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Manuscript title Metabolome, transcriptome and metabolic flux analysis of arabinose fermentation by engineered Saccharomyces cerevisiae.
PubMed ID 20816840
Journal Metabolic Engineering
Year 2010
Authors H. Wouter Wisselink, Chiara Cipollina, Bart Oud, BarbaraCrimi, Joseph J. Heijnen, Jack T. Pronk, Antonius J.A.van Maris
Affiliations Department of Biotechnology, Delft University of Technology, Julianalaan 67, 2628 BC Delft, The Netherlands.
Keywords Saccharomyces cerevisiae, Evolutionary engineering, Arabinose, Transcriptomics, Metabolomics, Metabolic flux analysis
Full text article Downloadarticle Wisselink_2010.pdf
Project name not specified

Experiment Description info

Organism Saccharomyces cerevisiae
Strain IMS0001 and IMS0002
Data type flux measurements
Data units normalised to the specific glucose uptake rate
Execution date not specified

Experimental Details info

Temperature (0C) 30
pH 5.0
Carbon source glucose and arabinose
Culture mode chemostat
Process condition anaerobic
Dilution rate (h-1) 0.03
Working volume (L) 1.0
Biomass concentration (g/L) Biomass yield (g g−1) = 0.066±0.001 (IMS0001, Glucose), 0.072±0.003 (IMS0002, Glucose) and 0.075±0.001 (IMS0002, Arabinose)
Medium composition

Cultures were performed in MY supplemented with 0.01 g l−1 ergosterol and 0.42 g l−1 Tween 80 dissolved in ethanol, silicon antifoam, vitamin solution and trace elements [7], and 20 g l−1 glucose (MYG) or arabinose (MYA).

General protocol information Flux analysis method: 13C constrained MFA

Platform: LC-MS

Methods description - Notes

Intracellular metabolic fluxes were calculated through metabolic flux balancing using a compartmented stoichiometric model that was previously developed for describing aerobic growth on glucose [1,2] and modified to describe anaerobic growth on glucose and arabinose. Reactions required for production and excretion of organic acids and for transport of oleate and palmitoleate (required for lipid biosynthesis) and the fumarate reductase reaction (TCA reductive branch) were introduced [1, 3]. Four reactions necessary for arabinose utilization were added to the model: arabinose transport, arabinose isomerase, l-ribulokinase and l-ribulose-5-phosphate-4-epimerase. The assumed macromolecular biomass composition was 45% (w/w) protein, 40.7% (w/w) polysaccharides, 6.3% (w/w) RNA, 0.4% (w/w) DNA, 2.9% (w/w) lipid, 2.5% (w/w) metals, 1.2% (w/w) water, 0.8% (w/w) phosphate and 0.2% (w/w) sulfate [2, 4]. Measured protein contents of IMS0001 and IMS0002 strains agreed with the assumed value (data not shown). Dedicated software (SPAD it, Nijmegen, The Netherlands) was used for metabolic flux balancing, the theory and practice of which have been thoroughly described elsewhere (4, 5, 6) and will not be repeated here. For each growth condition, specific rates of growth, substrate consumption and carbon-dioxide, glycerol, acetate, pyruvate, lactate and succinate production were measured. For the conversion of fluxes into mmoles per Cmole of biomass, a biomass C-molar weight of 25.87 g mol−1 was used [2, 4]. The number of measured rates was sufficient to result in an over-determined system, thus enabling data reconciliation. In all cases the degree of redundancy equaled 2. --------------------------------------------References---------------------------------------
[1] P. Daran-Lapujade, M.L. Jansen, J.M. Daran, W. van Gulik, J.H. de Winde, J.T. Pronk. J. Biol. Chem., 279 (2004), pp. 9125-9138. http://doi.org/dhhj2k
[2] Lange, H.C., 2002. Quantitative physiology of S. cerevisiae using metabolic network analysis. Ph.D. Thesis, Delft University of Technology, Department of Biotechnology, the Netherlands.
[3] K. Enomoto, Y. Arikawa, H. Muratsubaki. FEMS Microbiol. Lett., 215 (2002), pp. 103-108. http://doi.org/c6x6g6
[4] T.L. Nissen, U. Schulze, J. Nielsen, J. Villadsen.Microbiology, 143 (1997), pp. 203-218. http://doi.org/dwvx2q
[5] Vallino, J.J., Stephanopoulos, G., 1990. In: S.K. Skidar, M. Bier, P. Todd (Eds.), Frontiers in Bioprocess. CRC Press, Boca Raton, FL, pp. 205–219.
[6] W.M. van Gulik, J.J. Heijnen. Biotechnol. Bioeng., 48 (1995), pp. 681-698. http://doi.org/fsv4g6

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Entered by Administrator KiMoSysFirst name: Administrator
Affiliation: INESC-ID/IST
Homepage: http://kdbio.inesc-id.pt/kimosys
Interests: mathematical modeling, accessible data, use of data

Created 2018-08-28 16:39:16 UTC

Updated 2018-08-28 16:39:16 UTC

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