Quantitative Metabolomics and Instationary 13C-Metabolic Flux Analysis Reveals Impact of Recombinant Protein Production on Trehalose and Energy Metabolism in Pichia pastoris
| Autor: | Pau Ferrer, Joan Albiol, Marc Carnicer, Hugo Cueto Rojas, A. Wahl, Joel Jordà |
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| Předmět: |
Endocrinology
Diabetes and Metabolism lcsh:QR1-502 Rhizopus oryzae Xylose Biochemistry Article lcsh:Microbiology Pichia pastoris chemistry.chemical_compound Metabolomics Metabolic flux analysis Instationary 13C-metabolic flux analysis Lipase Molecular Biology trehalose recombinant protein production biology Trehalose biology.organism_classification Recombinant protein production LC-MS Citric acid cycle chemistry instationary 13C-metabolic flux analysis Metabolome biology.protein metabolome GC-MS |
| Zdroj: | Europe PubMed Central Dipòsit Digital de Documents de la UAB Universitat Autònoma de Barcelona Metabolites, Vol 4, Iss 2, Pp 281-299 (2014) Metabolites Metabolites; Volume 4; Issue 2; Pages: 281-299 Metabolites, 4 (2), 2014 |
| ISSN: | 2218-1989 |
| Popis: | Pichia pastoris has been recognized as an effective host for recombinant protein production. In this work, we combine metabolomics and instationary 13C metabolic flux analysis (INST 13C-MFA) using GC-MS and LC-MS/MS to evaluate the potential impact of the production of a Rhizopus oryzae lipase (Rol) on P. pastoris central carbon metabolism. Higher oxygen uptake and CO2 production rates and slightly reduced biomass yield suggest an increased energy demand for the producing strain. This observation is further confirmed by 13C-based metabolic flux analysis. In particular, the flux through the methanol oxidation pathway and the TCA cycle was increased in the Rol-producing strain compared to the reference strain. Next to changes in the flux distribution, significant variations in intracellular metabolite concentrations were observed. Most notably, the pools of trehalose, which is related to cellular stress response, and xylose, which is linked to methanol assimilation, were significantly increased in the recombinant strain. |
| Databáze: | OpenAIRE |
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