Interplay between glutathione, Atx1 and copper. 1. Copper(I) glutathionate induced dimerization of Atx1.
Copper is both an essential element as a catalytic cofactor and a toxic element because of its redox properties. Once in the cell, Cu(I) binds to glutathione (GSH) and various thiol-rich proteins that sequester and/or exchange copper with other intracellular components. Among them, the Cu(I) chaperone Atx1 is known to deliver Cu(I) to Ccc2, the Golgi Cu-ATPase, in yeast. However, the mechanism for Cu(I) incorporation into Atx1 has not yet been unraveled. We investigated here a possible role of GSH in Cu(I) binding to Atx1. Yeast Atx1 was expressed in Escherichia coli and purified to study its ability to bind Cu(I). We found that with an excess of GSH [at least two GSH/Cu(I)], Atx1 formed a Cu(I)-bridged dimer of high affinity for Cu(I), containing two Cu(I) and two GSH, whereas no dimer was observed in the absence of GSH. The stability constants (log beta) of the Cu(I) complexes measured at pH 6 were 15-16 and 49-50 for CuAtx1 and Cu (2) (I) (GS(-))(2)(Atx1)(2), respectively. Hence, these results suggest that in vivo the high GSH concentration favors Atx1 dimerization and that Cu (2) (I) (GS(-))(2)(Atx1)(2) is the major conformation of Atx1 in the cytosol.
Miras
Roger
R
Laboratoire de Chimie et Biologie des Métaux, CEA, DSV, iRTSV, 17 rue des Martyrs, Grenoble, 38054, France.
Morin
Isabelle
I
Jacquin
Olivier
O
Cuillel
Martine
M
Guillain
Florent
F
Mintz
Elisabeth
E
D000042:Absorption; D002352:Carrier Proteins / Q000737:chemistry* / Q000378:metabolism*; D003300:Copper / Q000378:metabolism*; D019281:Dimerization; D005978:Glutathione / Q000378:metabolism*; D011485:Protein Binding; D020836:Protein Structure, Quaternary; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000378:metabolism*; D013379:Substrate Specificity
C093341:ATX1 protein, S cerevisiae; D002352:Carrier Proteins; D029701:Saccharomyces cerevisiae Proteins; D003300:Copper; D005978:Glutathione