De Novo Synthesis of Tyramine in Engineered Escherichia coli Using Two-Stage Dissolved Oxygen-Controlled Fermentation

Metabolic regulation and fermentation strategies limit the synthesis of tyramine. Here, a de novo synthetic tyramine-producing strain, LAN 25, was constructed. First, tyramine-producing strain was obtained by modifying the key metabolic nodes of tyrosine synthesis and overexpressing the tyrosine decarboxylase gene from Lactobacillus brevis. Then, a two-stage dissolved oxygen (DO)-controlled fermentation process was established in a 5 L fed-batch bioreactor. In the first stage, sufficient DO was supplied to accumulate tyrosine precursors, while in the second stage, limited DO was provided to promote tyramine synthesis. Next, ldhA and adhE were deleted to improve the strain's robustness. To enhance the redox flux (ATP/ADP ratio) under limited DO conditions, the anaerobic promoter, Pvgb, was used to control the expression of a ppk-based ATP regeneration system in response to DO changes. Additionally, the tyrosine internal transport system was modified. Finally, a titer of 21.33 g/L of tyramine with a yield of 0.092 g/g glucose was obtained.