title: Localization and Spectroscopic Analysis of the Cu(I) Binding Site in Wheat Metallothionein Ec-1.
abstract: The early cysteine-labeled metallothionein (MT) from Triticum aestivum (common wheat), denoted Ec-1, features two structurally well-defined domains, γ and βE, coordinating two and four Zn(II) ions, respectively. While the protein is currently assumed to function mainly in zinc homeostasis, a low amount of copper ions was also recently detected in a native Ec-1 sample. To evaluate the observed copper binding in more detail, the recombinant Zn₆Ec-1 form was exposed to different amounts of Cu(I) ions and the resulting species characterized with spectroscopic methods. Data reveal that the first Cu(I) equivalent coordinates exclusively to the N-terminal γ-domain of the protein and replaces one Zn(II) ion. To analyze the ability of the γ-domain for coordination of monovalent metal ions in more detail, the γ-Ec-1 peptide fragment was incubated with increasing amounts of Cu(I) and the process monitored with UV-VIS, circular dichroism, and luminescence spectroscopy. Closely similar spectra are observed regardless if the apo- or the metal ion-loaded and, hence, pre-folded forms, were used for the titration experiments with Cu(I). The results indicate that low amounts of Cu(I) ions displace the two metal ions subsequently and stoichiometrically, despite the different coordination geometry requirements of Cu(I) and Zn(II).
authors.affiliation: Department of Chemistry, University of Zurich, 8057 Zurich, Switzerland. tarasava.katsiaryna@gmail.com.
authors.forename: Katsiaryna
authors.initials: K
authors.lastname: Tarasava
authors.affiliation: Department of Chemistry, University of Zurich, 8057 Zurich, Switzerland. jens.loebus@tosca-engineering.com.
authors.forename: Jens
authors.initials: J
authors.lastname: Loebus
authors.affiliation: Department of Chemistry, University of Zurich, 8057 Zurich, Switzerland. freisinger@chem.uzh.ch.
authors.forename: Eva
authors.initials: E
authors.lastname: Freisinger
mesh_terms: D001665:Binding Sites; D002942:Circular Dichroism; D003300:Copper / Q000737:chemistry / Q000378:metabolism*; D008668:Metallothionein / Q000737:chemistry* / Q000378:metabolism; D010940:Plant Proteins / Q000737:chemistry / Q000378:metabolism; D017434:Protein Structure, Tertiary; D014908:Triticum / Q000378:metabolism*; D015032:Zinc / Q000737:chemistry / Q000378:metabolism
chemical_list: D010940:Plant Proteins; D003300:Copper; D008668:Metallothionein; D015032:Zinc
keywords: copper ions; luminescence spectroscopy; metallothionein; spectroscopy; wheat; zinc ions