title: An unusual mode of iron-sulfur-cluster coordination in a teleost glutaredoxin.
abstract: Glutaredoxins that contain a Cys-X-X-Cys active site motif are glutathione-dependent thiol-disulfide oxidoreductases. Vertebrate glutaredoxin 2 is characterized by two extra cysteines that form an intra-molecular disulfide bridge. Zebrafish glutaredoxin 2 contains four additional cysteines that are conserved within the infraclass of bony fish (teleosts). Here, we present a biochemical and biophysical characterization of zebrafish glutaredoxin 2, focusing on iron-sulfur-cluster coordination. The coordination of [2Fe2S](2+)-clusters in monomers of this protein was revealed by both absorption and Mössbauer spectroscopy as well as size exclusion chromatography. All other holo-glutaredoxins represent [FeS]-cluster bridged dimers using two molecules of non-covalently bound glutathione and the N-terminal active site cysteines as ligands. These cysteine residues were not required for [FeS]-cluster coordination in zebrafish glutaredoxin 2. A crystal structure of the teleost protein revealed high structural similarity to its human homologue. The two vertebrate-specific cysteines as well as two of the teleost-specific cysteines are positioned within a radius of 7Å near the C-terminus suggesting a potential role in [FeS]-cluster coordination. Indeed, mutated proteins lacking these teleost-specific cysteines lost the ability to bind the cofactor. Hence, the apparent mode of [FeS]-cluster coordination in zebrafish glutaredoxin 2 could be different from all yet described [FeS]-glutaredoxins.
authors.affiliation: Division for Biochemistry, Department for Medical Biochemistry and Biophysics, Karolinska Institutet, 17177 Stockholm, Sweden.
authors.forename: Lars
authors.initials: L
authors.lastname: Bräutigam
authors.forename: Catrine
authors.initials: C
authors.lastname: Johansson
authors.forename: Bastian
authors.initials: B
authors.lastname: Kubsch
authors.forename: Michael A
authors.initials: MA
authors.lastname: McDonough
authors.forename: Eckhard
authors.initials: E
authors.lastname: Bill
authors.forename: Arne
authors.initials: A
authors.lastname: Holmgren
authors.forename: Carsten
authors.initials: C
authors.lastname: Berndt
mesh_terms: D020816:Amino Acid Motifs; D000818:Animals; D020134:Catalytic Domain; D003545:Cysteine / Q000737:chemistry; D004789:Enzyme Activation; D054477:Glutaredoxins / Q000737:chemistry*; D006801:Humans; D007506:Iron-Sulfur Proteins / Q000737:chemistry*; D008024:Ligands; D008969:Molecular Sequence Data; D011485:Protein Binding; D055503:Protein Multimerization; D017386:Sequence Homology, Amino Acid; D015027:Zebrafish / Q000378:metabolism*; D029961:Zebrafish Proteins / Q000737:chemistry*
chemical_list: D054477:Glutaredoxins; D007506:Iron-Sulfur Proteins; D008024:Ligands; D029961:Zebrafish Proteins; D003545:Cysteine
keywords: Crystal structure; GSH; GSSG; Glutathione; Grx; HED; Mössbauer spectroscopy; Redox; Trx; Zebrafish; glutaredoxin; glutathion; glutathione disulphide; hydroxyethyl disulphide; thioredoxin