Effect of gene disruptions of the TCA cycle on production of succinic acidin Saccharomyces cerevisiae

Succinate is the main taste component produced by yeasts during sake (Japan ese rice wine) fermentation. The pathway leading to accumulation of succina te was examined in liquid culture in the presence of a high concentration ( 15%) of glucose under aerobic and anaerobic conditions using a series of Sa ccharomyces cerevisiae strains in which various genes that encode the expre ssion of enzymes required in TCA cycle were disrupted. When cultured in YPD medium containing 15% glucose under aerobic conditions, the KGD1 (alpha-ke toglutarate dehydrogenase) gene disrupted mutant produced a lower level of succinate than the wild-type strain, while the SDH1 (succinate dehydrogenas e) gene-disrupted mutant produced an increased level of succinate. On the o ther hand,the FUM1 (fumarase) gene disrupted mutant produced significantly higher levels of fumarate but did not form malate at all. These results ind icate that succinate, fumarate and malate are mainly synthesized through th e TCA cycle (oxidative direction) even in the presence of glucose at a conc entration as high as 15%. When the growth condition was shifted from aerobi c to anaerobic, the increased level of succinate in SDH1 disruptants was no longer observed, whereas the decreased level of succinate in the KGD1 diru ptant was still observed. A double mutant of the two fumarate reductase iso zyme genes (OSM1 and FRDS) showed a succinate productivity of 50% as compar ed to the parent when cells were incubated in glucose buffered solution. Th ese results indicate that succinate could be synthesized through two pathwa ys, namely, alpha-ketoglutarate oxidation via the TCA cycle and fumarate re duction under anaerobic conditions. and fumarate reduction under anaerobic conditions.