Comparative Fe and Zn K-edge X-ray absorption spectroscopic study of the ferroxidase centres of human H-chain ferritin and bacterioferritin from Desulfovibrio desulfuricans |
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Authors: | L Toussaint M G Cuypers L Bertrand L Hue C V Romão L M Saraiva M Teixeira W Meyer-Klaucke M C Feiters R R Crichton |
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Institution: | 1. Unité de Biochimie, Institut des Sciences de la Vie, Université Catholique de Louvain, Croix du Sud, 4-5, boite 3, 1348, Louvain-la-Neuve, Belgium 2. Macromolecular Crystallography Group, ESRF, BP-220, 38043, Grenoble Cedex, France 3. Unité Hormones et Métabolisme, Université Catholique de Louvain, UCL 75-29, Avenue Hippocrate 75, 1200, Brussels, Belgium 4. Division of Cardiology, Université Catholique de Louvain, UCL 53-49, Avenue E. Mounier 52, 1200, Brussels, Belgium 5. Instituto de Tecnologia Química e Biológica, Universidade Nova de Lisboa, Av. República, EAN, Apartado 127, 2781-901, Oeiras, Portugal 6. EMBL, Outstation Hamburg, Notkestrasse 85, 22603, Hamburg, Germany 7. Department of Organic Chemistry, Institute for Molecules and Materials, Radboud University Nijmegen, 135 Heyendaalseweg, 6525 AJ, Nijmegen, The Netherlands
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Abstract: | Iron uptake by the ubiquitous iron-storage protein ferritin involves the oxidation of two Fe(II) ions located at the highly conserved dinuclear “ferroxidase centre” in individual subunits. We have measured X-ray absorption spectra of four mutants (K86Q, K86Q/E27D, K86Q/E107D, and K86Q/E27D/E107D, involving variations of Glu to Asp on either or both sides of the dinuclear ferroxidase site) of recombinant human H-chain ferritin (rHuHF) in their complexes with reactive Fe(II) and redox-inactive Zn(II). The results for Fe–rHuHf are compared with those for recombinant Desulfovibrio desulfuricans bacterioferritin (DdBfr) in three states: oxidised, reduced, and oxidised/Chelex®-treated. The X-ray absorption near-edge region of the spectrum allows the oxidation state of the iron ions to be assessed. Extended X-ray absorption fine structure simulations have yielded accurate geometric information that represents an important refinement of the crystal structure of DdBfr; most metal–ligand bonds are shortened and there is a decrease in ionic radius going from the Fe(II) to the Fe(III) state. The Chelex®-treated sample is found to be partly mineralised, giving an indication of the state of iron in the cycled-oxidised (reduced, then oxidised) form of DdBfr, where the crystal structure shows the dinuclear site to be only half occupied. In the case of rHuHF the complexes with Zn(II) reveal a surprising similarity between the variants, indicating that the rHuHf dinuclear site is rigid. In spite of this, the rHuHf complexes with Fe(II) show a variation in reactivity that is reflected in the iron oxidation states and coordination geometries. |
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Keywords: | Ferritin Iron-storage protein Non-haem iron Diiron ferroxidase centre Zinc-substituted protein |
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