TitleHistidine 103 in Fra2 is an iron-sulfur cluster ligand in the [2Fe-2S] Fra2-Grx3 complex and is required for in vivo iron signaling in yeast.
Publication TypeJournal Article
Year of Publication2011
AuthorsLi, H, Mapolelo, DT, Dingra, NN, Keller, G, Riggs-Gelasco, PJ, Winge, DR, Johnson, MK, Outten, CE
JournalJ Biol Chem
Volume286
Issue1
Pagination867-76
Date Published2011 Jan 7
ISSN1083-351X
KeywordsAmino Acid Sequence, Animals, Apoproteins, Histidine, Humans, Intracellular Signaling Peptides and Proteins, iron, Ligands, Mice, Molecular Sequence Data, mutagenesis, mutation, Oxidoreductases, Protein Multimerization, Protein Stability, Protein Structure, Quaternary, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Signal Transduction, Spectrum Analysis, sulfur, Transcription Factors
Abstract

The BolA homologue Fra2 and the cytosolic monothiol glutaredoxins Grx3 and Grx4 together play a key role in regulating iron homeostasis in Saccharomyces cerevisiae. Genetic studies indicate that Grx3/4 and Fra2 regulate activity of the iron-responsive transcription factors Aft1 and Aft2 in response to mitochondrial Fe-S cluster biosynthesis. We have previously shown that Fra2 and Grx3/4 form a [2Fe-2S](2+)-bridged heterodimeric complex with iron ligands provided by the active site cysteine of Grx3/4, glutathione, and a histidine residue. To further characterize this unusual Fe-S-binding complex, site-directed mutagenesis was used to identify specific residues in Fra2 that influence Fe-S cluster binding and regulation of Aft1 activity in vivo. Here, we present spectroscopic evidence that His-103 in Fra2 is an Fe-S cluster ligand in the Fra2-Grx3 complex. Replacement of this residue does not abolish Fe-S cluster binding, but it does lead to a change in cluster coordination and destabilization of the [2Fe-2S] cluster. In vivo genetic studies further confirm that Fra2 His-103 is critical for control of Aft1 activity in response to the cellular iron status. Using CD spectroscopy, we find that ∼1 mol eq of apo-Fra2 binds tightly to the [2Fe-2S] Grx3 homodimer to form the [2Fe-2S] Fra2-Grx3 heterodimer, suggesting a mechanism for formation of the [2Fe-2S] Fra2-Grx3 heterodimer in vivo. Taken together, these results demonstrate that the histidine coordination and stability of the [2Fe-2S] cluster in the Fra2-Grx3 complex are essential for iron regulation in yeast.

DOI10.1074/jbc.M110.184176
Alternate JournalJ. Biol. Chem.
PubMed ID20978135
PubMed Central IDPMC3013046
Grant ListES03817 / ES / NIEHS NIH HHS / United States
ES13780 / ES / NIEHS NIH HHS / United States
GM62524 / GM / NIGMS NIH HHS / United States
K22 ES013780 / ES / NIEHS NIH HHS / United States
P20RR016461 / RR / NCRR NIH HHS / United States
R01 GM062524-11 / GM / NIGMS NIH HHS / United States
R01 GM062524-12 / GM / NIGMS NIH HHS / United States
R01 GM083292-18 / GM / NIGMS NIH HHS / United States
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