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Yeast metallothionein. Sequence and metal-binding properties.

D R Winge, K B Nielson, W R Gray

    The Journal of Biological Chemistry
    |November 25, 1985
    PubMed
    Summary

    Yeast metallothionein, a 53-residue protein, binds 8 copper ions and exhibits distinct metal-binding configurations for copper and cadmium, similar to mammalian proteins.

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    Area of Science:

    • Biochemistry
    • Molecular Biology
    • Yeast Genetics

    Background:

    • The CUP1 locus in Saccharomyces cerevisiae encodes a copper-resistant protein.
    • Metallothioneins are metal-binding proteins involved in heavy metal detoxification and homeostasis.
    • Understanding yeast metallothionein structure and function provides insights into metal-binding mechanisms.

    Purpose of the Study:

    • To purify and characterize the protein product of the CUP1 locus in copper-resistant yeast.
    • To determine the primary structure and metal-binding properties of yeast metallothionein.
    • To compare the metal-binding configurations of yeast metallothionein with mammalian counterparts.

    Main Methods:

    • Protein purification from Saccharomyces cerevisiae.
    • Amino-terminal sequencing and molecular weight determination.
    • Metal ion binding studies (copper, silver, cadmium, zinc) using reconstitution and metal ion exchange assays.
    • Proteolysis protection assays.

    Main Results:

    • Yeast metallothionein is a 53-residue polypeptide with a molecular weight of 5655, lacking the first 8 predicted amino acids.
    • The protein binds 8 copper ions, coordinated by 12 cysteines per molecule.
    • Distinct binding stoichiometries were observed for different metal ions: 8 for Cu(I) and Ag(I), and 4 for Cd(II) and Zn(II).
    • Yeast metallothionein displays two distinct metal-binding configurations, analogous to mammalian metallothioneins.

    Conclusions:

    • Yeast metallothionein possesses a unique N-terminus and a defined structure for metal binding.
    • The protein exhibits differential binding affinities and stoichiometries for various metal ions.
    • The findings highlight conserved mechanisms of metal ion coordination in metallothioneins across different species.

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