Rational design of tryptophan hydroxylation 1 for improving 5-Hydroxytryptophan production

Abstract

5-Hydroxytryptophan (5-HTP) is a chemical precursor of serotonin, which synthesizes melatonin and serotonin in animals and regulates mood, sleep, and behavior. Tryptophan hydroxylase (TPH) uses tetrahydrobiopterin (BH4) as a cofactor to hydroxylate L-tryptophan (L-Trp) to 5-HTP, and the low catalytic activity of TPH limits the rate of hydroxylation of L-Trp. In this study, the catalytic mechanism and structural features of L-Trp-TPH1-BH4 were investigated, and the catalytic activity was improved using a rational design strategy. Then the S337A/F318Y beneficial mutation was obtained. Molecular dynamics simulations showed that the S337A/F318Y mutant formed a salt bridge with TPH1 while forming an additional hydrogen bond with the substrate indole ring, stabilizing the indole ring and enhancing the binding affinity of the variant to L-Trp. As a result, the yield of 5-HTP was increased by 2.06-fold, resulting in the production of 0.91 g/L of 5-HTP. The rational design of the TPH structure to improve the hydroxylation efficiency of L-Trp offers the prospect of green production of 5-HTP.