Eeffects Of Glyoxylate Cycle On Succinic Acid Dual-phase Production In Corynebacterium Glutamicum ATCC13032

To investigate the effects of glyoxylate cycle on succinic acid two-phase production in C. glutamicum ATCC13032, we constructed a high yield of succinic acid strain by knocking out ldhA gene, and then knocked out aceA gene to block the glyoxylate cycle, through a two stage fermentation, we studied the differences of carbon flow distribution in anaerobic metabolic pathways. The changing of aerobic culture temperature can also affect the carbon source shunt. By determining the enzyme activity of four key enzymes, including isocitrate lyase (ICL), phosphoenolpyruvate carboxylase (PPC), pyruvate carboxylase (PYC), and malate dehydrogenase (MDH) at the end of aerobic fermentation, we analyzed the specific effects of glyoxylate cycle on metabolic node. Polymerase chain reaction (PCR) was applied to clone genes from the chromosomal DNA of C. glutamicum ATCC13032and plasmid pK18mobsacB was used to carry the deletion fragment. We examined succinic acid production in dual-phase fermentations using high performance liquid chromatography (HPLC). The concentration of succinate in ldhA aceA double mutant was49.6g/L, decreased18%comparing with ldhA mutant. At the end of aerobic stage, the enzyme activity of ICL decreased70%, accordingly that of PPC decreased68%. When the cells were cultured in37℃, succinic acid yield decreased significantly, the activity of ICL increased by1times while the key enzyme PYC was reduced by37%. The results showed by genetic manipulation or changing the culture conditions, the flux of glyoxylate cycle will affect the activity of other metabolic pathways, with changing the carbon source shunt.