| Institution: | 1. College of Bioscience and Biotechnology, Yangzhou University, Yangzhou, Jiangsu, 225009 China
These authors contributed equally to this work.;2. College of Bioscience and Biotechnology, Yangzhou University, Yangzhou, Jiangsu, 225009 China;3. Testing Center, Yangzhou University, Yangzhou, Jiangsu, 225009 China;4. Department of Chemistry, Stanford University, Stanford, California, 94305 USA |
| Abstract: | Herein, we report an engineered enzyme that can monooxygenate unprotected tryptophan into the corresponding 3a-hydroxyhexahydropyrrolo2,3-b]indole-2-carboxylic acid (HPIC) in a single, scalable step with excellent turnover number and diastereoselectivity. Taking advantage of directed evolution, we analyzed the stepwise oxygen-insertion mechanism of tryptophan 2,3-dioxygenases, and transformed tryptophan 2,3-dioxygenase from Xanthomonas campestris into a monooxygenase for oxidative cyclization of tryptophans. It was revealed that residue F51 is vital in determining the product ratio of HPIC to N′-formylkynurenine. Our reactions and purification procedures use no organic solvents, resulting in an eco-friendly method to prepare HPICs for further applications. |
