On the reaction mechanism of an endoperoxide ring formation by fumitremorgin B endoperoxidase. The right arrangement makes a difference†
Fumitremorgin B endoperoxidase (FtmF) belongs to 2-oxoglutarate dependent dioxygenases and catalyzes an unusual oxidative reaction of endoperoxide formation at the final stage of biosynthesis of verruculogen – a mycotoxin produced by Aspergillus and Penicillinum strains. The published crystal structure of FtmF (PDB: 4ZON), which is of overall good quality, contains a model of the substrate bound in the active site, which, however, has very low occupancy and its conformation does not comply with the small molecule crystal structure. Moreover, a previous computational study that employed a model based on this crystal structure revealed a substantial reaction barrier, which might indicate that the model of FtmF/substrate complex can have serious errors. The purpose of this work was to model with computational methods the structure of the enzyme–substrate complex and to investigate the mechanisms of the enzymatic reaction. Docking, molecular dynamics simulation and DFT results, all indicate the substrate most likely binds in the active site in a configuration very different from that originally suggested. Moreover, for this newly proposed structure of the enzyme–substrate complex, the reaction energy profile is characterised exclusively by low barriers and it successfully explains the observed regiospecificity of the enzymatic process. Finally, a plausible binding site for ascorbate was found and it is suggested that ascorbate is involved in the final step of the FtmF reaction.