The So-Called Listeria innocua Ferritin Is a Dps Protein. Iron Incorporation, Detoxification, and DNA Protection Properties

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Abstract
Listeria innocua Dps (DNA binding protein from starved cells) affords protection to DNA against oxidative damage and can accumulate about 500 iron atoms within its central cavity through a process facilitated by a ferroxidase center. The chemistry of iron binding and oxidation in Listeria Dps (LiDps, formerly described as a ferritin) using H2O2 as oxidant was studied to further define the mechanism of iron deposition inside the protein and the role of LiDps in protecting DNA from oxidative damage. The relatively strong binding of 12 Fe2+ to the apoprotein (KD ∼ 0.023 μM) was demonstrated by isothermal titration calorimetry, fluorescence quenching, and pH stat experiments. Hydrogen peroxide was found to be a more efficient oxidant for the protein-bound Fe2+ than O2. Iron(II) oxidation by H2O2 occurs with a stoichiometry of 2 Fe2+/H2O2 in both the protein-based ferroxidation and subsequent mineralization reactions, indicating complete reduction of H2O2 to H2O. Electron paramagnetic resonance (EPR) spin-trapping experiments demonstrated that LiDps attenuates the production of hydroxyl radical by Fenton chemistry. DNA cleavage assays showed that the protein, while not binding to DNA itself, protects it against the deleterious combination of Fe2+ and H2O2. The overall process of iron deposition and detoxification by LiDps is described by the following equations. For ferroxidation, Fe2+ + DpsZ → [(Fe2+)-Dps]Z+1 + H+ (Fe2+ binding) and [(Fe2+)-Dps]Z+1 + Fe2+ + H2O2 → [(Fe3+)2(O)2-Dps]Z+1 + 2H+ (Fe2+ oxidation/hydrolysis). For mineralization, 2Fe2+ + H2O2 + 2H2O → 2Fe(O)OH(core) + 4H+ (Fe2+ oxidation/hydrolysis). These reactions occur in place of undesirable odd-electron redox processes that produce hydroxyl radical.