Abstract
Metallothioneins are small cysteine-rich proteins believed to play a role, among others, in the homeostasis of essential metal ions such as ZnII and CuI. Recently, we could show that wheat Ec-1 is coordinating its six ZnII ions in form of metal–thiolate clusters analogously to the vertebrate metallothioneins. Specifically, two ZnII ions are bound in the N-terminal and four in the C-terminal domain. In the following, we will present evidence for the relative independence of the two domains from each other with respect to their metal ion binding abilities, and uncover three intriguing peculiarities of the protein. Firstly, one ZnII ion of the N-terminal domain is relative resistant to complete replacement with CdII indicating the presence of a ZnII-binding site with increased stability. Secondly, the C-terminal domain is able to coordinate an additional fifth metal ion, though with reduced affinity, which went undetected so far. Finally, reconstitution of apoEc-1 with an excess of ZnII shows a certain amount of sub-stoichiometrically metal-loaded species. The possible relevance of these finding for the proposed biological functions of wheat Ec-1 will be discussed. In addition, extended X-ray absorption fine structure (EXAFS) measurements on both, the full-length and the truncated protein, provide final evidence for His participation in metal ion binding.
Peroza, E A