Metabolic Engineering Of Corynebacterium Glutamicum For Efficient Production Of(R)-Acetoin

In this research,acetate synthesis pathway gene cat and pqo,glycerin synthesis pathway gene nagD and acetoin synthesis pathway gene alsSD operon were metabolically modified in Corynebacterium glutamicum CGF2 to realize a high production of(R)-acetoin,combining with optimization of fermentation condition.Firstly,the yield of acetoin increased 10.34%,and the production of acetate reduced 59.78% by abolishing cat in flask cultivation.Further abolishing pqo had no effect on both acetoin and acetate.However,the fed-batch production of acetoin by cat mutation strain was not as good as the control strain in 5 L batch fermenter.Above result showed that the deletion of cat had a disadvantage on acetoin production under our fed-batch fermentation condition.Secondly,the yield of acetoin increased 17.24%,and the production of glycerin reduced 48.79% by abolishing nagD in flask cultivation.In fed-batch cultivation,the production of acetoin increased to 72.50 g/L comparing with the control strain,which accumulated 54.23 g/L acetoin.Above result showed that the deletion of nagD efficiently interrupted the synthesis of glycerin.As a result,more carbon was used in the synthesis of acetoin.Thirdly,through the insertion of alsSD operon in the genome of nagD mutation strain and the introducing of over-expression vector of alsSD operon without IPTG induction,the yield of acetoin increased 5.88% in flask cultivation.In fed-batch cultivation,the production of acetoin increased to 80.50 g/L.Above result showed that enhanceing the expressing of alsSD operon could result in more accumulation of acetoin.Finally,we studied the influence of dissolved oxygen and organic nitrogen on acetoin,and found that the highest yield of acetoin was acquired in high dissolved oxygen and 50 g/L corn steep liquor as organic nitrogen.Under optimizing fed-batch fermentation condition,the recombination strain achieved 96.2 g/L(R)-acetion(purity 95%)by the productivity of 1.30 g/(Lh).And the yield of acetion was 0.36 g/g glucose,which reached 73.8% of the maxium theoretic yield.