Chemical Engineering Research Center, School of Chemical Engineering and Technology, Tianjin University, Tianjin 300350, PR China.
Chemical Engineering Research Center, School of Chemical Engineering and Technology, Tianjin University, Tianjin 300350, PR China; Tianjin Key Laboratory of Membrane Science and Desalination Technology, Tianjin University, Tianjin 300072, PR China.
Enzyme Microb Technol. 2023 Apr;165:110198. doi: 10.1016/j.enzmictec.2023.110198. Epub 2023 Jan 16.
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.
5-羟色氨酸（5-HTP）是血清素的化学前体，它在动物中合成褪黑素和血清素，调节情绪、睡眠和行为。色氨酸羟化酶（TPH）使用四氢生物蝶呤（BH4）作为辅助因子将 L-色氨酸（L-Trp）羟化为 5-HTP，而 TPH 的低催化活性限制了 L-Trp 的羟化速率。在这项研究中，研究了 L-Trp-TPH1-BH4 的催化机制和结构特征，并使用合理的设计策略提高了催化活性。然后获得了 S337A/F318Y 有利突变。分子动力学模拟表明，S337A/F318Y 突变体与 TPH1 形成盐桥的同时与底物吲哚环形成额外的氢键，稳定了吲哚环并增强了变体与 L-Trp 的结合亲和力。结果，5-HTP 的产量提高了 2.06 倍，生产出 0.91g/L 的 5-HTP。通过合理设计 TPH 结构来提高 L-Trp 的羟化效率，为 5-HTP 的绿色生产提供了前景。
