QM/MM Molecular Dynamics Investigations of the Substrate Binding of Leucotriene A4 Hydrolase: Implication for the Catalytic Mechanism.

LTA4H is a monozinc bifunctional enzyme which exhibits both aminopeptidase and epoxide hydrolase activities. Its dual functions in anti- and pro-inflammatory roles have attracted wide attention to the inhibitor design. In this work, we tried to construct Michaelis complexes of LTA4H with both a native peptide substrate and LTA4 molecule using combined quantum mechanics and molecular mechanics molecular dynamics simulations. First of all, the zinc ion is coordinated by H295, H299, and E318. For its aminopeptidase activity, similar to conventional peptidases, the fourth ligand to the zinc ion is suggested to be an active site water, which is further hydrogen bonded with a downstream glutamic acid, E296. For the epoxide hydrolase activity, the fourth ligand to the zinc ion is found to be an epoxy oxygen atom. The potential of mean force calculation indicates about an 8.5 kcal/mol activation barrier height for the ring-opening reaction, which will generate a metastable carbenium intermediate. Subsequent frontier molecular orbital analyses suggest that the next step would be the nucleophilic attacking reaction at the C12 atom by a water molecule activated by D375. Our simulations also analyzed functions of several important residues like R563, K565, E271, Y383, and Y378 in the binding of peptide and LTA4.