Antigenicity of metallothionein.

The antigenic determinants of vertebrate metallothionein have been determined by a competitive-binding double-antibody radioimmunoassay to consist of two immunologically dominant regions in the NH2-terminal domain (residues 1-29). The COOH-terminal domain (residues 30-61) exhibited trivial immunoreactivity in competitive binding assays. The tryptic peptide encompassing residues 1-25 of the molecule competed with 125I-labeled metallothionein as effectively as the native protein. The crossreactivity was unaffected whether the protein was native or denatured. The antigenicity is thus independent of the degree of folding of the protein and, although all antigenic sites depend to some degree on conformation or topography, this favors a sequential (or continuous) rather than a discontinuous nature of the determinants. The two regions in metallothionein that appear to be important in the interaction of the molecule with the antisera include the NH2-terminal acetylated methionine and the cluster of lysines in the sequence from residues 20-25. These regions are homologous in the various vertebrate metallothioneins known to crossreact with rabbit anti-rat metallothionein antiserum. This implies that the induction in rabbits of antiserum to rat metallothionein is an autoimmune phenomenon. The results support the two-cluster model of metallothionein and are compatible with predictions of sites of antigenicity based on regions of high hydrophilicity.