Optimization of lipid production with a genome-scale model of Yarrowia lipolytica.
BACKGROUND
Yarrowia lipolytica is a non-conventional yeast that is extensively investigated for its ability to excrete citrate or to accumulate large amounts of storage lipids, which is of great significance for single cell oil production. Both traits are thus of interest for basic research as well as for biotechnological applications but they typically occur simultaneously thus lowering the respective yields. Therefore, engineering of strains with high lipid content relies on novel concepts such as computational simulation to better understand the two competing processes and to eliminate citrate excretion.


RESULTS
Using a genome-scale model (GSM) of baker's yeast as a scaffold, we reconstructed the metabolic network of Y. lipolytica and optimized it for use in flux balance analysis (FBA), with the aim to simulate growth and lipid production phases of this yeast. We validated our model and found the predictions of the growth behavior of Y. lipolytica in excellent agreement with experimental data. Based on these data, we successfully designed a fed-batch strategy to avoid citrate excretion during the lipid production phase. Further analysis of the network suggested that the oxygen demand of Y. lipolytica is reduced upon induction of lipid synthesis. According to this finding we hypothesized that a reduced aeration rate might induce lipid accumulation. This prediction was indeed confirmed experimentally. In a fermentation combining these two strategies lipid content of the biomass was increased by 80%, and lipid yield was improved more than four-fold, compared to standard conditions.


CONCLUSIONS
Genome scale network reconstructions provide a powerful tool to predict the effects of genetic modifications and the metabolic response to environmental conditions. The high accuracy and the predictive value of a newly reconstructed GSM of Y. lipolytica to optimize growth conditions for lipid accumulation are demonstrated. Based on these findings, further strategies for engineering Y. lipolytica towards higher efficiency in single cell oil production are discussed.
Institute of Molecular Biosciences, BioTechMed Graz, University of Graz, Humboldtstrasse 50/II, 8010, Graz, Austria. martin.kavscek@uni-graz.at.
Martin
M
Kavšček
Institute of Molecular Biosciences, BioTechMed Graz, University of Graz, Humboldtstrasse 50/II, 8010, Graz, Austria. govindprasad.bhutada@uni-graz.at.
Govindprasad
G
Bhutada
Institute of Molecular Biology and Biochemistry, Medical University of Graz, Graz, Austria. t.madl@tum.de.|Omics Center Graz, BioTechMed Graz, Graz, Austria. t.madl@tum.de.|Center for Integrated Protein Science Munich (CIPSM) at the Department of Chemistry, Technische Universität München, Garching, Germany. t.madl@tum.de.|Institute of Structural Biology, Helmholtz Zentrum München, Neuherberg, Germany. t.madl@tum.de.
Tobias
T
Madl
Institute of Molecular Biosciences, BioTechMed Graz, University of Graz, Humboldtstrasse 50/II, 8010, Graz, Austria. klaus.natter@uni-graz.at.
Klaus
K
Natter
D057919:Biological Oxygen Demand Analysis; D018533:Biomass; D019343:Citric Acid / Q000378:metabolism; D005285:Fermentation; D005818:Genetic Engineering; D016681:Genome, Fungal; D050356:Lipid Metabolism / Q000235:genetics*; D064688:Metabolic Flux Analysis; D053858:Metabolic Networks and Pathways; D008962:Models, Theoretical; D009249:NADP / Q000096:biosynthesis; D009584:Nitrogen / Q000378:metabolism; D030781:Organisms, Genetically Modified / Q000378:metabolism; D010427:Pentose Phosphate Pathway; D025062:Yarrowia / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism
D019343:Citric Acid; D009249:NADP; D009584:Nitrogen