engleski

Molecular Modeling of Acetylacetone Dioxygenase from Acinobacter Jonsoii

Acetylacetone dioxygenase from Acinetobacter jonsoii, Dke1, is a non-heme Fe2+ dependent enzyme that catalyzes the oxidative C-C bond scission of β-dicarbonyl compounds by molecular oxygen. In nature it comes as homo-tetramer with each subunit organized in a single domain β-barrel fold characteristic of the Cupin superfamily of proteins. In order to better understand the enzymatic mechanism of O2 reduction in Dke1, and to find a possible correlation between the enzymatic activity and the role of outer shell waters and hydrophilicity in the active site, mutants of Dke1 were constructed with the aim to create space for waters in the active site. Therefore, the 9 mutants F51A, F59A, Y70A, R80A, E98N, E98A, F115A, F119A, and F125A were constructed, and the respective O2 reduction rates were measured and correlated with active site hydrophilicity. Therefore, we utilized several different molecular modeling approaches. First we determined the low energy positions for a water molecule in the Fe2+ binding site of the WT protein and its mutants. Interestingly, the number of possible positions roughly correlates with the measured O2 reduction rates. Furthermore we docked the 2, 4-pentanedione anion (-1) into the one subunit of the Dke1 tetramer and performed Molecular Dynamics (MD) simulations. During 2 ns of unconstrained MD simulation several water molecules entered into the protein metal binding sites, but no one water molecules entered into the metal binding site of the subunit to which the substrate was bound. This is in accord with the hypothesis proposed for the O2 activation mechanism generally proposed for mononuclear nonheme Fe(II) enzymes and may be a valuable approach for the in-silico design for Mononuclear nonheme Fe2+ enzymes in general.

modeling; macromolecule; protein; cleaving

nije evidentirano