Glutamate is industrially produced by fermentation using Corynebacterium glutamicum. The key factor for efficient glutamate production by this microorganism has been considered to be a metabolic change at the 2-oxoglutarate dehydrogenase (ODH) branch point caused by a decrease in ODH activity under glutamate-overproducing conditions. However, this change would be insufficient because the ODH branch is merely the final branch in the glutamate biosynthetic pathway, and efficient glutamate production requires a balanced supply of acetyl-CoA and oxaloacetate (OAA), which are condensed to form a precursor of glutamate, namely, citrate. Therefore, there must be another (other) change(s) in metabolic flux. In this study, we demonstrated that a decrease in pyruvate dehydrogenase (PDH) activity catalyzes the conversion of pyruvate to acetylCoA. It is speculated that carbon flux from pyruvate to acetyl-CoA decreases under glutamate-overproducing conditions. Furthermore, an increase in pyruvate carboxylase (PC) activity, which catalyzes the reaction of pyruvate to OAA, is evident under glutamate-overproducing conditions, except under biotin-limited condition, which may lead to an increase in carbon flux from pyruvate to OAA. These data suggest that a novel metabolic change occurs at the pyruvate node, leading to a high yield of glutamate through adequate partitioning of the carbon flux.