Directed Evolution of a Tryptophan 2,3‐Dioxygenase for the Diastereoselective Monooxygenation of Tryptophans

Abstract

Herein, we report an engineered enzyme that can monooxygenate unprotected tryptophan into the corresponding 3a‐hydroxyhexahydropyrrolo[2,3‐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.