The ferroxidase activity of yeast frataxin

Frataxin is required for maintenance of normal mitochondrial iron levels and respiration. The mature form of yeast frataxin (mYfh1p) assembles stepwise into a multimer of 840 kDa (α₄₈) that accumulates iron in a water-soluble form. Here, two distinct iron oxidation reactions are shown to take place during the initial assembly step (α → α₃). A ferroxidase reaction with a stoichiometry of 2 Fe(II)/O₂ is detected at Fe(II)/mYfh1p ratios of ≤0.5. Ferroxidation is progressively overcome by autoxidation at Fe(II)/mYfh1p ratios of >0.5. Gel filtration analysis indicates that an oligomer of mYfh1p, α₃, is responsible for both reactions. The observed 2 Fe(II)/O₂ stoichiometry implies production of H₂O₂ during the ferroxidase reaction. However, only a fraction of the expected total H₂O₂ is detected in solution. Oxidative degradation of mYfh1p during the ferroxidase reaction suggests that most H₂O₂ reacts with the protein. Accordingly, the addition of mYfhlp to a mixture of Fe(II) and H₂O₂ results in significant attenuation of Fenton chemistry. Multimer assembly is fully inhibited under anaerobic conditions, indicating that mYfh1p is activated by Fe(II) in the presence of O₂. This combination induces oligomerization and mYfh1p-catalyzed Fe(II) oxidation, starting a process that ultimately leads to the sequestration of as many as 50 Fe(II)/subunit inside the multimer.