Frataxin (Yfh1 in yeast) is a conserved
protein and deficiency leads to the
neurodegenerative disease Friedreich's ataxia.
Frataxin is a critical
protein for Fe-S cluster assembly in mitochondria, interacting with other components of the Fe-S cluster machinery, including
cysteine desulfurase Nfs1, Isd11 and the Isu1 scaffold
protein. Yeast Isu1 with the
methionine to
isoleucine substitution (M141I), in which the E. coli
amino acid is inserted at this position, corrected most of the phenotypes that result from lack of Yfh1 in yeast. This suppressor Isu1 behaved as a genetic dominant. Furthermore
frataxin-bypass activity required a completely functional Nfs1 and correlated with the presence of efficient scaffold function. A screen of random Isu1 mutations for
frataxin-bypass activity identified only M141 substitutions, including
Ile, Cys, Leu, or Val. In each case, mitochondrial Nfs1 persulfide formation was enhanced, and mitochondrial Fe-S cluster assembly was improved in the absence of
frataxin. Direct targeting of the entire E. coli
IscU to ∆yfh1 mitochondria also ameliorated the mutant phenotypes. In contrast, expression of
IscU with the reverse substitution i.e.
IscU with Ile to Met change led to worsening of the ∆yfh1 phenotypes, including severely compromised growth, increased sensitivity to
oxygen, deficiency in Fe-S clusters and
heme, and impaired
iron homeostasis. A bioinformatic survey of eukaryotic Isu1/prokaryotic
IscU database entries sorted on the
amino acid utilized at the M141 position identified unique groupings, with virtually all of the eukaryotic scaffolds using Met, and the preponderance of prokaryotic scaffolds using other
amino acids. The
frataxin-bypassing
amino acids Cys, Ile, Leu, or Val, were found predominantly in prokaryotes. This
amino acid position 141 is unique in Isu1, and the
frataxin-bypass effect likely mimics a conserved and ancient feature of the prokaryotic Fe-S cluster assembly machinery.