A Quantum Mechanical/Molecular Mechanical Study of the Hydroxylation of Phenol and Halogenated Derivatives by Phenol Hydroxylase

Abstract

A combined quantum mechanical and molecular mechanical (QM/MM) method (AM1/CHARMM) was used to investigate the mechanism of the aromatic hydroxylation of phenol by a flavin dependent phenol hydroxylase (PH), an essential reaction in the degradation of a wide range of aromatic compounds. The model for the reactive flavin intermediate (C4a-hydroperoxyflavin) bound to PH was constructed on the basis of the crystal structure of the enzyme−substrate complex. A potential energy surface (PES) was calculated as a function of the reaction coordinates for hydroxylation of phenol (on C6) and for proton transfer from phenol (O1) to an active-site base Asp54 (OD1). The results support a reaction mechanism in which phenol is activated through deprotonation by Asp54, after which the phenolate is hydroxylated through an electrophilic aromatic substitution. Ab initio test calculations were performed to verify these results of the QM/MM model. Furthermore, the variation in the calculated QM/MM activation energies for hydro...