Oxidative dimerization in metallothionein is a result of intermolecular disulphide bonds between cysteines in the alpha-domain

Upon storage under aerobic conditions metallothioneins (MTs) form a new spe cies, which is characterized by a molecular mass approximately twice the si ze of monomeric MT and shifted Cd-113/111- and H-1-NMR resonances. The inve stigation of this oxidative dimerization process by NMR spectroscopy allowe d us to structurally characterize this NIT species that has been described to occur in vivo and might be synthesized under conditions of oxidative str ess. The oxidative dimer was characterized by the formation of an intermole cular cysteine disulphide bond involving the a-domain, and a detailed analy sis of chemical shift changes and intermolecular nuclear Overhauser effects points towards a disulphide bond involving Cys(36). In contrast to the met al-bridged (non-oxidative) dimerization, the metal-cysteine cluster structu res in both MT domains remain intact and no conformational exchange or meta l-metal exchange was observed. Also in contrast to the many recently report ed oxidative processes which involve the beta -domain cysteine groups and r esult in the increased dynamics of the bound metal ions in this N-terminal domain, we found no evidence for any increased dynamics in the alpha -domai n metals following this oxidation. Therefore these findings provide additio nal corroboration that metal binding in the C-terminal alpha -domain is rat her tight, even under conditions of a changing cellular oxidation potential , compared with the more labile/dynamic nature of the metals in the N-termi nal beta -domain cluster under similar conditions.