Iron Incorporation in Streptococcus suis Dps-like Peroxide Resistance Protein Dpr Requires Mobility in the Ferroxidase Center and Leads to the Formation of a Ferrihydrite-like Core

The Dps-like peroxide resistance protein (Dpr) is a dodecameric protein that protects the human and swine pathogen Streptococcus suis from hydrogen peroxide by removing free Fe 2+ from the cytosol. Subsequent oxidation of iron by Dpr results in the deposition of Fe 3+ inside the protein's central cavity. Structural changes that occur in the ferroxidase center were studied by X-ray crystallography after soaking Dpr crystals with Fe 2+ in the presence of sodium dithionite. Twelve iron-binding sites were identified with each site formed by residues Asp74 and Glu78 from one subunit, and Asp63, His47 and His59 from a 2-fold symmetry-related subunit. Compared to the iron-free Dpr, Asp74 and Glu78 were found to be the most flexible amino acid residues and able to adopt a variety of conformations in different subunits. The crystal structure of an Asp74Ala Dpr mutant soaked with a Fe 2+ -solution revealed variations in the Asp63 position and no iron bound to the ferroxidase center. These results indicate an intrinsic flexibility in the active site that may be important for the catalytic reaction and subsequent nucleation events. Two iron cores with remarkably different features were identified in Dpr using X-ray absorption spectroscopy. Purified Dpr was found to have a small-size iron core with only ∼ 16 iron atoms/dodecamer forming a ferritin-like ferrihydrite structure. Because of its size, this core represents the smallest iron core identified so far in ferritins and other Dps-like proteins. A large-size core (∼ 180 iron atoms/dodecamer) formed after incubating the protein with a ferrous solution shows differences in iron coordination compared to the small size core. Characterization of the two iron cores in Dpr could provide insights into nucleation events and the mechanism of iron core growth in the Dps family of proteins.