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13 C-Metabolic flux analysis ( 13 C-MFA) traditionally assumes that kinetic isotope effects from isotopically labeled compounds do not appreciably alter cellular growth or metabolism, despite indications that some biochemical reactions can be non-negligibly impacted. Here, populations of  Escherichia coli  were adaptively evolved for ~1000 generations on uniformly labeled  13 C-glucose, a commonly used isotope for  13 C-MFA. Phenotypic characterization of these evolved strains revealed ~40% increases in growth rate, with no significant difference in fitness when grown on either labeled ( 13 C) or unlabeled ( 12 C) glucose. The evolved strains displayed decreased biomass yields, increased glucose and oxygen uptake, and increased acetate production, mimicking what is observed after adaptive evolution on unlabeled glucose. Furthermore, full genome re-sequencing revealed that the key genetic changes underlying these phenotypic alterations were essentially the same as those acquired during adaptive evolution on unlabeled glucose. Additionally, glucose competition experiments demonstrated that the wild-type exhibits no isotopic preference for unlabeled glucose, and the evolved strains have no preference for labeled glucose. Overall, the results of this study indicate that there are no significant differences between  12 C and  13 C-glucose as a carbon source for  E .  coli  growth.

Evolution of E. coli on [U-13C]Glucose Reveals a Negligible Isotopic Influence on Metabolism and Physiology