Alteration of the Catalytic Reaction Trajectory of a Vicinal Oxygen Chelate Enzyme by Directed Evolution. Issue 26 (27th April 2022)

• Record Type: Journal Article
• Authors: Wang, Yi Shuang; Zheng, Wan; Jiang, Nan; Jin, Yun Xia; Meng, Zi Kang; Sun, Meng Xin; Zong, Yu Liang; Xu, Tong; Zhu, Jiapeng; Tan, Ren Xiang
• Is Part Of: Angewandte Chemie international edition. Volume 61:Issue 26(2022)
• Page Start: n/a
• Page End: n/a
• Publication Date: 2022-04-27
• Subjects: Bifunctional Enzyme -- Decarboxylation -- Dioxygenase -- Directed Evolution -- VOC Enzyme Family; Chemistry -- Periodicals; 540
• DOI: 10.1002/anie.202201321
• Languages: English
• ISSNs: 1433-7851

Abstract

Abstract: The vicinal oxygen chelate (VOC) metalloenzyme superfamily catalyzes a highly diverse set of reactions with the mechanism characterized by the bidentate coordination of vicinal oxygen atoms to metal ion centers, but there remains a lack of a platform to steer the reaction trajectories, especially for o ‐quinone metabolizing pathways. Herein, we present the directed‐evolution‐enabled bifunctional turnover of ChaP, which is a homotetramer and represents an unprecedented VOC enzyme class. Unlike the ChaP catalysis of extradiol‐like o ‐quinone cleavage and concomitant α‐keto acid decarboxylation, a group of ChaP variants (CVs) catalyze intradiol‐like o ‐quinone deconstruction and CO2 liberation from the resulting o ‐hydroxybenzoic acid scaffolds with high regioselectivity. Enzyme crystal structures, labeling experiments and computational simulations corroborated that the D49L mutation allows the metal ion to change its coordination with the tyrosine phenoxy atoms in different monomers, thereby altering the reaction trajectory with the regiospecificity further improved by the follow‐up replacement of the Y92 residue with any of alanine, glycine, threonine, and serine. The study highlights the unpredicted catalytic versatility and enzymatic plasticity of VOC enzymes with biotechnological significance. Abstract : Directed evolution of ChaP, a novel vicinal oxygen chelate (VOC) metalloenzyme, afforded bifunctional enzymes designated as "ChaP variants", which catalyzed regioselective intradiol‐like o‐ quinone cleavage and o ‐hydroxybenzoic acid decarboxylation of the ring‐opened molecules. This activity expands the VOC catalytic repertoire to bring us closer to artificial VOC enzymes of broad biotechnological significance.