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. 2001 Oct 15;359(Pt 2):353–360. doi: 10.1042/0264-6021:3590353

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

K Zangger 1, G Shen 1, G Oz 1, J D Otvos 1, I M Armitage 1
PMCID: PMC1222153  PMID: 11583581

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 (113/111)Cd- and (1)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 vivo 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 alpha-domain, and a detailed analysis of chemical shift changes and intermolecular nuclear Overhauser effects points towards a disulphide bond involving Cys(36). In contrast to the metal-bridged (non-oxidative) dimerization, the metal-cysteine cluster structures in both MT domains remain intact and no conformational exchange or metal-metal exchange was observed. Also in contrast to the many recently reported oxidative processes which involve the beta-domain cysteine groups and result 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-domain metals following this oxidation. Therefore these findings provide additional corroboration that metal binding in the C-terminal alpha-domain is rather tight, even under conditions of a changing cellular oxidation potential, compared with the more labile/dynamic nature of the metals in the N-terminal beta-domain cluster under similar conditions.

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Selected References

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  1. Arseniev A., Schultze P., Wörgötter E., Braun W., Wagner G., Vasák M., Kägi J. H., Wüthrich K. Three-dimensional structure of rabbit liver [Cd7]metallothionein-2a in aqueous solution determined by nuclear magnetic resonance. J Mol Biol. 1988 Jun 5;201(3):637–657. doi: 10.1016/0022-2836(88)90644-4. [DOI] [PubMed] [Google Scholar]
  2. Cherian M. G., Howell S. B., Imura N., Klaassen C. D., Koropatnick J., Lazo J. S., Waalkes M. P. Role of metallothionein in carcinogenesis. Toxicol Appl Pharmacol. 1994 May;126(1):1–5. doi: 10.1006/taap.1994.1083. [DOI] [PubMed] [Google Scholar]
  3. Cuajungco M. P., Lees G. J. Zinc metabolism in the brain: relevance to human neurodegenerative disorders. Neurobiol Dis. 1997;4(3-4):137–169. doi: 10.1006/nbdi.1997.0163. [DOI] [PubMed] [Google Scholar]
  4. Furey W. F., Robbins A. H., Clancy L. L., Winge D. R., Wang B. C., Stout C. D. Crystal structure of Cd,Zn metallothionein. Experientia Suppl. 1987;52:139–148. doi: 10.1007/978-3-0348-6784-9_7. [DOI] [PubMed] [Google Scholar]
  5. Hamer D. H. Metallothionein. Annu Rev Biochem. 1986;55:913–951. doi: 10.1146/annurev.bi.55.070186.004405. [DOI] [PubMed] [Google Scholar]
  6. Jacob C., Maret W., Vallee B. L. Control of zinc transfer between thionein, metallothionein, and zinc proteins. Proc Natl Acad Sci U S A. 1998 Mar 31;95(7):3489–3494. doi: 10.1073/pnas.95.7.3489. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Jiang L. J., Maret W., Vallee B. L. The glutathione redox couple modulates zinc transfer from metallothionein to zinc-depleted sorbitol dehydrogenase. Proc Natl Acad Sci U S A. 1998 Mar 31;95(7):3483–3488. doi: 10.1073/pnas.95.7.3483. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Kröncke K. D., Fehsel K., Schmidt T., Zenke F. T., Dasting I., Wesener J. R., Bettermann H., Breunig K. D., Kolb-Bachofen V. Nitric oxide destroys zinc-sulfur clusters inducing zinc release from metallothionein and inhibition of the zinc finger-type yeast transcription activator LAC9. Biochem Biophys Res Commun. 1994 Apr 29;200(2):1105–1110. doi: 10.1006/bbrc.1994.1564. [DOI] [PubMed] [Google Scholar]
  9. Kägi J. H., Kojima Y. Chemistry and biochemistry of metallothionein. Experientia Suppl. 1987;52:25–61. doi: 10.1007/978-3-0348-6784-9_3. [DOI] [PubMed] [Google Scholar]
  10. Maret W., Jacob C., Vallee B. L., Fischer E. H. Inhibitory sites in enzymes: zinc removal and reactivation by thionein. Proc Natl Acad Sci U S A. 1999 Mar 2;96(5):1936–1940. doi: 10.1073/pnas.96.5.1936. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Maret W., Larsen K. S., Vallee B. L. Coordination dynamics of biological zinc "clusters" in metallothioneins and in the DNA-binding domain of the transcription factor Gal4. Proc Natl Acad Sci U S A. 1997 Mar 18;94(6):2233–2237. doi: 10.1073/pnas.94.6.2233. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Maret W., Vallee B. L. Thiolate ligands in metallothionein confer redox activity on zinc clusters. Proc Natl Acad Sci U S A. 1998 Mar 31;95(7):3478–3482. doi: 10.1073/pnas.95.7.3478. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Nettesheim D. G., Engeseth H. R., Otvos J. D. Products of metal exchange reactions of metallothionein. Biochemistry. 1985 Nov 19;24(24):6744–6751. doi: 10.1021/bi00345a003. [DOI] [PubMed] [Google Scholar]
