Domain nature of metallothionein

• Published in: The Journal of biological chemistry Vol. 257; no. 7; pp. 3471 - 3476
• Main Authors: Winge, D R, Miklossy, K A
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 10.04.1982; American Society for Biochemistry and Molecular Biology
• Subjects: Amino Acid Sequence; Amino Acids - analysis; Animals; Cadmium - pharmacology; Cadmium Poisoning - metabolism; Edetic Acid; Kinetics; Liver - metabolism; Male; Metalloproteins - isolation & purification; Metallothionein - isolation & purification; Metallothionein - metabolism; Peptide Fragments - analysis; Protein Binding; Protein Conformation; Rats; Rats, Inbred Strains; Spectrophotometry, Ultraviolet
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/S0021-9258(18)34802-6

Metallothionein purified from the livers of rats injected with CdCl3 was cleaved by proteolysis into a 32-residue polypeptide that contained 4 bound Cd ions. Appearance of this fragment designated alpha requires prior treatment of metallothionein with EDTA to remove the Zn ions and destabilize the 3-metal cysteine cluster in the other domain. The half-molecule domain was not efficiently produced by proteolysis of native metallothionein. The Cd4-alpha fragment is asymmetric in shape, as is the parent molecule. NH2-terminal sequence analysis revealed that the alpha fragment starts at Lys 30. Since the same amino acids are released from the COOH terminus of intact thionein and the alpha fragment by carboxypeptidase Y, the alpha domain generated by digestion with subtilisin therefore comprises residues 30 through 61. The amino acid composition of the alpha polypeptide is consistent with the structure of the 4-metal cysteine cluster proposed by Otvos and Armitage ((1980) Proc. Natl. Acad. Sci. U. S. A. 77, 7094-7098). Metallothionein appears to consist of a 3-metal cysteine domain in the NH2-terminal half of the thionein molecule and the 4-metal cysteine domain in the COOH-terminal half.