111Cd NMR Studies of the Domain Specificity of Ag+ and Cu+ Binding to Metallothionein

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Abstract
Metal displacement reactions of Cd7MT with Ag+ or Cu+ and interprotein metal exchange reactions between Cd7MT and Ag12MT or Cu12MT were studied by 111Cd NMR. Titration of 111Cd7MT with Ag+ indicates that Ag+ binds preferentially to the β-domain of the protein to form the metal hybrid species, (Cd4)α(Ag6)βMT. Once the β-domain is filled, additional Ag+ ions displace Cd2+ from the α-domain to form (Ag6)α(Ag6)βMT. The metal displacement reaction is cooperative and the two domains react independently of one another. The (Cd4)α(Ag6)βMT hybrid protein is also formed as the major product of direct interprotein metal exchange between Cd7MT and Ag12MT. Cu+ reacts with Cd7MT in a manner similar to Ag+, with addition of 6 equiv of Cu+ leading to preferential formation of (Cd4)α(Cu6)βMT, and 12 equiv of Cu+ to formation of (Cu6)α(Cu6)βMT. However, unlike Ag+, Cu+ appears to produce intermediate species that may contain mixed-metal clusters. Interprotein metal exchange between Cu12MT and Cd7MT leads to the net transfer of Cd2+ into the α-domain and Cu+ into the β-domain. The differential affinities of the two domains for monovalent and divalent metal ions plus the availability of facile pathways for metal exchange may be features that enable MT to function simultaneously in the metabolism of different metal ions.