Multidimensional Metabolic Engineering for Green and Sustainable Bioproduction of Serotonin from Glucose in Bacillus licheniformis.

Serotonin (5-hydroxytryptamine, 5-HT) plays a critical role in maintaining intestinal health, regulating mood and sleep, and enhancing crop stress tolerance. Here, a Bacillus licheniformis cell factory was developed for the sustainable and efficient synthesis of 5-HT. First, a tryptophan-producing chassis strain, TRP18, was obtained by blocking competitive pathways, modifying the tryptophan operon, and rewiring carbon metabolic flux, and the Trp titer was increased by 23.94-fold over the starting strain, TRP0. Subsequently, a heterologous 5-HT biosynthetic pathway was established, and tryptophan hydroxylase Luz15 was optimized by protein engineering. Finally, the 5-HT pathway was optimized by a dual-cassette system, and an optimal strain, HT-12, was attained, producing 1.23 g/L 5-HT in a 5-L bioreactor, representing the highest titer of 5-HT from glucose in Bacillus species to date. This study provides an efficient and sustainable process for 5-HT production in B. licheniformis, laying the foundation for industrial-scale manufacturing of 5-HT and other Trp-derived biochemicals.