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 (alpha(48)) that accumulates iron in a water-soluble form. Here, two distinct iron oxidation reactions are shown to take place during the initial assembly step (alpha --> alpha(3)). A ferroxidase reaction with a stoichiometry of 2 Fe(II)/O(2) is detected at Fe(II)/mYfh1p ratios of < or = 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, alpha(3), is responsible for both reactions. The observed 2 Fe(II)/O(2) stoichiometry implies production of H(2)O(2) during the ferroxidase reaction. However, only a fraction of the expected total H(2)O(2) is detected in solution. Oxidative degradation of mYfh1p during the ferroxidase reaction suggests that most H(2)O(2) reacts with the protein. Accordingly, the addition of mYfh1p to a mixture of Fe(II) and H(2)O(2) 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 O(2). 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.
Highlights
The two major iron-utilizing processes in the cell, production of heme by ferrochelatase and the iron-sulfur cluster biosynthetic pathway, reside in the mitochondrial matrix
Recent studies have identified a handful of inner mitochondrial membrane proteins involved in mitochondrial iron transport (6 – 8) as well as a mitochondrial matrix ferritin involved in iron storage [9]
The mitochondrial matrix protein frataxin has been implicated in mitochondrial iron homeostasis [10, 11], but its precise function is still not known
Summary
Mouse models in which the frataxin gene is selectively inactivated in neuronal or cardiac tissue present multiple respiratory enzyme deficits and accumulate iron in mitochondria in a time-dependent manner [16] In agreement with these observations, evidence of abnormal cellular iron homeostasis, increased oxidative damage, and respiratory enzyme deficits has been reported for the human disease (reviewed in Ref. 17). Titration of the mature form of yeast frataxin (mYfh1p) with Fe(II) under aerobic conditions results in stepwise assembly of a 48-subunit multimer with a molecular mass of 840 kDa and a hydrodynamic radius of 11 nm that sequesters 50 atoms of iron/subunit and forms iron-rich cores with a diameter of 2– 4 nm [23, 24]. Our results support a direct role for frataxin in iron metabolism
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