EPR and Mössbauer spectroscopy of intact mitochondria isolated from Yah1p-depleted Saccharomyces cerevisiae.
Yah1p, an [Fe 2S 2]-containing ferredoxin located in the matrix of Saccharomyces cerevisiae mitochondria, functions in the synthesis of Fe/S clusters and heme a prosthetic groups. EPR, Mossbauer spectroscopy, and electron microscopy were used to characterize the Fe that accumulates in Yah1p-depleted isolated intact mitochondria. Gal- YAH1 cells were grown in standard rich media (YPD and YPGal) under O 2 or argon atmospheres. Mitochondria were isolated anaerobically, then prepared in the as-isolated redox state, the dithionite-treated state, and the O 2-treated state. The absence of strong EPR signals from Fe/S clusters when Yah1p was depleted confirms that Yah1p is required in Fe/S cluster assembly. Yah1p-depleted mitochondria, grown with O 2 bubbling through the media, accumulated excess Fe (up to 10 mM) that was present as 2-4 nm diameter ferric nanoparticles, similar to those observed in mitochondria from yfh1Delta cells. These particles yielded a broad isotropic EPR signal centered around g = 2, characteristic of superparamagnetic relaxation. Treatment with dithionite caused Fe (3+) ions of the nanoparticles to become reduced and largely exported from the mitochondria. Fe did not accumulate in mitochondria isolated from cells grown under Ar; a significant portion of the Fe in these organelles was in the high-spin Fe (2+) state. This suggests that the O 2 used during growth of Gal- YAH1 cells is responsible, either directly or indirectly, for Fe accumulation and for oxidizing Fe (2+) --> Fe (3+) prior to aggregation. Models are proposed in which the accumulation of ferric nanoparticles is caused either by the absence of a ligand that prevents such precipitation in wild-type mitochondria or by a more oxidizing environment within the mitochondria of Yah1p-depleted cells exposed to O 2. The efficacy of reducing accumulated Fe along with chelating it should be considered as a strategy for its removal in diseases involving such accumulations.
Miao
Ren
R
Department of Chemistry, Texas A&M University, College Station, Texas 77843-3255, USA.
Martinho
Marlène
M
Morales
Jessica Garber
JG
Kim
Hansoo
H
Ellis
E Ann
EA
Lill
Roland
R
Hendrich
Michael P
MP
Münck
Eckard
E
Lindahl
Paul A
PA
D000325:Adrenodoxin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004578:Electron Spin Resonance Spectroscopy; D007501:Iron / Q000737:chemistry / Q000378:metabolism; D008928:Mitochondria / Q000378:metabolism* / Q000648:ultrastructure*; D010100:Oxygen / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015204:Spectroscopy, Mossbauer
D029701:Saccharomyces cerevisiae Proteins; C120375:YAH1 protein, S cerevisiae; D000325:Adrenodoxin; D007501:Iron; D010100:Oxygen