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  • Changes in substrate availability in Escherichia coli lead to rapid metabolite, flux and growth rate responses.

Changes in substrate availability in Escherichia coli lead to rapid metabolite, flux and growth rate responses.

Metabolic engineering (2013-02-02)
Hilal Taymaz-Nikerel, Marjan De Mey, Gino Baart, Jo Maertens, Joseph J Heijnen, Walter van Gulik
ABSTRACT

The interactions between the intracellular metabolome, fluxome and growth rate of Escherichia coli after sudden glycolytic/gluconeogenic substrate shifts are studied based on pulses of different substrates to an aerobic glucose-limited steady-state (dilution rate=0.1h(-1)). After each added glycolytic (glucose) and gluconeogenic (pyruvate and succinate) substrate pulse, no by-products were secreted and a pseudo steady state in flux and metabolites was achieved in about 30-40s. In the pulse experiments a large oxygen uptake capacity of the cells was observed. The in vivo dynamic responses showed massive reorganization and flexibility (1/100-14-fold change) of extra/intracellular metabolic fluxes, matching with large changes in the concentrations of intracellular metabolites, including reversal of reaction rate for pseudo/near equilibrium reactions. The coupling of metabolome and fluxome could be described by Q-linear kinetics. Remarkably, the three different substrate pulses resulted in a very similar increase in growth rate (0.13-0.3h(-1)). Data analysis showed that there must exist as yet unknown mechanisms which couple the protein synthesis rate to changes in central metabolites.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Succinic acid, SAJ special grade, ≥99.5%
Supelco
Succinic acid, certified reference material, TraceCERT®, Manufactured by: Sigma-Aldrich Production GmbH, Switzerland
Sigma-Aldrich
Pyruvic acid, natural, ≥95%, FG
Sigma-Aldrich
Pyruvic acid, 95%
Sigma-Aldrich
Pyruvic acid, 98%
Sigma-Aldrich
Succinic acid, ACS reagent, ≥99.0%
Sigma-Aldrich
Pyruvic acid, ≥97%, FG
Sigma-Aldrich
Succinic acid, purum p.a., ≥99.0% (T)
Sigma-Aldrich
Succinic acid, BioXtra, BioRenewable, ≥99.0%
Sigma-Aldrich
Succinic acid, ReagentPlus®, BioRenewable, ≥99.0%
Sigma-Aldrich
Succinic acid, puriss. p.a., ACS reagent, ≥99.5% (T)
Sigma-Aldrich
Succinic acid, BioReagent, BioRenewable, suitable for cell culture, suitable for insect cell culture