Reaction of human metallothionein-3 with cisplatin and transplatin

Human metallothioneins, small cysteine- and metal-rich proteins, play an important role in the acquired resistance to platinum-based anticancer drugs. These proteins contain a M(II)4(CysS)11 cluster and a M(II)3(CysS)9 cluster localized in the alpha-domain and the beta-domain, respectively. The noninducible isoform metallothionein-3 (Zn7MT-3) is mainly expressed in the brain, but was found overexpressed in a number of cancer tissues. Since the structural properties of this isoform substantially differ from those of the ubiquitously occurring Zn7MT-1/Zn7MT-2 isoforms, the reactions of cis-diamminedichloridoplatinum(II) (cisplatin) and trans-diamminedichloridoplatinum(II) (transplatin) with human Zn7MT-3 were investigated and the products characterized. A comparison of the reaction kinetics revealed that transplatin reacts with cysteine ligands of Zn7MT-3 faster than cisplatin. In both binding processes, stoichiometric amounts of Zn(II) were released from the protein. Marked differences between the reaction rates of cisplatin and transplatin binding to Zn7MT-3 and the formation of the Pt-S bonds suggest that the binding of both Pt(II) compounds is a complex process, involving at least two subsequent binding steps. The electrospray ionization mass spectrometry characterization of the products showed that whereas all ligands in cisplatin were replaced by cysteine thiolates, transplatin retained its carrier ammine ligands. The 113Cd NMR studies of Pt1 113Cd6MT-3 revealed that cisplatin binds preferentially to the beta-domain of the protein. The rates of reaction of cisplatin and transplatin with Zn7MT-3 were much faster than those of cisplatin and transplatin with Zn7MT-2. The biological consequences of a substantially higher reactivity of cisplatin toward Zn7MT-3 than Zn7MT-2 in the acquired resistance to platinum-based drugs are discussed.