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

Abstract

Upon storage under aerobic conditions metallothioneins (MTs) form a new species, which is characterized by a molecular mass approximately twice the size of monomeric MT and shifted ""$/"""Cd-and "H-NMR resonances. The investigation of this oxidative dimerization process by NMR spectroscopy allowed us to structurally characterize this MT species that has been described to occur in i o and might be synthesized under conditions of oxidative stress. The oxidative dimer was characterized by the formation of an intermolecular cysteine disulphide bond involving the α-domain, and a detailed analysis of chemical shift changes and intermolecular nuclear Overhauser effects points towards a disulphide bond involving Cys$'. In contrast to the metal-bridged (non-oxidative) dimerization, the metalcysteine cluster structures in both MT domains remain intact and

Metallothionein Disulfides Are Present in Metallothionein-overexpressing Transgenic Mouse Heart and Increase under Conditions of Oxidative Stress

“…2) ␤-domain of MT has been proposed to be important for zinc and copper homeostasis (31). However, an intermolecular disulfide bond in the ␣-domain of MT has been demonstrated in vitro under oxidative conditions (19). Our results suggest that both the ␣-and ␤-domains in MT may be involved in disulfide bond formation under both physiological and oxidative stress conditions.…”

“…A number of in vitro studies have shown that MT can form dimers through disulfide bond formation under oxidative conditions (17,19,24), although intramolecular oxidation of the cysteine thiolates and mixed disulfide formation has also been demonstrated (28). Intramolecular disulfide bonds following treatment of MT with S-nitrosocysteine, which generates nitric oxide, were also detected by Raman spectroscopy (16).…”

“…MT nitrosylation leads to disulfide bond formation in the ␤-domain (11). Upon storage under aerobic conditions, MT becomes dimerized through a disulfide bridge between two cysteine residues in thr C-terminal ␣-domain (19). Although the Cu 4 cluster in the ␤-domain of Cu 4 Zn 4 -MT-3 was stable to air oxidation, the Zn 4 cluster in the ␣-domain was found to be air sensitive resulting in the release of one zinc ion, presumably accompanied by the formation of disulfide bonds.…”

Role of metallothionein in cadmium traffic and toxicity in kidneys and other mammalian organs

“…Due to numerous cysteines with thiol (SH) groups, MTs exhibit high-affinity/highcapacity binding properties for various reactive metal ions, such as Zn, Cd, Hg, Cu, Pb, Ni, Co, Fe, Ag, and Au; one molecule of apo-MT can bind 7-9 Zn or Cd ions (or any combination of these two), up to 12 Cu ions, and up to 18 Hg ions (Palumaa et al 2002(Palumaa et al , 2003reviewed in Stillman 1995). The reactive thiol groups promote spontaneous oligomerization of the thionein molecules, and such aggregates from the rabbit liver and horse kidney can bind much more than usual 7 g atom Cd/mol apo-MT (Zangger et al 2001;Wilhelmsen et al 2002). The binding affinity of MTs for metal ions is metal-dependent, but several in vitro studies, using rat liver common MTs, showed a slightly different relative order of affinities for metals; in the study by Waalkes et al (1984), the order was Cd [ Pb [ Cu [ Hg [ Zn [ Ag [ Ni [ Co, in the study by Nielson et al (1985), the order was Hg [ Cu [ Cd [ Zn [ Ni = Co, whereas Hamer (1986) reported the order:…”

Partial oxidation and oxidative polymerization of metallothionein

“…Disulfide-containing MT has been isolated from tissues of mice that overexpress the protein [19]. Isolated MT forms intermolecular or intramolecular disulfide bridges [20,21]. Thus, MT exists in states that differ in metal load and degree of oxidation.…”