Investigating the influence of histidine residues on the metal ion binding ability of the wheat metallothionein γ-Ec-1 domain

• Published in: Journal of inorganic biochemistry 2015-12, Vol.153, p.197-203
• Main Authors: Tarasava, Katsiaryna, Freisinger, Eva
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Binding Sites; Cadmium; Cadmium - pharmacology; Histidine; Histidine - chemistry; Histidine - genetics; Metal ion specificity; Metallothionein; Metallothionein - chemistry; Metallothionein - genetics; Metallothionein - metabolism; Molecular Sequence Data; Mutation; Protein Binding; Zinc; Zinc - pharmacology
• ISSN: 0162-0134; 1873-3344
• DOI: 10.1016/j.jinorgbio.2015.08.009

While Zn(II) and Cd(II) have similar geochemical and environmental properties, their biological properties are distinctively different as Cd(II) ions have very limited metabolic significance and are mostly even toxic, while Zn(II) ions belong to the most essential micronutrients. One of the key proteins involved in intracellular Zn(II) and Cd(II) binding are metallothioneins (MTs), small cysteine-rich proteins ubiquitously found in many different organisms. In the past two decades, also MT sequences from diverse species that contain histidine residues have been found, and His-metal ion coordination has been shown. It is not clear, however, why in some MTs parts of the Cys residues are replaced by His, while most other MTs only contain Cys residues for metal ion binding. To address this question, we used the γ-domain of the early-cysteine labeled (Ec-1) metallothionein from common wheat as a model system because its enclosed M2Cys6 cluster represents the smallest metal-thiolate cluster possible with divalent metal ions. Based on the known three-dimensional structure of the γ-domain we set about to investigate the influence of a single Cys-to-His mutation on the structure and metal ion binding abilities of this domain. Combined data obtained by mass spectrometry, UV, as well as NMR spectroscopy suggest a preference for Zn(II) versus Cd(II) ions in the histidine containing binding site. The γ-domain of the wheat Ec-1 metallothionein contains a M(II)2Cys6 cluster. Mutation of one of the Cys residues to His increases the metal ion selectivity of the binding sites resulting in a mixed ZnCd cluster with Zn(II) nearly exclusively coordinated to the Cys3His site in accordance with the HSAB principle. [Display omitted] •We studied the influence of a Cys-to-His mutation on the M(II)2Cys6 cluster of a wheat MT.•Wild-type and C20H-γ-Ec-1 bind both two Zn(II) or Cd(II) ions.•We observed selective formation of ZnCys3His and CdCys4 sites in the mixed ZnCd species.•In contrast to the wild-type, the C20H mutant forms mostly the mixed ZnCd species.•Data are supported by UV and NMR spectroscopy as well as mass spectrometry.