Sustainable production of 2-phenylethanol from agro-industrial wastes by metabolically engineered Bacillus licheniformis

2-Phenylethanol (2-PE) is a kind of bulk flavor and fragrance, widely applied in food, fragrance, and cosmetic industries. In this work, a sustainable platform for 2-PE biosynthesis was developed by engineering Bacillus licheniformis using agro-industrial wastes as feedstocks. Firstly, the endogenous sucrose catabolism pathway of B. licheniformis was replaced by introducing CscB from Escherichia coli W and SucP from Bifdobacterium adolescentis. The 2-PE titer and yield of the resulting strain PE24 increased to 6.52 ± 0.20 g/L and 0.16 g/g sucrose, respectively, which increased by 16% compared to the control strain PE23 using 40 g/L sucrose as the carbon source. Subsequently, the effects of molasses and crude glycerol as the feedstocks on 2-PE production were investigated. By optimizing the molasses pretreatment methods and medium components, the 2-PE production was increased from 2.21 g/L to 5.74 g/L using 50 g/L pretreated molasses, 15 g/L crude glycerol, and 7 g/L corn steep liquor powder as the raw materials. Finally, we characterized the PE24 strain by fed-batch fermentation in 5-L fermenter and achieved a titer of 6.43 g/L of 2-PE, representing the highest level of 2-PE production. This study provided a cost-effective process for 2-PE sustainable biosynthesis.