Significance of alpha-fragment of metallothionein in cadmium binding

In order to evaluate the significance of alpha- and beta-fragments of metallothionein with regard to Cd binding in biosynthetic processes, the Cd-binding ability of four mutant metallothioneins was examined using the Escherichia coli expression system. The features of the mutant metallothioneins were proteins in which cysteine residues in the alpha- or beta-fragment were replaced with alanine residues, or that the sequential order of the fragments was altered. The expressed mutant metallothioneins having an intact alpha-fragment showed the constructive abilities of the Cd-thiolate cluster. On the other hand, mutant metallothionein having an alpha-fragment lacking metal-binding sites exhibited no Cd-binding ability. The condition for maintaining the Cd-binding capability of metallothionein was that the alpha-fragment remains intact irrespective of the sequential order of the two fragments. The alpha-fragment is an indispensable component in metal-binding processes of Cd-metallothionein.