TitleMonothiol glutaredoxins can bind linear [Fe3S4]+ and [Fe4S4]2+ clusters in addition to [Fe2S2]2+ clusters: spectroscopic characterization and functional implications.
Publication TypeJournal Article
Year of Publication2013
AuthorsZhang, B, Bandyopadhyay, S, Shakamuri, P, Naik, SG, Huynh, BHanh, Couturier, J, Rouhier, N, Johnson, MK
JournalJ Am Chem Soc
Volume135
Issue40
Pagination15153-64
Date Published2013 Oct 9
ISSN1520-5126
KeywordsAconitate Hydratase, Apoenzymes, Enzyme Activation, Glutaredoxins, Glutathione, iron, Protein Binding, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Spectrum Analysis, sulfur
Abstract

Saccharomyces cerevisiae mitochondrial glutaredoxin 5 (Grx5) is the archetypical member of a ubiquitous class of monothiol glutaredoxins with a strictly conserved CGFS active-site sequence that has been shown to function in biological [Fe2S2](2+) cluster trafficking. In this work, we show that recombinant S. cerevisiae Grx5 purified aerobically, after prolonged exposure of the cell-free extract to air or after anaerobic reconstitution in the presence of glutathione, predominantly contains a linear [Fe3S4](+) cluster. The excited-state electronic properties and ground-state electronic and vibrational properties of the linear [Fe3S4](+) cluster have been characterized using UV-vis absorption/CD/MCD, EPR, Mössbauer, and resonance Raman spectroscopies. The results reveal a rhombic S = 5/2 linear [Fe3S4](+) cluster with properties similar to those reported for synthetic linear [Fe3S4](+) clusters and the linear [Fe3S4](+) clusters in purple aconitase. Moreover, the results indicate that the Fe-S cluster content previously reported for many monothiol Grxs has been misinterpreted exclusively in terms of [Fe2S2](2+) clusters, rather than linear [Fe3S4](+) clusters or mixtures of linear [Fe3S4](+) and [Fe2S2](2+) clusters. In the absence of GSH, anaerobic reconstitution of Grx5 yields a dimeric form containing one [Fe4S4](2+) cluster that is competent for in vitro activation of apo-aconitase, via intact cluster transfer. The ligation of the linear [Fe3S4](+) and [Fe4S4](2+) clusters in Grx5 has been assessed by spectroscopic, mutational, and analytical studies. Potential roles for monothiol Grx5 in scavenging and recycling linear [Fe3S4](+) clusters released during protein unfolding under oxidative stress conditions and in maturation of [Fe4S4](2+) cluster-containing proteins are discussed in light of these results.

DOI10.1021/ja407059n
Alternate JournalJ. Am. Chem. Soc.
PubMed ID24032439
PubMed Central IDPMC3836218
Grant ListGM47295 / GM / NIGMS NIH HHS / United States
GM62524 / GM / NIGMS NIH HHS / United States
R01 GM062524 / GM / NIGMS NIH HHS / United States
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