title: Methionine motifs of copper transport proteins provide general and flexible thioether-only binding sites for Cu(I) and Ag(I).
abstract: Cellular acquisition of copper in eukaryotic organisms is primarily accomplished through high-affinity copper transport proteins (Ctr). The extracellular N-terminal regions of both human and yeast Ctr1 contain multiple methionine residues organized in copper-binding Mets motifs. These motifs comprise combinations of methionine residues arranged in clusters of MXM and MXXM, where X can be one of several amino acids. Model peptides corresponding to 15 different Mets motifs were synthesized and determined to selectively bind Cu(I) and Ag(I), with no discernible affinity for divalent metal ions. These are rare examples of biological thioether-only metal binding sites. Effective dissociation constant (KD) values for the model Mets peptides and Cu(I) were determined by an ascorbic acid oxidation assay and validated through electrospray ionization mass spectrometry and range between 2 and 11 microM. Affinity appears to be independent of pH, the arrangement of the motif, and the composition of intervening amino acids, all of which reveal the generality and flexibility of the MX1-2MX1-2M domain. Circular dichroism spectroscopy, 1H-NMR spectroscopy, and X-ray absorption spectroscopy were also used to characterize the binding event. These results are intended to aid the development of the still unknown mechanism of copper transport across the cell membrane.
authors.affiliation: Department of Chemistry, Duke University, P.O. Box 90346, Durham, NC 27708, USA.
authors.forename: Jeffrey T
authors.initials: JT
authors.lastname: Rubino
authors.forename: Pamela
authors.initials: P
authors.lastname: Riggs-Gelasco
authors.forename: Katherine J
authors.initials: KJ
authors.lastname: Franz
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D003300:Copper / Q000737:chemistry*; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008715:Methionine / Q000737:chemistry* / Q000235:genetics; D008969:Molecular Sequence Data; D015394:Molecular Structure; D010084:Oxidation-Reduction; D010455:Peptides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012834:Silver / Q000737:chemistry*; D021241:Spectrometry, Mass, Electrospray Ionization; D013057:Spectrum Analysis / Q000379:methods
chemical_list: D002352:Carrier Proteins; D005656:Fungal Proteins; D010455:Peptides; D012834:Silver; D003300:Copper; D008715:Methionine