Mechanisms of metabolic robustness in Escherichia coli in response to altered levels of glucose-6-phosphate dehydrogenase activity. A system-level investigation

Cécile Nicolas, Brice Enjalbert, Fabien Létisse, Stéphane Massou & Jean-Charles Portais

Laboratory of BioSystems & Chemical Engineering, UMR5504, UMR792, CNRS, INRA, INSA – LISBP/INSA 135, avenue de Rangueil, F-31077 Toulouse cedex 4, France

Microorganisms have an efficient capacity for adapting their metabolism in response to genetic or environmental changes. Therefore, understanding metabolic robustness has become an emergent issue for both basic and applied purposes. The application of ¹³C-Metabolic flux analysis (¹³C-fluxomics) to mutants lacking key metabolic enzymes has provided direct evidence that part of this robustness originates from the network organization of metabolic systems, where the interplay between all available biochemical reactions provides alternative mechanisms for compensating the perturbations. Accordingly, we have recently used ¹³C-fluxomics to show that Escherichia coli can efficiently adapt its metabolism to both the lack and 15-fold increase of glucose-6-phosphate dehydrogenase (G6PdH) activity (Nicolas et al., FEBS lett., 2007), an enzyme that plays a key role in the partitioning of carbon between glycolysis and the Pentose-Phosphate pathway (PPP). To get further comprehensive understanding of the mechanisms underlying such metabolic behaviour, we have investigated in more details the response of E. coli to quantitative alterations in G6PdH. A set of mutants with fixed levels of G6PdH activity was generated, with levels ranging from 2% to 1500% the WT strain activity. Combination of transcriptomics, metabolomics and fluxomics approaches were carried out on each mutant to provide detailed information on the nature, origin, and extent of the compensatory mechanisms. Because the activity of a single enzyme is tuned at different levels in the mutants, this investigation provides a situation that mimics gene-level regulation of metabolism. The results show that both hierarchical and metabolic mechanisms are involved in the control of carbon fluxes.