Engineering Escherichia coli for Efficient Production of L-Tryptophan

Abstract

L-tryptophan is an essential amino acid widely used in animal feed, food additives, and pharmaceuticals. Although the biosynthesis of L-tryptophan from renewable raw materials is sustainable and environmentally friendly, its long synthetic pathway and complex regulatory mechanisms often result in low production efficiency. In this study, multiple strategies were employed to improve L-tryptophan production in Escherichia coli. These strategies included three main ones: the introduction of anti-feedback inhibition mutations, the overexpression of key rate-limiting enzymes, and the blockage of competitive and catabolic pathways. Additionally, the glucose transport system was modified by replacing it with a non-phosphoenolpyruvate (PEP)-dependent system, thereby reducing PEP consumption. In addition, overexpression of an L-tryptophan transporter protein enhanced L-tryptophan efflux and alleviated product inhibition. Consequently, an efficient L-tryptophan-producing E. coli strain was constructed. Furthermore, dynamic regulation of L-tryptophan synthesis was initially explored by employing a temperature-sensitive promoter to modulate the PEP metabolic pathway. Scale-up fermentation in a 5-L bioreactor demonstrated that the final strain (TY9) produced 34.1 g·L⁻¹ L-tryptophan under fed-batch conditions.