Mononuclear non-heme-iron (MNH-Fe) enzymes activate O2 for a stunning array of biomedically, agriculturally, and environmentally important oxidation reactions. Our past decade's work, supported (in part) by this grant, established the intermediacy of iron(IV)-oxo (ferryl) complexes in the reactions of seven different MNH-Fe enzymes. Five of these complexes generate substrate radicals by abstracting hydrogen (H?) from unactivated aliphatic carbons, initiating formation of new C-O, C- Cl/Br, and C-S bonds. Energized by our recent success in rationalizing the divergent outcomes mediated by the (halo)ferryl complexes in the ?-ketoglutarate(?KG)-dependent aliphatic hydroxylases and halogenases, we now aim to understand even more complex ferryl-mediated transformations, including those exhibited by the enzymes: (1) hydroxypropylphosponate epoxidase (HppE), which catalyzes the 1,3-dehydrogenation of an alcohol to an epoxide, using hydrogen peroxide as the oxidant, in the biosynthesis of the antibiotic, fosfomycin;(2) carbapenem synthase (CarC), which uses one or more tyrosyl radical in concert with the presumptive ferryl complex to promote stereoinversion of a chiral carbon and desaturation of a C-C bond two atoms removed from the stereocenter, reportedly in a single O2 activation event, to produce the core of an important class of antibiotics;and (3) 2-hydroxyethylphosponate (2-HEP) dioxygenase (HEPD) and methylphosphonate synthase (MPnS), a pair of related enzymes that use ferryl complexes to cleave the C-C bond of 2- HEP in distinct 4-e- oxidation reactions, producing a precursor to the herbicide phosphinothricin (HEPD) and a major store of oceanic methane (MPnS). Our past studies on myo-inositol oxygenase and isopenicillin N synthase demonstrated a fundamentally distinct manifold for enzymatic O2 and C- H activation, involving H? abstracting FeIII-superoxo complexes. This manifold obviates the requirement for a reducing co-substrate (e.g., ?KG), enabling four-electron (4-e-) oxidations. HEPD and MPnS are likely also to employ this manifold on the pathways to their ferryl intermediates, a hypothesis that we will test here. We will elucidate the mechanisms of these fascinating enzymes to develop an integrated understanding of their complex oxidation chemistry.

Public Health Relevance

Soil microbes synthesize a bewildering array of known and yet-to-be-discovered natural products with herbicidal, antibiotic, antifungal, antiviral, and anticancer activities. Enzymes that use mononuclear non-heme iron cofactors to activate dioxygen (or in some cases hydrogen peroxide) and cleave strong carbon-hydrogen bonds play key roles in the construction, combinatorial elaboration, and degradation of many of these natural products, and functionally related enzymes in humans have roles in control of transcription and differentiation, detection of and response to oxygen insufficiency (hypoxia), synthesis of connective tissue, and repair of cellular genetic material (DNA). An integrated understanding of the diverse chemistry mediated by members of this important enzyme family, which is sought in this project, would inform design of drugs targeting the enzymes, discovery of new bioactive natural products, and development of new biomimetic chemical processes.

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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Pennsylvania State University
Schools of Arts and Sciences
University Park
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Martinie, Ryan J; Pollock, Christopher J; Matthews, Megan L et al. (2017) Vanadyl as a Stable Structural Mimic of Reactive Ferryl Intermediates in Mononuclear Nonheme-Iron Enzymes. Inorg Chem 56:13382-13389
Peck, Spencer C; Wang, Chen; Dassama, Laura M K et al. (2017) O-H Activation by an Unexpected Ferryl Intermediate during Catalysis by 2-Hydroxyethylphosphonate Dioxygenase. J Am Chem Soc 139:2045-2052
Tamanaha, Esta; Zhang, Bo; Guo, Yisong et al. (2016) Spectroscopic Evidence for the Two C-H-Cleaving Intermediates of Aspergillus nidulans Isopenicillin N Synthase. J Am Chem Soc 138:8862-74
Srnec, Martin; Wong, Shaun D; Matthews, Megan L et al. (2016) Electronic Structure of the Ferryl Intermediate in the ?-Ketoglutarate Dependent Non-Heme Iron Halogenase SyrB2: Contributions to H Atom Abstraction Reactivity. J Am Chem Soc 138:5110-22
Martinie, Ryan J; Livada, Jovan; Chang, Wei-chen et al. (2015) Experimental Correlation of Substrate Position with Reaction Outcome in the Aliphatic Halogenase, SyrB2. J Am Chem Soc 137:6912-9
Matthews, Megan L; Chang, Wei-chen; Layne, Andrew P et al. (2014) Direct nitration and azidation of aliphatic carbons by an iron-dependent halogenase. Nat Chem Biol 10:209-15
Chang, Wei-chen; Guo, Yisong; Wang, Chen et al. (2014) Mechanism of the C5 stereoinversion reaction in the biosynthesis of carbapenem antibiotics. Science 343:1140-4
Krebs, Carsten; Dassama, Laura M K; Matthews, Megan L et al. (2013) Novel Approaches for the Accumulation of Oxygenated Intermediates to Multi-Millimolar Concentrations. Coord Chem Rev 257:
Wong, Shaun D; Srnec, Martin; Matthews, Megan L et al. (2013) Elucidation of the Fe(IV)=O intermediate in the catalytic cycle of the halogenase SyrB2. Nature 499:320-3
Wang, Chen; Chang, Wei-chen; Guo, Yisong et al. (2013) Evidence that the fosfomycin-producing epoxidase, HppE, is a non-heme-iron peroxidase. Science 342:991-5

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