Molecular dynamics study of oxygenation reactions catalysed by the enzyme p-hydroxybenzoate hydroxylase

Combined quantum mechanical and molecular mechanical (QM/MM) methods have been used to model the rate-determining step of the title reaction for three different substrates, i.e., p-oxybenzoate dianion 1, p-hydroxybenzoate anion 2 and p-oxybenzoic acid anion 3. The system studied includes the enzyme, the cofactor, the substrate, and crystal water (ca. 7000 atoms total, ca. 100 QM atoms). Molecular dynamics simulations at the AM1/GROMOS level confirm that the dianion mechanism is favoured over both monoanion mechanisms, with a computed free energy barrier of 12 kcal mol⁻¹. Geometry optimisations of model systems (QM) and of the complete enzyme (QM/MM) provide additional information on the intrinsic barriers and transition states