  14. O'Brien P., Salacinski H. J. Evidence that the reactions of cadmium in the presence of metallothionein can produce hydroxyl radicals. Arch Toxicol. 1998 Nov;72(11):690–700. doi: 10.1007/s002040050562. [DOI] [PubMed] [Google Scholar]
  15. Otvos J. D., Engeseth H. R., Nettesheim D. G., Hilt C. R. Interprotein metal exchange reactions of metallothionein. Experientia Suppl. 1987;52:171–178. doi: 10.1007/978-3-0348-6784-9_10. [DOI] [PubMed] [Google Scholar]
  16. Otvos J. D., Engeseth H. R., Wehrli S. Preparation and 113Cd NMR studies of homogeneous reconstituted metallothionein: reaffirmation of the two-cluster arrangement of metals. Biochemistry. 1985 Nov 19;24(24):6735–6740. doi: 10.1021/bi00345a001. [DOI] [PubMed] [Google Scholar]
  17. Oz G., Pountney D. L., Armitage I. M. NMR spectroscopic studies of I = 1/2 metal ions in biological systems. Biochem Cell Biol. 1998;76(2-3):223–234. doi: 10.1139/bcb-76-2-3-223. [DOI] [PubMed] [Google Scholar]
  18. Palmiter R. D. The elusive function of metallothioneins. Proc Natl Acad Sci U S A. 1998 Jul 21;95(15):8428–8430. doi: 10.1073/pnas.95.15.8428. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Palumaa P., Mackay E. A., Vasák M. Nonoxidative cadmium-dependent dimerization of Cd7-metallothionein from rabbit liver. Biochemistry. 1992 Feb 25;31(7):2181–2186. doi: 10.1021/bi00122a040. [DOI] [PubMed] [Google Scholar]
  20. Palumaa P., Vaher M. Metal-induced dimerization of Cd7-metallothionein. Role of anions. Ann Clin Lab Sci. 1996 May-Jun;26(3):264–268. [PubMed] [Google Scholar]
  21. Palumaa P., Vasák M. Binding of inorganic phosphate to the cadmium-induced dimeric form of metallothionein from rabbit liver. Eur J Biochem. 1992 May 1;205(3):1131–1135. doi: 10.1111/j.1432-1033.1992.tb16882.x. [DOI] [PubMed] [Google Scholar]
  22. Palumaa P., Zerbe O., Vasák M. Formation and spectroscopic characterization of a novel monomeric cadmium- and phosphate-containing form of metallothionein. Biochemistry. 1993 Mar 23;32(11):2874–2879. doi: 10.1021/bi00062a019. [DOI] [PubMed] [Google Scholar]
  23. Robbins A. H., McRee D. E., Williamson M., Collett S. A., Xuong N. H., Furey W. F., Wang B. C., Stout C. D. Refined crystal structure of Cd, Zn metallothionein at 2.0 A resolution. J Mol Biol. 1991 Oct 20;221(4):1269–1293. [PubMed] [Google Scholar]
  24. Sato M., Bremner I. Oxygen free radicals and metallothionein. Free Radic Biol Med. 1993 Mar;14(3):325–337. doi: 10.1016/0891-5849(93)90029-t. [DOI] [PubMed] [Google Scholar]
  25. Schultze P., Wörgötter E., Braun W., Wagner G., Vasák M., Kägi J. H., Wüthrich K. Conformation of [Cd7]-metallothionein-2 from rat liver in aqueous solution determined by nuclear magnetic resonance spectroscopy. J Mol Biol. 1988 Sep 5;203(1):251–268. doi: 10.1016/0022-2836(88)90106-4. [DOI] [PubMed] [Google Scholar]
  26. Stohs S. J., Bagchi D. Oxidative mechanisms in the toxicity of metal ions. Free Radic Biol Med. 1995 Feb;18(2):321–336. doi: 10.1016/0891-5849(94)00159-h. [DOI] [PubMed] [Google Scholar]
  27. Suzuki K. T., Ohnuki R., Yaguchi K. Post-mortem and in vitro dimerization of metallothionein in cadmium-accumulated rat liver and kidney. Toxicol Lett. 1983 Apr;16(1-2):77–84. doi: 10.1016/0378-4274(83)90013-9. [DOI] [PubMed] [Google Scholar]
  28. Thornalley P. J., Vasák M. Possible role for metallothionein in protection against radiation-induced oxidative stress. Kinetics and mechanism of its reaction with superoxide and hydroxyl radicals. Biochim Biophys Acta. 1985 Jan 21;827(1):36–44. doi: 10.1016/0167-4838(85)90098-6. [DOI] [PubMed] [Google Scholar]
  29. Thévenod F., Friedmann J. M. Cadmium-mediated oxidative stress in kidney proximal tubule cells induces degradation of Na+/K(+)-ATPase through proteasomal and endo-/lysosomal proteolytic pathways. FASEB J. 1999 Oct;13(13):1751–1761. doi: 10.1096/fasebj.13.13.1751. [DOI] [PubMed] [Google Scholar]
  30. Wishart D. S., Bigam C. G., Yao J., Abildgaard F., Dyson H. J., Oldfield E., Markley J. L., Sykes B. D. 1H, 13C and 15N chemical shift referencing in biomolecular NMR. J Biomol NMR. 1995 Sep;6(2):135–140. doi: 10.1007/BF00211777. [DOI] [PubMed] [Google Scholar]
  31. Zangger K., Oz G., Haslinger E., Kunert O., Armitage I. M. Nitric oxide selectively releases metals from the amino-terminal domain of metallothioneins: potential role at inflammatory sites. FASEB J. 2001 May;15(7):1303–1305. doi: 10.1096/fj.00-0641fje. [DOI] [PubMed] [Google Scholar]
  32. Zangger K., Oz G., Otvos J. D., Armitage I. M. Three-dimensional solution structure of mouse [Cd7]-metallothionein-1 by homonuclear and heteronuclear NMR spectroscopy. Protein Sci. 1999 Dec;8(12):2630–2638. doi: 10.1110/ps.8.12.2630. [DOI] [PMC free article] [PubMed] [Google Scholar]

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