understand the impact of hypoxia on lipid metabolism. Chemostat cultures operated under glucose-limiting conditions under normoxic and hypoxic conditions were analyzed in terms of intra/extracellular product distribution and lipid composition. Integrated analysis of lipidome and transcriptome datasets allowed us to demonstrate an important remodeling of the lipid metabolism under limited oxygen availability. Additionally, cells with reduced amounts of ergosterol through fluconazole treatment were also included in the study to observe the impact on protein secretion and its lipid composition.ConclusionsOur results show that cells adjust their membrane composition in response to oxygen limitation mainly by changing their sterol and sphingolipid composition. Although fluconazole treatment results a different lipidome profile than hypoxia, both conditions result in higher recombinant protein secretion levels.

【 授权许可】

CC BY   
© The Author(s) 2017

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