Human GLRX5 (glutaredoxin 5) is an evolutionarily conserved thiol–disulfide oxidoreductase that has a direct role in the maintenance of normal cytosolic and mitochondrial iron homoeostasis, and its expression affects haem biosynthesis and erythropoiesis. We have crystallized the human GLRX5 bound to two [2Fe–2S] clusters and four GSH molecules. The crystal structure revealed a tetrameric organization with the [2Fe–2S] clusters buried in the interior and shielded from the solvent by the conserved β1-α2 loop, Phe69 and the GSH molecules. Each [2Fe–2S] cluster is ligated by the N-terminal activesite cysteine (Cys67) thiols contributed by two protomers and two cysteine thiols from two GSH. The two subunits co-ordinating the cluster are in a more extended conformation compared with iron–sulfur-bound human GLRX2, and the intersubunit interactions are more extensive and involve conserved residues among monothiol GLRXs. Gel-filtration chromatography and analytical ultracentrifugation support a tetrameric organization of holo-GLRX5, whereas the apoprotein is monomeric. MS analyses revealed glutathionylation of the cysteine residues in the absence of the [2Fe–2S] cluster, which would protect them from further oxidation and possibly facilitate cluster transfer/acceptance. Apo-GLRX5 reduced glutathione mixed disulfides with a rate 100 times lower than did GLRX2 and was active as a glutathione-dependent electron donor for mammalian ribonucleotide reductase.
The crystal structure of human GLRX5: iron–sulfur cluster co-ordination, tetrameric assembly and monomer activity
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Catrine Johansson, Annette K. Roos, Sergio J. Montano, Rajib Sengupta, Panagis Filippakopoulos, Kunde Guo, Frank von Delft, Arne Holmgren, Udo Oppermann, Kathryn L. Kavanagh; The crystal structure of human GLRX5: iron–sulfur cluster co-ordination, tetrameric assembly and monomer activity. Biochem J 15 January 2011; 433 (2): 303–311. doi: https://doi.org/10.1042/BJ20101286
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