In vivo kinetics of primary metabolism in Saccharomyces cerevisiae studied through prolonged chemostat cultivation

Liang Wu, Mlawule R. Mashego, Angela M. Proell, Jacobus L. Vinke, Cor Ras, Jan Van Dam, Wouter A. Van Winden, Walter M. Van Gulik, Joseph J. Heijnen

Research output: Contribution to journalArticleScientificpeer-review

25 Citations (Scopus)

Abstract

In this study, prolonged chemostat cultivation is applied to investigate in vivo enzyme kinetics of Saccharomyces cerevisiae. S. cerevisiae was grown in carbon-limited aerobic chemostats for 70-95 generations, during which multiple steady states were observed, characterized by constant intracellular fluxes but significant changes in intracellular metabolite concentrations and enzyme capacities. We provide evidence for two relevant kinetic mechanisms for sustaining constant fluxes: in vivo near-equilibrium of reversible reactions and tight regulation of irreversible reactions by coordinated changes of metabolic effectors. Using linear-logarithmic kinetics, we illustrate that these multiple steady-state measurements provide linear constraints between elasticity parameters instead of their absolute values. Upon perturbation by a glucose pulse, glucose uptake and ethanol excretion in prolonged cultures were remarkably lower, compared to a reference culture perturbed at 10 generations. Metabolome measurements during the transient indicate that the differences might be due to a reduced ATP regeneration capacity in prolonged cultures.

Original languageEnglish
Pages (from-to)160-171
Number of pages12
JournalMetabolic Engineering
Volume8
Issue number2
DOIs
Publication statusPublished - Mar 2006

Keywords

  • Adaptation
  • Equilibrium
  • In vivo kinetics
  • Lin-log kinetics
  • Metabolome
  • Prolonged chemostat cultivation
  • S. cerevisiae
  • Stimulus-response experiment

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