Metabolic Engineering Of Corynebacterium Glutamicum For 2,3-Butaneniol And Acetoin Production

2,3-butanediol and acetoin are important platform chemicals,which are widely used for food, pharmaceutical engineering, energy and other industries. Corynebacterium glutamicum is non-pathogenic and one of the typical gram-positive bacteria. In sufficient oxygen condition, it can reach a high cell density with fast substrate consumption and ideal productivity. In this paper, C. glutamicum ATCC 13032 wild-type WT and SAT7 with gene ldhA, pta and ack knocked out were used as the original strains. By introducing the synthetic pathways, the strains could produce 2,3-butanediol and acetoin.Gene butA encoding meso-2,3-butanediol dehydrogenase in C. glutamicum was deleted for identifing the exogenous gene performance, which constructed initial strains WT?butA / SAT7?butA. Operon alsSD from B. subtilis 168 was introduced for acetoin production which constructed strains CGF3-10. Gene bdhA encoding D-2,3- butanediol dehydrogenase from B. Subtilis was introduced to construct strains CGF11-18. Gene budC encoding meso-2,3- butanediol dehydrogenase from Klebsiella pneumoniae was introduced to construct strains CGF19-26. Gene butA in C. glutamicum was overexpressed for meso-2,3- butanediol production.The intracellular NAD+/NADH ratio was regulated through cofactor engineering. The gene nox from Lactobacillus brevis CICC 6239 encodes NADH oxidase, which transfers excessive NADH to NAD+. It can provide sufficient oxidizing force for acetoin production. The gene udhA from E. coli W1485 encodes synthetic transhydrogenase, which can transfers NADPH produced in pentose phosphate pathway into NADH to increase 2,3-butanediol production.With the optimized shaker speed 150 rpm and 20 g/L initial glucose concentration, three different media were tested for enhanced acetoin / 2,3-butanediol production. The metabolic products were examined by HPLC. The results below were acquired: Strain CGF5 produced 5.94 g/L acetoin in CGIII medium after 50 hours with a yield of 60%. Strain CGF18 produced 4.65 g/L D-2,3-butaneniol in CGXII medium after 30 hours with a yield of 46% and a productivity of 0.15 g/Lh. Strain CGF28 produced 4.73 g/L meso-2,3-butaneniol in CGXIIY medium after 30 hours with a yield of 47% and a productivity of 0.16 g/Lh.It proved that C. glutamicum with metabolic engineering was able to produce 2,3-butaneniol and acetoin. Gene bdhA plays an important role in the synthesis of D-2,3- butanediol in C. glutamicum. The meso-2,3-butanediol dehydrogenase coded by butA has a better performance than that coded by budC.