title: Energetics of copper trafficking between the Atx1 metallochaperone and the intracellular copper transporter, Ccc2.
abstract: The Atx1 metallochaperone protein is a cytoplasmic Cu(I) receptor that functions in intracellular copper trafficking pathways in plants, microbes, and humans. A key physiological partner of the Saccharomyces cerevisiae Atx1 is Ccc2, a cation transporting P-type ATPase located in secretory vesicles. Here, we show that Atx1 donates its metal ion cargo to the first N-terminal Atx1-like domain of Ccc2 in a direct and reversible manner. The thermodynamic gradient for metal transfer is shallow (K(exchange) = 1.4 +/- 0.2), establishing that vectorial delivery of copper by Atx1 is not based on a higher copper affinity of the target domain. Instead, Atx1 allows rapid metal transfer to its partner. This equilibrium is unaffected by a 50-fold excess of the Cu(I) competitor, glutathione, indicating that Atx1 also protects Cu(I) from nonspecific reactions. Mechanistically, we propose that a low activation barrier for transfer between partners results from complementary electrostatic forces that ultimately orient the metal-binding loops of Atx1 and Ccc2 for formation of copper-bridged intermediates. These thermodynamic and kinetic considerations suggest that copper trafficking proteins overcome the extraordinary copper chelation capacity of the eukaryotic cytoplasm by catalyzing the rate of copper transfer between physiological partners. In this sense, metallochaperones work like enzymes, carefully tailoring energetic barriers along specific reaction pathways but not others.
authors.affiliation: Department of Chemistry, Northwestern University, Evanston, Illinois 60208-3113, USA.
authors.forename: D L
authors.initials: DL
authors.lastname: Huffman
authors.forename: T V
authors.initials: TV
authors.lastname: O'Halloran
mesh_terms: D001692:Biological Transport; D002352:Carrier Proteins; D027682:Cation Transport Proteins; D003001:Cloning, Molecular; D003300:Copper / Q000378:metabolism*; D000079922:Copper Transport Proteins; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D005978:Glutathione / Q000378:metabolism; D018832:Molecular Chaperones / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins
chemical_list: C093341:ATX1 protein, S cerevisiae; C092974:CCC2 protein, S cerevisiae; D002352:Carrier Proteins; D027682:Cation Transport Proteins; D000079922:Copper Transport Proteins; D005656:Fungal Proteins; D018832:Molecular Chaperones; D029701:Saccharomyces cerevisiae Proteins; D003300:Copper; D005978:Glutathione