Human frataxin has a vital role in the biosynthesis of iron–sulfur (Fe-S) clusters in mitochondria and its deficiency causes the neurodegenerative disease Friedreich’s ataxia. Proposed functions for frataxin in the Fe-S pathway include iron donation to the Fe-S cluster machinery and regulation of cysteine desulfurase activity to control the rate of Fe-S production, although further molecular detail is required to distinguish these two possibilities. It is well established that frataxin can coordinate iron using glutamate and aspartate side chains on the protein surface; however, in this work we identify a new iron coordinating residue in the Nterminus of human frataxin using complementary spectroscopic and structural approaches. Further, we demonstrate that His86 in this N-terminal region is required for high affinity iron coordination and iron assembly of Fe-S clusters by ISCU as part of the Fe-S cluster biosynthetic complex. If a binding site that includes His86 is important for Fe-S cluster synthesis as part of its chaperone function, this raises the possibility that either iron binding at the acidic surface of frataxin may be spurious or that it is required for protein–protein interactions with the Fe-S biosynthetic quaternary complex. Our data suggest that iron coordination to frataxin may be significant to the Fe-S cluster biosynthesis pathway in mitochondria.
The progressive neurodegenerative disease Friedreich's ataxia is caused by a decreased level of expression of frataxin, a putative iron chaperone. Frataxin is thought to transiently interact with ISU, the scaffold protein onto which iron-sulfur clusters are assembled, to deliver ferrous iron. Photoreactive heterotrifunctional chemical cross-linking confirmed the interaction between frataxin and ISU in the presence of iron and validated that transient interactions can be covalently trapped with this method. Because frataxin may participate in transient interactions with other mitochondrial proteins, this cross-linking approach may reveal new protein partners and pathways in which it interacts and help deduce direct, downstream consequences of its deficiency.
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