Combinatorial metabolic engineering and tolerance evolving of Escherichia coli for high production of 2'-fucosyllactose

Abstract

2'-Fucosyllactose (2'-FL) is an important functional ingredient of advanced infant formula. Here, Escherichia coli MG1655 was engineered for achieving high 2'-FL production. The expressions of 2'-FL synthesis pathway genes were finely regulated with single or multi copies according to rate-limiting enzyme diagnosis. On this basic, the branch pathway genes were deleted, and the overexpression of the 2'-FL efflux protein SetA and the fructose-1,6-bisphosphatase GlpX were tuned. The resulting strain produced 46.06 ± 1.28 g/L 2'-FL in a 5-L fermenter. Furtherly, adaptive laboratory evolution was conducted. A rpoC gene mutation was obtained which could improve the cell tolerance and the 2'-FL production up to 61.06 ± 1.93 g/L, with the highest productivity of 1.70 g/L/h among E. coli strains by now. Taken together, this work provides a combinatorial strategy to improve 2'-FL accumulation including rational fine-tuning pathway genes expressions and irrational adaptive laboratory evolution. This study should be helpful for constructing high level 2'-FL producers.

Keywords: 2′-fucosyllactose; Adaptive laboratory evolution; Escherichia coli; Metabolic engineering.