title: Macrophomate synthase: QM/MM simulations address the Diels-Alder versus Michael-Aldol reaction mechanism.
abstract: Macrophomate synthase (MPS) of the phytopathogenic fungus Macrophoma commelinae catalyzes the transformation of 2-pyrone derivatives to the corresponding benzoate analogues. The transformation proceeds through three separate chemical reactions, including decarboxylation of oxalacetate to produce pyruvate enolate, two C-C bond formations between 2-pyrone and pyruvate enolate that form a bicyclic intermediate, and final decarboxylation with concomitant dehydration. Although some evidence suggests that the second step of the reaction catalyzed by MPS is a Diels-Alder reaction, definite proof that the C-C bond formations occur via a concerted mechanism is still required. An alternative route for formation of the C-C bonds is a stepwise Michael-aldol reaction. In this work, mixed quantum and molecular mechanics (QM/MM) combined with Monte Carlo simulations and free-energy perturbation (FEP) calculations were performed to investigate the relative stabilities of the transition states (TS) for both reaction mechanisms. The key results are that the Diels-Alder TS model is 17.7 and 12.1 kcal/mol less stable than the Michael and aldol TSs in the stepwise route, respectively. Therefore, this work indicates that the Michael-aldol mechanism is the route used by MPS to catalyze the second step of the overall transformation, and that the enzyme is not a natural Diels-Alderase, as claimed by Ose and co-workers (Nature 2003, 422, 185-189; Acta Crystallogr. 2004, D60, 1187-1197). A modified link-atom treatment for the bonds at the QM/MM interface is also presented.
authors.affiliation: Department of Chemistry, Yale University, 225 Prospect Street, New Haven, Connecticut 06520-8107, USA.
authors.forename: Cristiano Ruch Werneck
authors.initials: CR
authors.lastname: Guimarães
authors.forename: Marina
authors.initials: M
authors.lastname: Udier-Blagović
authors.forename: William L
authors.initials: WL
authors.lastname: Jorgensen
mesh_terms: D001565:Benzoates / Q000737:chemistry / Q000378:metabolism; D003198:Computer Simulation; D005656:Fungal Proteins / Q000737:chemistry* / Q000378:metabolism*; D003904:Mitosporic Fungi / Q000201:enzymology; D008958:Models, Molecular; D009097:Multienzyme Complexes / Q000737:chemistry* / Q000378:metabolism*; D011753:Pyrones / Q000737:chemistry / Q000378:metabolism; D011789:Quantum Theory; D013816:Thermodynamics
chemical_list: D001565:Benzoates; D005656:Fungal Proteins; D009097:Multienzyme Complexes; D011753:Pyrones; C408144:macrophomate synthase