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K A Miklossy

Publications and source records attributed to K A Miklossy.

3 recordsLinked to original sources
PubMed1982

113Cd NMR study of a metallothionein fragment. Evidence for a two-domain structure.

A 32-residue polypeptide fragment, designated alpha I, of rat liver metallothionein obtained by subtilisin digestion was studied by 113Cd NMR. The amino acid composition of the fragment corresponded to residues 30-61 of the metallothionein primary structure, and it contained 3.4 g atoms of Cd2+/mol of alpha I-fragment. Four 113Cd resonances were observed, three of which had identical chemical shifts to those assigned to the four-metal cluster in human liver metallothionein-2 under the same pH and buffer conditions. The 5-ppm chemical shift difference between the remaining resonance assigned to the four-metal cluster in the intact protein can be explained to result from the removal of the NH2-terminal polypeptide fragment containing the three-metal cluster. These results provide unambiguous evidence for the two-domain structure of metallothionein, containing a separate three- and a four-metal cluster.

Animals↗
PubMed1982

Domain nature of metallothionein.

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.

Amino Acid Sequence↗