Metallothionein (MT) is a suitable model for investigating molecular intera
ctions relating to the handling of metals in cells. However, the production
of functional MT proteins in microorganisms has been limited because of th
e instability of MT-the thiol group of cysteine is easily oxidized and prot
eolysis occurs. To increase the binding ability and to stabilize MT, we des
igned genes for dimeric and tetrameric MT and the genes were successfully o
verexpressed in Escherichia coli to generate functional oligomeric MTs. A h
uman MT-H (hMT-II) synthesized with prokaryotic codons, a linker encoding a
glycine tripeptide, and Met-deficient hMT-II was ligated to create a dimer
ic MT, from which a tetrameric MT was then constructed. The increased molec
ular size of the constructs resulted in improved stability and productivity
in E. coli. Cells of E. coli carrying the oligomeric MT genes showed resis
tance toward Zn and Cd toxicity. The oligomeric proteins formed inclusion b
odies, which were dissolved with dithiothreitol, and the purified apo-MTs w
ere reconstituted with Cd or Zn ions under reducing conditions. The dimeric
and tetrameric MT proteins exhibited both Cd and Zn binding activities tha
t were, respectively, two and four times higher than those of the hMT-II mo
nomer protein. These stable oligomeric MTs have potential as a biomaterial
for uses such as detoxification and bioremediation for heavy metals.