Computational exploration of the catalytic mechanism of dopamine beta-monooxygenase: modeling of its mononuclear copper active sites.

Dopamine hydroxylation by the copper-superoxo, -hydroperoxo, and -oxo species of dopamine beta-monooxygenase (DBM) is investigated using theoretical calculations to identify the active species in its reaction and to reveal the key functions of the surrounding amino acid residues in substrate binding. A 3D model of rat DBM is constructed by homology modeling using the crystal structure of peptidylglycine alpha-hydroxylating monooxygenase (PHM) with a high sequence identity of 30% as a template. In the constructed 3D model, the CuA site in domain 1 is coordinated by three histidine residues, His265, His266, and His336, while the CuB site in domain 2 is coordinated by two histidine residues, His415 and His417, and by a methionine residue, Met490. The three Glu268, Glu369, and Tyr494 residues are suggested to play an important role in the substrate binding at the active site of DBM to enable the stereospecific hydrogen-atom abstraction. Quantum mechanical/molecular mechanical (QM/MM) calculations are performed to determine the structure of the copper-superoxo, -hydroperoxo, and -oxo species in the whole-enzyme model with about 4700 atoms. The reactivity of the three oxidants is evaluated in terms of density-functional-theory calculations with small models extracted from the QM region of the whole-enzyme model.