Binuclear copper enzymes perform O2 activation for the monooxygenation of neuropeptides and hormones, reactions that are essential for neurochemistry for all higher eukaryotes. Binuclear copper monoogygenases can be classified into "coupled" and "non-coupled" based on the magnetic interaction between the two copper centers. The coupled binuclear enzymes (for example, tyrosinase (Ty) and catechol oxidase (CaO)) have two strongly coupled copper sites that reduce O2 to a dicopper peroxide intermediate that effects electrophilic aromatic substitution (EAS). Non-coupled binuclear copper monooxygenases (peptidylglycine ?-hydroxylating monooxygenase (PHM), dopamine ?-monooxygenase (D?M), and tyramine ?-monooxygenase (T?M)) feature two copper centers that are distant, and exhibit no magnetic exchange interaction. These non-coupled enzymes generate a reactive monocopper-O2 species that reacts through hydrogen-atom abstraction (HAA). However, the means by which electronic structure and exchange coupling influence binuclear copper sites towards either EAS or HAA reactivity are not known. In contrast to the more well-studied coupled binuclear enzymes, direct spectroscopic probes of reaction intermediates in the non-coupled binuclear copper monooxygenase family have not provided sufficient information to explain the HAA and subsequent hydroxylation mechanisms. Recent advances in PHM and T?M expression systems provide the opportunity to prepare active-site mutants for the first time which have the potential to allow kinetic trapping of key reaction intermediates for spectroscopic analysis. These studies require application of advanced spectroscopies such as resonance Raman, electron paramagnetic resonance, magnetic circular dichroism, and synchrotron-based methods, and in combination with computational methods, particularly density functional theory, the results will be related to enzymatic catalysis and provide a rationale for active site structural elements in the non-coupled binuclear copper oxygenases. Combined with previous results in the coupled binuclear family, an electronic structure/function model will be developed to explain the differences in reactivity between coupled and non- coupled binuclear copper enzymes. These studies will yield new details concerning the activation of O2 by copper enzymes, insight that is useful for understanding the biochemistry of copper on a molecular level.

Public Health Relevance

Copper-dependent monooxygenases are essential for neurochemistry, and also play critical roles in a range of health issues, including breast,1 lung,2 and prostate3 cancers, Alzheimer's disease,4 and tumor growth.5 Understanding the biochemistry of copper on a molecular level provides the means to inhibit or improve these problems.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Postdoctoral Individual National Research Service Award (F32)
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Special Emphasis Panel (ZRG1-F04-D (20))
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Barski, Oleg
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Stanford University
Schools of Arts and Sciences
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Peterson, Ryan L; Ginsbach, Jake W; Cowley, Ryan E et al. (2013) Stepwise protonation and electron-transfer reduction of a primary copper-dioxygen adduct. J Am Chem Soc 135:16454-67
Ginsbach, Jake W; Peterson, Ryan L; Cowley, Ryan E et al. (2013) Correlation of the electronic and geometric structures in mononuclear copper(II) superoxide complexes. Inorg Chem 52:12872-4