Stimulation of acetoin production in metabolically engineered Lactococcus lactis by increasing ATP demand.

Abstract

Having a sufficient supply of energy, usually in the form of ATP, is essential for all living organisms. In this study, however, we demonstrate that it can be beneficial to reduce ATP availability when the objective is microbial production. By introducing the ATP hydrolyzing F1-ATPase into a Lactococcus lactis strain engineered into producing acetoin, we show that production titer and yield both can be increased. At high F1-ATPase expression level, the acetoin production yield could be increased by 10%; however, because of the negative effect that the F1-ATPase had on biomass yield and growth, this increase was at the cost of volumetric productivity. By lowering the expression level of the F1-ATPase, both the volumetric productivity and the final yield could be increased by 5% compared to the reference strain not overexpressing the F1-ATPase, and in batch fermentation, it was possible to convert 176mM (32g/L) of glucose into 146.5mM (12.9g/L) acetoin with a yield of 83% of the theoretical maximum. To further demonstrate the potential of the cell factory developed, we complemented it with the lactose plasmid pLP712, which allowed for growth and acetoin production from a dairy waste stream, deproteinized whey. Using this cheap and renewable feedstock, efficient acetoin production with a titer of 157mM (14g/L) acetoin was accomplished.