The thermodynamic and kinetic properties of the glycolytic pathway have been analyzed in order to characterize the ultimate limits for the metabolite concentrations that may be evolutionarily reached in response to a selective pressure in the direction of increased glycolytic reaction flux. The results indicate that the chemical potential for conversion of glucose into pyruvate at presently observed levels of these two metabolites (and of NADH, NAD+, ATP, ADP and inorganic phosphate) is high enough to allow ultimately for an accumulation of fructose 1,6-bisphosphate at concentrations which may be considered as indefinitely high from a practical point of view. This means that non-kinetic factors such as metabolite solubility put a definite limit to the glycolytic reaction flux that may be evolutionarily reached.
With the reasonable assumption that evolution eventually may raise the concentration of fructose 1,6-bisphosphate minimally to the same level as that presently attained by glucose, analytical evidence is presented to show that catalytic improvement of enzymes in the glycolytic pathway ultimately may result in glycerone phosphate levels that are at least two orders of magnitude higher than those presently observed. This argues strongly against the idea that triose-phosphate isomerase represents a perfectly evolved catalyst in the sense that it should have reached the end of its evolutionary development.