Abstract:[Objective] D-pantothenic acid (vitamin B5) is a water-soluble vitamin widely used in feed, food, chemical, pharmaceutical and other industries. At present, vitamin B5 is mainly produced by chemical-enzymatic methods, while this production process generates a large amount of cyanide-containing wastewater. Cell factories have the characteristics of being green and environmentally friendly. Therefore, it is necessary to develop a biotransformation system for producing D-pantothenic acid with low value substrates. [Method] With the β-alanine synthesis strain constructed in the previous study as the starting strain, a three-enzyme system was constructed by coupling L-aspartase from Corynebacterium glutamicum and pantothenate synthase (PanC) from Bacillus subtilis. The D-pantothenic acid biosynthesis pathway was validated and optimized in vitro and then reconstructed in cells. By optimizing the ratio of three enzymes and introducing an ATP cycle system, the system optimization and large-scale production of D-pantothenic acid were finally achieved in a 5 L bioreactor. [Result] Under the optimal biotransformation conditions, 70 g/L of fumaric acid and 89 g/L of D-pantolactone were converted to 128.6 g/L of D-pantothenic acid through whole cell catalysis, with a molar conversion rate of 97.3%. [Conclusion] An effective biotransformation system for producing D-pantothenic acid from fumaric acid and D-pantolactone is successfully developed.
