. The goal of this proposal is to understand how dioxygen is activated by biological diiron centers in metabolically critical transformations. Nonheme diiron proteins and enzymes perform a variety of essential functions involving dioxygen, including dioxygen transport (hemerythrin), DMA biosynthesis (ribonucleotide reductase), iron storage (ferritin), and oxidations of organic substrates (methane monooxygenase, fatty acid desaturases, alkane and arene hydroxylases, myo-inositol oxygenase, deoxyhypusine hydroxylase). In general, dioxygen activation is proposed to entail a common mechanism involving diiron(lll)-peroxo intermediates and high-valent iron-oxo species derived therefrom. The project goal will be accomplished using a combination of biomimetic and spectroscopic approaches. Building on past accomplishments in modeling structural and spectroscopic properties of such sites, it is proposed to synthesize precursor complexes of tripodal ligands, to react them with O2 or peroxides, and to characterize the metastable intermediates derived therefrom. Of great interest are intermediates such as O2 adducts of diiron(ll) complexes (either iron(ll)iron(lll)-superoxo or diiron(lll)-peroxo species), and species with Fe(lll-)Fe(-IV) and Fe(-IV)Fe(-lV) oxidation states. These complexes will be characterized by X-ray crystallography whenever possible and by a variety of techniques such as NMR, EPR, UV-vis-NIR, Raman, Mossbauer, electrospray mass spectrometry, electrochemistry, and EXAFS. Both stopped-flow and conventional kinetic methods will be used to characterize their mechanisms of formation and decomposition. The oxidative reactivities of these transient complexes towards a range of substrates will be investigated and compared with those of enzyme active sites. Also to be synthesized are complexes that can serve as precedents for the novel oxygen activation mechanism recently proposed for myo-inositol oxygenase entailing an iron(ll)iron(lll) center that binds O2 and a diiron(lll)-superoxo species that acts as the initial oxidant. Parallel to these efforts, our spectroscopic expertise will be applied to elucidating the diiron site structures of methane monooxygenase intermediates and deoxyhypusine hydroxylase. Relevance. Nonheme diiron enzymes perform a variety of metabolically critical functions that require dioxygen activation. Understanding how these enzymes work can lead to the development of new drug strategies for treating some human diseases. For example, myo-inositol oxygenase may be connected to the many complications associated with diabetes mellitus, while deoxyhypusine hydroxylase is required for the formation of mature eukaryotic elongation factor 5a that is essential for cell proliferation and may thus serve as the target for anti-tumor or anti-HIV therapy.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Research Project (R01)
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Macromolecular Structure and Function A Study Section (MSFA)
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Anderson, Vernon
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University of Minnesota Twin Cities
Schools of Arts and Sciences
United States
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Castillo, Rebeca G; Banerjee, Rahul; Allpress, Caleb J et al. (2017) High-Energy-Resolution Fluorescence-Detected X-ray Absorption of the Q Intermediate of Soluble Methane Monooxygenase. J Am Chem Soc 139:18024-18033
Komor, Anna J; Rivard, Brent S; Fan, Ruixi et al. (2017) CmlI N-Oxygenase Catalyzes the Final Three Steps in Chloramphenicol Biosynthesis without Dissociation of Intermediates. Biochemistry 56:4940-4950
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Komor, Anna J; Rivard, Brent S; Fan, Ruixi et al. (2016) Mechanism for Six-Electron Aryl-N-Oxygenation by the Non-Heme Diiron Enzyme CmlI. J Am Chem Soc 138:7411-21
Jasniewski, Andrew J; Engstrom, Lisa M; Vu, Van V et al. (2016) X-ray absorption spectroscopic characterization of the diferric-peroxo intermediate of human deoxyhypusine hydroxylase in the presence of its substrate eIF5a. J Biol Inorg Chem 21:605-18
Jasniewski, Andrew J; Knoot, Cory J; Lipscomb, John D et al. (2016) A Carboxylate Shift Regulates Dioxygen Activation by the Diiron Nonheme ?-Hydroxylase CmlA upon Binding of a Substrate-Loaded Nonribosomal Peptide Synthetase. Biochemistry 55:5818-5831
Ye, Shengfa; Xue, Genqiang; Krivokapic, Itana et al. (2015) Magnetic circular dichroism and computational study of mononuclear and dinuclear iron(IV) complexes. Chem Sci 6:2909-2921

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