CARBON FLUX DISTRIBUTIONS AT THE GLUCOSE-6-PHOSPHATE BRANCH POINT IN CORYNEBACTERIUM-GLUTAMICUM DURING LYSINE OVERPRODUCTION

Analyses indicate that the lysine yield in Corynebacterium glutamicum is limited by suboptimal flux partitioning at either the glucose 6-pho sphate (Glc6P), phosphoenolpyruvate (PEP), or pyruvate (Pyr) branch po ints (or a combination thereof), which results in disproportionate pro duction of the required lysine precursors. Suboptimal flux partitionin g at a metabolic branch point can result from the inadequate enzyme ac tivity of a subordinate branch (a weakly rigid branch point) or from t he active feedback regulation of the flux partitioning (strongly rigid branch point). In order to examine branch-point characteristics, we h ave utilized metabolite balances, constrained by biochemistry, to esti mate flux distributions in the primary metabolic network of C. glutami cum from measured, extracellular, metabolite accumulation rates. These flux distributions, in combination with local metabolic perturbations , were used to infer branch-point characteristics. This study examines the flexibility of the Glc6P branch point, which could limit lysine y ield via inadequate NADPH production, by perturbations induced from at tenuation of Glc6P isomerase activity (first branch-point enzyme of gl ycolysis) and by fermentation on gluconate, which effectively bypasses the Glc6P branch point. Results from the analyses of these perturbati ons, as well as the flux distributions at the Glc6P branch point durin g the different phases of a control lysine fermentation, indicate that the Glc6P branch point is neither weakly nor strongly rigid, so that limitations in lysine yield must result from rigidity at either the PE P or Pyr branch point.