13C-Labeled metabolic flux analysis of a fed-batch culture of elutriated Saccharomyces cerevisiae

被引:24
作者
Costenoble, Roeland
Mueller, Dirk
Barl, Timo
van Gulik, Walter M.
van Winden, Wouter A.
Reuss, Matthias
Heijnen, Joseph J.
机构
[1] Delft Univ Technol, Dept Biotechnol, NL-2628 BC Delft, Netherlands
[2] Univ Stuttgart, Inst Biochem Engn, D-7000 Stuttgart, Germany
关键词
synchronized culture; C-13-labeling experiment; Saccharomyces cerevisiae; LC-MS analysis; metabolic flux analysis; cell division cycle;
D O I
10.1111/j.1567-1364.2006.00199.x
中图分类号
Q81 [生物工程学(生物技术)]; Q93 [微生物学];
学科分类号
071005 ; 0836 ; 090102 ; 100705 ;
摘要
This study addresses the question of whether observable changes in fluxes in the primary carbon metabolism of Saccharomyces cerevisiae occur between the different phases of the cell division cycle. To detect such changes by metabolic flux analysis, a C-13-labeling experiment was performed with a fed-batch culture inoculated with a partially synchronized cell population obtained through centrifugal elutriation. Such a culture exhibits dynamic changes in the fractions of cells in different cell cycle phases over time. The mass isotopomer distributions of free intracellular metabolites in central carbon metabolism were measured by liquid chromatography-mass spectrometry. For four time points during the culture, these distributions were used to obtain the best estimates for the metabolic fluxes. The obtained flux fits suggested that the optimally fitted split ratio for the pentose phosphate pathway changed by almost a factor of 2 up and down around a value of 0.27 during the experiment. Statistical analysis revealed that some of the fitted flux distributions for different time points were significantly different from each other, indicating that cell cycle-dependent variations in cytosolic metabolic fluxes indeed occurred.
引用
收藏
页码:511 / 526
页数:16
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