A de novo evolved domain improves the cadmium detoxification capacity of limpet metallothioneins.

Metallothioneins (MTs) constitute an important family of metal binding proteins. Mollusk MTs, in particular, have been used as model systems to better understand the evolution of their metal binding features and functional adaptation. In the present study two recombinantly produced MTs, LgiMT1 and LgiMT2, and their de novo evolved γ domain, of the marine limpet Lottia gigantea, were analyzed by electronic spectroscopy and mass spectrometry. Both MT proteins, as well as their γ domains, exhibit a strong binding specificity for Cd(II), but not for Zn(II) or Cu(I). The LgiMTs' γ domain renders an M(SCys) cluster with an increased Cd stoichiometry (binding 4 instead of 3 Cd ions), representing a novel structural element in the world of MTs, probably featuring an adamantane 3D structure. This cluster significantly improves the Cd(II)-binding performance of the full length proteins and thus contributes to the particularly high Cd coping capacity observed in free-living limpets.