Abstact

Dioxygen (O(2)) is a potent oxidant used by aerobic organisms for energy transduction and critical biosynthetic processes. Numerous metalloenzymes harness O(2) to mediate C-H bond hydroxylation reactions, but most commonly feature iron or copper ions in their active site cofactors. In contrast, many manganese-activated enzymes horizontal line such as glutamine synthetase and isocitrate lyase horizontal line perform redox neutral chemical transformations and very few are known to activate O(2) or C-H bonds. Here, we report that the dimanganese-metalated form of the cambialistic monooxygenase SfbO (Mn(2)-SfbO) can efficiently mediate enzymatic C-H bond hydroxylation. The activity of the dimanganese form of SfbO toward substrate hydroxylation is comparable to that of its heterobimetallic Mn/Fe form but exhibits distinct kinetic profiles. Kinetic, spectroscopic, and structural studies invoke a mixed-valent dimanganese cofactor (Mn(II)Mn(III)) in O(2) activation and evidence a stoichiometric role for superoxide in maturing an O(2)-inert Mn(II)(2) cofactor. Computational studies support a hypothesis wherein superoxide addition to the Mn(II)(2) cofactor installs a critical bridging hydroxide ligand that stabilizes higher-valent manganese oxidation states. These findings establish the viability of proteinaceous dimanganese cofactors in mediating complex, multistep redox transformations.