Construction of cell factory capable of efficiently converting L-tryptophan into 5-hydroxytryptamine

Background: L-Tryptophan (L-Trp) derivatives such as 5-hydroxytryptophan (5-HTP) and 5-hydroxytryptamine (5-HT), N-Acetyl-5-hydroxytryptamine and melatonin are important molecules with pharmaceutical interest. Among, 5-HT is an inhibitory neurotransmitter with proven benefits for treating the symptoms of depression. At present, 5-HT depends on plant extraction and chemical synthesis, which limits its mass production and causes environmental problems. Therefore, it is necessary to develop an efficient, green and sustainable biosynthesis method to produce 5-HT. Results: Here we propose a one-pot production of 5-HT from L-Trp via two enzyme cascades for the first time. First, a chassis cell that can convert L-Trp into 5-HTP was constructed by heterologous expression of tryptophan hydroxylase from Schistosoma mansoni (SmTPH) and an artificial endogenous tetrahydrobiopterin (BH4) module. Then, dopa decarboxylase from Harminia axyridis (HaDDC), which can specifically catalyse 5-HTP to 5-HT, was used for 5-HT production. The cell factory, E. coli BL21(DE3)Delta tnaA/BH4/HaDDC-SmTPH, which contains SmTPH and HaDDC, was constructed for 5-HT synthesis. The highest concentration of 5-HT reached 414.5 +/- 1.6 mg/L (with conversion rate of 25.9 mol%) at the optimal conditions (substrate concentration,2 g/L; induced temperature, 25 degrees C; IPTG concentration, 0.5 mM; catalysis temperature, 30 degrees C; catalysis time, 72 h). Conclusions: This protocol provided an efficient one-pot method for converting. L-Trp into 5-HT production, which opens up possibilities for the practical biosynthesis of natural 5-HT at an industrial scale.

Automated summary

This study proposes a one-pot method for the production of 5-HT from L-Trp via two enzyme cascades. A chassis cell capable of converting L-Trp to 5-HTP was constructed by expressing tryptophan hydroxylase from Schistosoma mansoni (SmTPH) and an artificial endogenous tetrahydrobiopterin (BH4) module. Then, dopa decarboxylase from Harminia axyridis (HaDDC) was used for the specific catalysis of 5-HTP to 5-HT. The constructed cell factory, E. coli BL21(DE3)Delta tnaA/BH4/HaDDC-SmTPH, achieved the highest 5-HT concentration of 414.5 +/- 1.6 mg/L (conversion rate of 25.9 mol%) under optimal conditions (substrate concentration of 2 g/L, induced temperature of 25 degrees C, IPTG concentration of 0.5 mM, catalysis temperature of 30 degrees C, catalysis time of 72 h).