Mechanistic studies of functional mononuclear and binuclear non-heme iron enzyme model complexes using variable temperature stopped-flow UV/vis spectroscopy
Gregor, Lauren Christine
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Variable-temperature stopped-flow (VT-SF) electronic spectroscopy (-85 to -50°C) was utilized to study the reactivity properties of a family of synthetic mononuclear and binuclear non-heme iron enzyme active site analogs. This technique was used to investigate the mechanisms of interactions of two diiron complexes, the diferrous [FeII2(H2Hbamb)2(NMI)2] and the mixed valent [FeII,FeIII(H2Hbamb)2]+, with either oxygen-atom donor (OAD) molecules or the mechanistic probe peroxide, 2-methyl-1-phenylprop-2-yl hydroperoxide (MPPH), and substrates containing weak C-H and O-H bonds. Single turnover studies with 9,10-dihydroanthracene (9,10-DHA) and the deuterated analog, d4-9,10-DHA allowed for the determination of kinetic isotope effects (KIE) which show an inverse KIE and evidence of a disproportionation mechanism. Previous investigations showed the rate of catalytic oxidation of cyclohexane to cyclohexanol by [FeII2(H2Hbamb)2(NMI)2] and MPPH decreased over time. Current VT-SF data show evidence of product inhibition by means of a pre-equilibrium process that inhibits the reaction of the oxidant with the [FeII,FeII] complex. Also examined is the ability of the [FeII,FeIII(H2Hbamb)2]+ complex to catalytically oxidize phenols to phenoxyl radicals via a putative [FeIV=O] species. The reactivity properties of substituted phenols that vary in their oxidation potentials and bond dissociation energies (BDE) was investigated by VT-SF electronic spectroscopic studies to gain insight into the mechanism of oxidation by the [FeII,FeIII] complex. Mechanistic studies were also performed utilizing a mononuclear non-heme iron complex [FeII(N2O1)(CH3OH)Cl2], which can bind alpha-keto acids (e.g. alpha-ketoglutarate, benzoylformate) in a bidentate fashion. Reactivity studies utilizing O2 shows coupled decarboxylation of the alpha-keto acid with catalytic oxidation of the methanol solvent formaldehyde (285 turnovers). In non-reactive solvents, the [FeII(N2O1)(alpha-KG)] adduct complex is capable of catalytically oxidizing a variety of substrates such as 9,10-dihydroanthracene, 2,4-di-tert-butyl phenol, cyclohexene, and cyclooctane at 25°C utilizing O2 as the oxidant. Investigations to the binding of alpha-keto acids to the mononuclear iron complex in the absence of O2 by VT-SF as well as binding studies with NO are discussed. Finally, VT-SF studies were performed to probe the reaction of O2 with [FeII(N2O1)(alpha-KG)(CH3OH)] and the proposed mechanism is discussed. The relevance of these data to non-heme iron enzymes like soluble Methane Monooxygenase, Ribonucleotide Reductase, and Taurine Dioxygenase is discussed.