The Research Of Cofactor Metabolic Regulation For Succinate Production By Escherichia Coli

Redox cofactor NADH is able to be a driving force for more effective productionof many bio-based products. ATP is the main energy intracellular for maintaining cellgrowth and vitality. This paper focuses on improving susccinate production inEscherichia coli based on reduce cofactor (NADH) regeneration, energy supply.Succinate yield was increased from1.12mol/mol glucose to1.56mol/mol glucose bydisturbance of PP pathway, enhance of the conversion bettween NADH and NADPHand deletion of competive pathway.In E. coli, PP patheway is the main sources of NADPH and NADPH can beconverted into NADH by transhydrogenase SthA. Thus improving the flux of PPpathway can provides cofactor NADH for succinate production in anaerobic condition.In order to improve the carbon flux of PP pathwy, the enzyme mutation genes zwf(243)and gnd(361) from Corynebacterium glutamicum were integrated into E. coli ZTKgenome, and the yield of succinate increased from1.12to1.26mol/mol glucose. Wecontinued to overexprees three genes belonging to PP pathway, and the succinate yieldfurther increased to1.3mol/mol glucose. There is soluble transhydrogenase (SthA) inE. coli, which catalyzes the transfer from NADPH to NADH. Overexpression sthA geneincreased succinate yield to1.44mol/mol glucose,while the cell growth rate andsuccinte productivity reduce significantly.We replaced native ppc gene with A. succinogenes pck gene and overexpressednative pck for regenerating ATP. Fermentation results indicated that the ATP supplyrecovered cell growth rate and succinate productivity, meanwhile the succinate yieldwas maintained at1.42mol/mol glucose. Contrary to previous report, the deletion ofack-pta just decreased the fitness of cell growth instead of abolishing cell growth inanaerobic condition and succinate yield increased to1.56mol/mol glucose, andmaximum productivity is2.44mmol/L/h.