title: Superoxide reductase from Giardia intestinalis: structural characterization of the first SOR from a eukaryotic organism shows an iron centre that is highly sensitive to photoreduction.
abstract: Superoxide reductase (SOR), which is commonly found in prokaryotic organisms, affords protection from oxidative stress by reducing the superoxide anion to hydrogen peroxide. The reaction is catalyzed at the iron centre, which is highly conserved among the prokaryotic SORs structurally characterized to date. Reported here is the first structure of an SOR from a eukaryotic organism, the protozoan parasite Giardia intestinalis (GiSOR), which was solved at 2.0 Å resolution. By collecting several diffraction data sets at 100 K from the same flash-cooled protein crystal using synchrotron X-ray radiation, photoreduction of the iron centre was observed. Reduction was monitored using an online UV-visible microspectrophotometer, following the decay of the 647 nm absorption band characteristic of the iron site in the glutamate-bound, oxidized state. Similarly to other 1Fe-SORs structurally characterized to date, the enzyme displays a tetrameric quaternary-structure arrangement. As a distinctive feature, the N-terminal loop of the protein, containing the characteristic EKHxP motif, revealed an unusually high flexibility regardless of the iron redox state. At variance with previous evidence collected by X-ray crystallography and Fourier transform infrared spectroscopy of prokaryotic SORs, iron reduction did not lead to dissociation of glutamate from the catalytic metal or other structural changes; however, the glutamate ligand underwent X-ray-induced chemical changes, revealing high sensitivity of the GiSOR active site to X-ray radiation damage.
authors.affiliation: Instituto de Tecnologia Química e Biológica, António Xavier Universidade Nova de Lisboa, Avenida da República, 2780-157 Oeiras, Portugal.
authors.forename: Cristiana M
authors.initials: CM
authors.lastname: Sousa
authors.affiliation: Structural Biology Group, ESRF - The European Synchrotron, CS40220, 38043 Grenoble CEDEX 9, France.
authors.forename: Philippe
authors.initials: P
authors.lastname: Carpentier
authors.affiliation: Instituto de Tecnologia Química e Biológica, António Xavier Universidade Nova de Lisboa, Avenida da República, 2780-157 Oeiras, Portugal.
authors.forename: Pedro M
authors.initials: PM
authors.lastname: Matias
authors.affiliation: Department of Biochemical Sciences, Istituto Pasteur-Fondazione Cenci Bolognetti, Sapienza University of Rome, 00185 Rome, Italy.
authors.forename: Fabrizio
authors.initials: F
authors.lastname: Testa
authors.affiliation: Instituto de Biologia Experimental e Tecnológica, Apartado 12, 2781-901 Oeiras, Portugal.
authors.forename: Filipa
authors.initials: F
authors.lastname: Pinho
authors.affiliation: Department of Biochemical Sciences, Istituto Pasteur-Fondazione Cenci Bolognetti, Sapienza University of Rome, 00185 Rome, Italy.
authors.forename: Paolo
authors.initials: P
authors.lastname: Sarti
authors.affiliation: CNR Institute of Molecular Biology and Pathology, 00185 Rome, Italy.
authors.forename: Alessandro
authors.initials: A
authors.lastname: Giuffrè
authors.affiliation: Instituto de Tecnologia Química e Biológica, António Xavier Universidade Nova de Lisboa, Avenida da República, 2780-157 Oeiras, Portugal.
authors.forename: Tiago M
authors.initials: TM
authors.lastname: Bandeiras
authors.affiliation: Instituto de Tecnologia Química e Biológica, António Xavier Universidade Nova de Lisboa, Avenida da República, 2780-157 Oeiras, Portugal.
authors.forename: Célia V
authors.initials: CV
authors.lastname: Romão
mesh_terms: D000595:Amino Acid Sequence; D020134:Catalytic Domain / Q000528:radiation effects; D018360:Crystallography, X-Ray; D016829:Giardia lamblia / Q000737:chemistry / Q000201:enzymology*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D010088:Oxidoreductases / Q000737:chemistry* / Q000378:metabolism*; D011487:Protein Conformation; D016415:Sequence Alignment; D014965:X-Rays
chemical_list: D010088:Oxidoreductases; C121849:superoxide reductase
keywords: eukaryotes; oxidative stress; radiation damage; superoxide reductase; visible spectroscopy