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Réaction CuI-Tyr → CuII-radical Tyr° catalysée par O2 dans les systèmes biologiques: Des preuves spectroscopiques et cinétiques dans des complexes modèles. – CuO2-rad

Submission summary

The aim of the project is to draw connections between two independent research areas, namely the Cu(I)/ O2 interaction and the phenoxyl radical chemistries. These two chemistries are unambiguously connected in biological systems. In particular, some enzymes utilize very efficiently the Cu(I)/ O2 entity to generate post-translationally modified endogenous amino acids and catalytically essential radicals. - It has been proposed in Galactose Oxidase (GO) that the cross-link between the cysteine and the tyrosyl radical cofactor is initiated by copper(I) binding to a pre-form of the enzyme (Tyr). Using well-established dioxygen activation pathways in synthetic Cu(I) complexes, some mechanisms have been proposed. The copper(I) pre-form of the enzyme (CuI/ Tyr) reacts with O2, affording the fully reduced form of the mature GO (CuI/ Tyr-Cys). It reacts again with O2, in a path similar to that proposed for catalytic turnover, affording the CuII/ radical Tyr-Cys° active form and H2O2. - These studies evidence a crucial role for the dioxygen copper(I) interaction in biological radical cofactors formation. - The proposed mechanisms in biological systems are however highly speculative in nature, as the proposed intermediates have not been observed (due to the high reactivity of the copper(I) form of the enzyme with O2). Detailed mechanistic investigations imply to work at cryogenic temperatures with fast kinetic apparatus, what is obviously not conceivable with biological samples. Model complexes are thus expected to play an essential role in a better knowledge of this oxygenation step. - Previous biomimetic approaches have been developed in the following contexts: - - Cu(I)/ O2 interaction: the rates of O2 reactions with Cu(I) complexes of polypyridyl (and related) ligands are quite rapid and the resulting complexes are often thermally unstable species. A general mechanistic framework involves initial binding of dioxygen to a Cu(I) center to form a transient 1:1 Cu/O2 adduct (only implicated in cryostopped-flow kinetic studies as it is a short-lived intermediates) that reacts with a second Cu(I) complex to form a well-defined (because more stable) binuclear 2:1 Cu/O2 complex. While this general picture adequately describes most of the oxygenation pathways, the ligand structure has been shown to influence significantly the dioxygen activation mechanisms and kinetics. In particular, one of the rare (only two reports to date) phenol(ate)-CuI complex affords, in the presence of O2, a surprisingly thermodynamically stable 1:1 Cu/O2 adduct. - - Cu(II)/ phenoxyl complexes: the spectral signatures, structural attributes and reactivity of the Cu(II)/ phenoxyl entity are well known. - - To date, no connections between these research fields could be established, and no biomimetic models reproduce the CuI-phenol(ate) + O2 ' CuII-phenoxyl + H2O2 reaction. Only two attempts to model this reaction were reported in literature. Cu(II)/ O2 adducts were evidenced, but Cu(II)/ phenoxyl radicals could not be observed due to the inability of the systems to manage proton transfers. - - The aim of the project is to provide the first evidence for the CuI-phenol(ate) + O2 ' CuII-phenoxyl + H2O2 reaction and address the mechanistic part of the Cu(I)/ O2 reaction by using cryo-stopped flow fast kinetic spectrophotometry. Three major steps could be discerned: - Models for the radical cofactor generation (from Cu(I) complexes + O2) - The copper(I) complexes of phenol-containing ligands will be synthesized and their reactivity towards O2 investigated. Since the aspect of the phenol moiety being an H+ source is the key to get phenoxyl radicals as final product, the pro-radical ligands will be designed to manage efficiently proton transfers. - Models for the biogenesis of the Tyr-Cys cross-link - With aim of modelling the first stage in the cofactor generation of GO (Tyr + Cys ' Tyr-Cys), ligands lacking the substituent at the ortho and / or para position will be synthesized an...

Project coordination

Fabrice THOMAS (Organisme de recherche)

The author of this summary is the project coordinator, who is responsible for the content of this summary. The ANR declines any responsibility as for its contents.

Partner

Help of the ANR 150,000 euros
Beginning and duration of the scientific project: - 36 Months

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