Oxygenic ligands of transient metal complexes play pivotal role in determining catalytic mechanisms and thermodynamic properties at the key steps in the reaction cycles of redox-active iron enzymes. Axial ligands of porphyrin cofactors have been extensively studied by Raman spectroscopy owning to major resonance enhancement via the electronic transition of the porphyrin macrocycle. Much less is known about oxygenic ligands in transient species of more structurally diverse non-heme iron enzymes. Vibrational studies in this class of enzymes can potentially yield critical structural information, but they are hindered by weak Raman enhancement of non-heme chromophores, photosensitivity, and magnetic interactions in dinuclear clusters. We utilize stable isotopes of dioxygen, bulk water, and substrates in time-resolved resonance Raman studies to detect and characterize transient species in several mono- and di-nuclear oxygen-activating iron enzymes. We analyze vibrational signatures of such species in comparison with similar isotopic substitutions on natural static or artificial analog species in order to derive complete vibration definition of the metal cluster, including oxygen- and solvent-derived ligands, terminal and bridging ligands in di-iron clusters, their exchangeability and protonation states. Application of this information to reconstruct mechanisms of the C-H bond activation by mono-nuclear taurine dioxygenase TauD and the O-O bond cleavage by di-nuclear soluble methane monooxygenase will be presented. Our results provide first evidence that proton affinities of oxygenic ligands in heme and non-heme metalloenzymes can deviate from p<i>K</i><sub>a</sub> in corresponding model compounds by over 15 pH units. Time-resolved data on compounds Q and T of sMMO are complemented by static and steady-state isotopic studies on di-ferric species of aldehyde deformylating oxygenase and di-iron urease UreA2B2. In addition to specific implications for intriguing catalytic mechanisms of each individual enzyme, isotope-difference Raman spectroscopy provides an unrivaled insight into the role of oxygenic ligands in reactivity of iron enzymes.

Oxygenic ligands in the catalysis by iron enzymes

251st National Meeting (2016)

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