Rational Engineering of Escherichia coli for High-Level Production of Riboflavin

Riboflavin is widely used as a food additive. Here, multiple strategies were used to increase riboflavin production in Escherichia coli LS31T. First, purR deletion and co-overexpression of fbp, purF, prs, gmk, and ndk genes resulted in an increase of 18.6% in riboflavin titer (reaching 729.7 mg/L). Second, optimization of reduced nicotinamide adenine dinucleotide phosphate/ nicotinamide adenine dinucleotide ratio and respiratory chain activity in LS31T increased the titer up to 1020.2 mg/L. Third, the expression level of the guaC gene in LS31T was downregulated by ribosome binding site replacement, and the riboflavin production was increased by 10.6% to 658.5 mg/L. Then, all the favorable modifications were integrated together, and the resulting strain LS72T produced 1339 mg/L of riboflavin. Moreover, the riboflavin titer of LS72T reached 21 g/L in fed-batch cultivation, with a yield of 110 mg riboflavin/g glucose. To our knowledge, both the riboflavin titer and yield obtained in fed-batch fermentation are the highest ones among all the rationally engineered strains.

Automated summary

Riboflavin production in Escherichia coli LS31T was significantly increased by employing multiple strategies and optimizations, leading to the highest titer and yield achieved in fed-batch fermentation among all rationally engineered strains.