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Redox sulfur chemistry of the copper chaperone Atox1 is regulated by the enzyme glutaredoxin 1, the reduction potential of the glutathione couple GSSG/2GSH and the availability of Cu(I)

Brose, Jens, La Fontaine, Sharon, Wedd, Anthony G. and Xiao, Zhiguang 2014, Redox sulfur chemistry of the copper chaperone Atox1 is regulated by the enzyme glutaredoxin 1, the reduction potential of the glutathione couple GSSG/2GSH and the availability of Cu(I), Metallomics, vol. 6, no. 4, pp. 793-808, doi: 10.1039/c4mt00020j.

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Title Redox sulfur chemistry of the copper chaperone Atox1 is regulated by the enzyme glutaredoxin 1, the reduction potential of the glutathione couple GSSG/2GSH and the availability of Cu(I)
Author(s) Brose, Jens
La Fontaine, Sharon
Wedd, Anthony G.
Xiao, Zhiguang
Journal name Metallomics
Volume number 6
Issue number 4
Start page 793
End page 808
Total pages 16
Publisher Royal Society of Chemistry Publications
Place of publication Cambridge, Eng.
Publication date 2014-04
ISSN 1756-591X
Keyword(s) Copper
Glutaredoxins
Glutathione
Humans
Metallochaperones
Models, Molecular
Oxidation-Reduction
Sulfur
Thermodynamics
Science & Technology
Life Sciences & Biomedicine
Biochemistry & Molecular Biology
NMR SOLUTION STRUCTURE
ESCHERICHIA-COLI
MIXED DISULFIDE
S-GLUTATHIONYLATION
THIOREDOXIN FAMILY
DISEASE PROTEIN
BLOOD-CELLS
THIOLTRANSFERASE
PURIFICATION
TRAFFICKING
Summary Glutaredoxins have been characterised as enzymes regulating the redox status of protein thiols via cofactors GSSG/GSH. However, such a function has not been demonstrated with physiologically relevant protein substrates in in vitro experiments. Their active sites frequently feature a Cys-xx-Cys motif that is predicted not to bind metal ions. Such motifs are also present in copper-transporting proteins such as Atox1, a human cytosolic copper metallo-chaperone. In this work, we present the first demonstration that: (i) human glutaredoxin 1 (hGrx1) efficiently catalyses interchange of the dithiol and disulfide forms of the Cys(12)-xx-Cys(15) fragment in Atox1 but does not act upon the isolated single residue Cys(41); (ii) the direction of catalysis is regulated by the GSSG/2GSH ratio and the availability of Cu(I); (iii) the active site Cys(23)-xx-Cys(26) in hGrx1 can bind Cu(I) tightly with femtomolar affinity (K(D) = 10(-15.5) M) and possesses a reduction potential of E(o)' = -118 mV at pH 7.0. In contrast, the Cys(12)-xx-Cys(15) motif in Atox1 has a higher affinity for Cu(I) (K(D) = 10(-17.4) M) and a more negative potential (E(o)' = -188 mV). These differences may be attributed primarily to the very low pKa of Cys23 in hGrx1 and allow rationalisation of conclusion (ii) above: hGrx1 may catalyse the oxidation of Atox1(dithiol) by GSSG, but not the complementary reduction of the oxidised Atox1(disulfide) by GSH unless Cu(aq)(+) is present at a concentration that allows binding of Cu(I) to reduced Atox1 but not to hGrx1. In fact, in the latter case, the catalytic preferences are reversed. Both Cys residues in the active site of hGrx1 are essential for the high affinity Cu(I) binding but the single Cys(23) residue only is required for the redox catalytic function. The molecular properties of both Atox1 and hGrx1 are consistent with a correlation between copper homeostasis and redox sulfur chemistry, as suggested by recent cell experiments. These proteins appear to have evolved the features necessary to fill multiple roles in redox regulation, Cu(I) buffering and Cu(I) transport.
Language eng
DOI 10.1039/c4mt00020j
Field of Research 030201 Bioinorganic Chemistry
Socio Economic Objective 970103 Expanding Knowledge in the Chemical Sciences
HERDC Research category C1 Refereed article in a scholarly journal
ERA Research output type C Journal article
Copyright notice ©2014, The Authors
Free to Read? Yes
Use Rights Creative Commons Attribution non-commercial licence
Persistent URL http://hdl.handle.net/10536/DRO/DU:30072396

Document type: Journal Article
Collections: School of Life and Environmental Sciences
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Every reasonable effort has been made to ensure that permission has been obtained for items included in DRO. If you believe that your rights have been infringed by this repository, please contact drosupport@deakin.edu.au.