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Direct Evidence for a Peroxide Intermediate and a Reactive Enzyme-Substrate-Dioxygen Configuration in a Cofactor-free Oxidase

DOI: 10.1002/anie.201405485 DOI Help
PMID: 25314114 PMID Help

Authors: Soi Bui (King's College London) , David Von Stetten (ESRF) , Pablo G. Jambrina (King's College London) , Thierry Prangé (Université Paris Descartes-CNRS) , Nathalie Colloc'h (Normandie Université) , Daniele De Sanctis (ESRF) , Antoine Royant (ESRF) , Edina Rosta (King's College London) , Roberto A Steiner (King's College London)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Angewandte Chemie International Edition , VOL 53 (50) , PAGES 13710 - 13714

State: Published (Approved)
Published: December 2014

Abstract: Cofactor-free oxidases and oxygenases promote and control the reactivity of O2 with limited chemical tools at their disposal. Their mechanism of action is not completely understood and structural information is not available for any of the reaction intermediates. Near-atomic resolution crystallography supported by in crystallo Raman spectroscopy and QM/MM calculations showed unambiguously that the archetypical cofactor-free uricase catalyzes uric acid degradation via a C5(S)-(hydro)peroxide intermediate. Low X-ray doses break specifically the intermediate C5OO(H) bond at 100 K, thus releasing O2 in situ, which is trapped above the substrate radical. The dose-dependent rate of bond rupture followed by combined crystallographic and Raman analysis indicates that ionizing radiation kick-starts both peroxide decomposition and its regeneration. Peroxidation can be explained by a mechanism in which the substrate radical recombines with superoxide transiently produced in the active site.

Subject Areas: Biology and Bio-materials


Instruments: I02-Macromolecular Crystallography

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