title: Arabidopsis chloroplastic glutaredoxin C5 as a model to explore molecular determinants for iron-sulfur cluster binding into glutaredoxins.
abstract: Unlike thioredoxins, glutaredoxins are involved in iron-sulfur cluster assembly and in reduction of specific disulfides (i.e. protein-glutathione adducts), and thus they are also important redox regulators of chloroplast metabolism. Using GFP fusion, AtGrxC5 isoform, present exclusively in Brassicaceae, was shown to be localized in chloroplasts. A comparison of the biochemical, structural, and spectroscopic properties of Arabidopsis GrxC5 (WCSYC active site) with poplar GrxS12 (WCSYS active site), a chloroplastic paralog, indicated that, contrary to the solely apomonomeric GrxS12 isoform, AtGrxC5 exists as two forms when expressed in Escherichia coli. The monomeric apoprotein possesses deglutathionylation activity mediating the recycling of plastidial methionine sulfoxide reductase B1 and peroxiredoxin IIE, whereas the dimeric holoprotein incorporates a [2Fe-2S] cluster. Site-directed mutagenesis experiments and resolution of the x-ray crystal structure of AtGrxC5 in its holoform revealed that, although not involved in its ligation, the presence of the second active site cysteine (Cys(32)) is required for cluster formation. In addition, thiol titrations, fluorescence measurements, and mass spectrometry analyses showed that, despite the presence of a dithiol active site, AtGrxC5 does not form any inter- or intramolecular disulfide bond and that its activity exclusively relies on a monothiol mechanism.
authors.affiliation: Unité Mixte de Recherches 1136, Institut National de la Recherche Agronomique-Nancy Université, Interactions Arbres Microorganismes, Institut Fédératif de Recherche 110 Ecosystèmes Forestiers, Agroressources, Biomolécule et Alimentation, 54506 Vandoeuvre-lès-Nancy Cedex, France.
authors.forename: Jérémy
authors.initials: J
authors.lastname: Couturier
authors.forename: Elke
authors.initials: E
authors.lastname: Ströher
authors.forename: Angela-Nadia
authors.initials: AN
authors.lastname: Albetel
authors.forename: Thomas
authors.initials: T
authors.lastname: Roret
authors.forename: Meenakumari
authors.initials: M
authors.lastname: Muthuramalingam
authors.forename: Lionel
authors.initials: L
authors.lastname: Tarrago
authors.forename: Thorsten
authors.initials: T
authors.lastname: Seidel
authors.forename: Pascale
authors.initials: P
authors.lastname: Tsan
authors.forename: Jean-Pierre
authors.initials: JP
authors.lastname: Jacquot
authors.forename: Michael K
authors.initials: MK
authors.lastname: Johnson
authors.forename: Karl-Josef
authors.initials: KJ
authors.lastname: Dietz
authors.forename: Claude
authors.initials: C
authors.lastname: Didierjean
authors.forename: Nicolas
authors.initials: N
authors.lastname: Rouhier
mesh_terms: D017360:Arabidopsis / Q000235:genetics / Q000378:metabolism*; D002736:Chloroplasts / Q000378:metabolism*; D003001:Cloning, Molecular; D018360:Crystallography, X-Ray; D054477:Glutaredoxins / Q000235:genetics / Q000378:metabolism*; D007506:Iron-Sulfur Proteins / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D021241:Spectrometry, Mass, Electrospray Ionization; D013347:Subcellular Fractions / Q000378:metabolism
chemical_list: D054477:Glutaredoxins; D007506:Iron-Sulfur Proteins