Rapid perturbation experiments are highly relevant to elaborate the in vivo kinetics for mathematical models of metabolism, which are needed for selecting gene targets for metabolic engineering. Perturbations were applied to chemostat-cultivated biomass (D=0.05 h(-1), aerobic glucose/ethanol-limited) using the BioScope of Saccharomyces cerevisiae CEN. PK 113-7D over time span of 90 and 180 s. The availability of the external electron acceptor oxygen was decreased from fully aerobic to anaerobic conditions. It was observed that the changes in metabolome response under these conditions were limited to the pyruvate node. Acetaldehyde supply was used as an extra external electron acceptor during glucose perturbation under fully aerobic conditions. This had a strong effect on the metabolome dynamics and resulted in a significantly higher initial glycolytic flux. Dynamic response of the adenine nucleotides indicated that their behavior is not dictated by the glycolytic flux but is much more coupled to the cytosolic NADH/NAD(+) ratio through the equilibrium pool of fructose 1,6-bisphosphate and 2/3-phosphoglycerate. Also, the electron donor availability (glucose) was decreased. This did not result in significant changes in the concentrations of the glycolytic and tricarboxylic acid cycle metabolites, whereas the adenine nucleotides, especially ADP and AMP, showed the opposite response to that observed in a glucose pulse experiment. Surprisingly, trehalose was not mobilized in the time frame of 180 s.

• PMID: 17311584
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Journal Article | Research Support, Non-U.S. Gov't

Authors

Mashego MR, van Gulik WM, Heijnen JJ

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