The ferroxidase activity of yeast frataxin

Abstract

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.