Engineering corynebacterium glutamicum for the production of pyruvate

A Corynebacterium glutamicum strain with inactivated pyruvate dehydrogenase complex and a deletion of the gene encoding the pyruvate:quinone oxidoreductase produces about 19 mM L-valine, 28 mM L-alanine and about 55 mM pyruvate from 150 mM glucose. Based on this double mutant C. glutamicum ^ΔaceE ^Δpqo, we engineered C. glutamicum for efficient production of pyruvate from glucose by additional deletion of the ldhA gene encoding NAD ⁺-dependent L-lactate dehydrogenase (LdhA) and introduction of a attenuated variant of the acetohydroxyacid synthase ( ^ΔC-T IlvN). The latter modification abolished overflow metabolism towards L-valine and shifted the product spectrum to pyruvate production. In shake flasks, the resulting strain C. glutamicum ^ΔaceE ^Δpqo ^ΔldhA ^ΔC-T ilvN produced about 190 mM pyruvate with a YP/S of 1.36 mol per mol of glucose; however, it still secreted significant amounts of L-alanine. Additional deletion of genes encoding the transaminases AlaT and AvtA reduced L-alanine formation by about 50%. In fed-batch fermentations at high cell densities with adjusted oxygen supply during growth and production (0-5% dissolved oxygen), the newly constructed strain C. glutamicum ^ΔaceE ^Δpqo ^ΔldhA ^ΔC-T ilvN ^ΔalaT ^ΔavtA produced more than 500 mM pyruvate with a maximum yield of 0.97 mol per mole of glucose and a productivity of 0.92 mmol g(CDW) ^{-1 h-1} (i.e., 0.08 g g _(CDW) ^{-1 h-1}) in the production phase.