Enhanced production of 2′-fucosyllactose in engineered Escherichia coli BL21star(DE3) by modulation of lactose metabolism and fucosyltransferase

•2′-fucosyllactose (2-FL) is a value-added human milk oligosaccharide in terms of nutraceutical and pharmaceutical purpose.•To minimize the metabolism of lactose, the endogenous lac operon was replaced by the modified lac operon containing lacZ△M15.•Replacement of lac operon leads alleviation of β-galactosidase activity and thereby, 2-FL yield was increased by 3 fold.•Improved soluble expression of FucT2 by fusion of the three aspartate tag might lead to higher 2-FL production with enhanced yield than the control strain.•Engineered E. coli BL21star(DE3) expressing GDP-l-fucose biosynthetic enzymes and three aspartate tagged fucosyltransferase produced 6.4g/L 2-FL in fed-batch fermentation using glycerol pH-stat. 2′-Fucosyllactose (2-FL) is one of most abundant functional oligosaccharides in human milk, which is involved in many biological functions for human health. To date, most microbial systems for 2-FL production have been limited to use Escherichia coli JM strains since they cannot metabolize lactose. In this study, E. coli BL21star(DE3) was engineered through deletion of the whole endogenous lactose operon and introduction of the modified lactose operon containing lacZ△M15 from E. coli K-12. Expression of genes for guanosine 5′-diphosphate (GDP)-l-fucose biosynthetic enzymes and heterologous α-1,2-fucosyltransferase (FucT2) from Helicobacter pylori allowed the engineered E. coli BL21star(DE3) to produce 2-FL with 3-times enhanced yield than the non-engineered E. coli BL21star(DE3). In addition, the titer and yield of 2-FL were further improved by adding the three aspartate molecules at the N-terminal of FucT2. Overall, 6.4g/L 2-FL with the yield of 0.225g 2-FL/g lactose was obtained in fed-batch fermentation of the engineered E. coli BL21star(DE3) expressing GDP-l-fucose biosynthetic enzymes and three aspartate tagged FucT2.