A simple strategy to effectively produce D-lactate in crude glycerol-utilizing Escherichia coli

Background: Fed-batch fermentation has been conventionally implemented for the production of lactic acid with a high titer and high productivity. However, its operation needs a complicated control which increases the production cost. Results: This issue was addressed by simplifying the production scheme. Escherichia coli was manipulated for its glycerol dissimilation and D-lactate synthesis pathways and then subjected to adaptive evolution under high crude glycerol. Batch fermentation in the two-stage mode was performed by controlling the dissolved oxygen (DO), and the evolved strain deprived of poxB enabled production of 100 g/L D-lactate with productivity of 1.85 g/L/h. To increase productivity, the producer strain was further evolved to improve its growth rate on crude glycerol. The fermentation was performed to undergo the aerobic growth with low substrate, followed by the anaerobic production with high substrate. Moreover, the intracellular redox of the strain was balanced by fulfillment of the anaerobic respiratory chain with nitrate reduction. Without controlling the DO, the microbial fermentation resulted in the homofermentative production of D-lactate (ca. 0.97 g/g) with a titer of 115 g/L and productivity of 3.29 g/L/h. Conclusions: The proposed fermentation strategy achieves the highest yield based on crude glycerol and a comparable titer and productivity as compared to the approach by fed-batch fermentation. It holds a promise to sustain the continued development of the crude glycerol-based biorefinery.