Is hydrogen peroxide produced during iron(II) oxidation in mammalian apoferritins?

The ferritins are a class of iron storage and detoxification proteins that play a central role in the biological management of iron. These proteins ha ve a catalytic site, "the ferroxidase site", located on the H-type subunit that facilitates the oxidation of Fe(II) to Fe(III) by O-2. Measurements du ring the past 10 years on a number of vertebrate ferritins have provided ev idence that H2O2 is produced at this diiron ferroxidase site. Recently repo rted experiments using three different analytical methods with horse spleen ferritin (HoSF) have failed to detect H2O2 production in this protein [Lin dsay, S., Brosnahan, D., and Watt, G.D. (2001) Biochemistry 40, 3340-3347]. These findings contrast with earlier results reporting H2O2 production in HoSF [Xu, B., and Chasteen, N.D. (1991) J. Biol. Chem. 266, 19965-19970]. H ere a sensitive fluorescence assay and an assay based on O-2 evolution in t he presence of catalase were used to demonstrate that H2O2 is produced in H oSF as previously reported. However, because of the relatively few H-chain ferroxidase sites in HoSF and the reaction of H2O2 with the protein, H2O2 i s more difficult to detect in this ferritin than in recombinant human H-cha in ferritin (EuHF). The proper sequence of addition of reagents is importan t for measurement of the total amount of H2O2 produced during the ferroxida tion reaction.