Yeast frataxin sequentially chaperones and stores iron by coupling protein assembly with iron oxidation

We have investigated the mechanism of frataxin, a conserved mitochondrial protein involved in iron metabolism and neurodegenerative disease. Previous studies revealed that the yeast frataxin homologue (mYfh1p) is activated by Fe(II) in the presence of O-2 and assembles stepwise into a 48-subunit multimer (alpha(48)) that sequesters > 2000 atoms of iron in 2-4-nm cores structurally similar to ferritin iron cores. Here we show that mYfh1p assembly is driven by two sequential iron oxidation reactions: A ferroxidase reaction catalyzed by mYfh1p induces the first assembly step (alpha --> alpha(3)), followed by a slower autoxidation reaction that promotes the assembly of higher order oligomers yielding alpha(48). Depending on the ionic environment, stepwise assembly is associated with accumulation of 50 - 75 Fe(II)/ subunit. Initially, this Fe( II) is loosely bound to mYfh1p and can be readily mobilized by chelators or made available to the mitochondrial enzyme ferrochelatase to synthesize heme. Transfer of mYfh1p-bound Fe( II) to ferrochelatase occurs in the presence of citrate, a physiologic ferrous iron chelator, suggesting that the transfer involves an intermolecular interaction. If mYfh1p-bound Fe( II) is not transferred to a ligand, iron oxidation, and mineralization proceed to completion, Fe(III) becomes progressively less accessible, and a stable iron-protein complex is formed. Iron oxidation-driven stepwise assembly is a novel mechanism by which yeast frataxin can function as an iron chaperone or an iron store.