Skip to main content
SpringerLink
Log in

Effect of α-domain substitution on the structure, property and function of human neuronal growth inhibitory factor

  • Original Paper
  • Published:
JBIC Journal of Biological Inorganic Chemistry Aims and scope Submit manuscript

Abstract

Human metallothionein-3 (hMT3), also named human neuronal growth inhibitory factor (hGIF), is attractive due to its distinct neuronal growth inhibitory activity, which is not shown by other human MT isoforms. It has been reported that the neuronal growth inhibitory activity arises from the N-terminal β-domain rather than its C-terminal α-domain. However, previous bioassay results have shown that the single β-domain is less effective at inhibiting the neuron growth than that in intact hMT3 on a molar basis, which suggests that the α-domain is indispensable to the neuronal growth inhibitory activity of hMT3. In order to confirm this assumption, we constructed two domain-hybrid mutants, the β(MT3)–β(MT3) mutant and the β(MT3)–α(MT1) mutant, and investigated their structural and metal binding properties by UV-vis spectroscopy, CD spectroscopy, pH titration, DTNB reaction, EDTA reaction, etc. The results showed that stability of the Cd3S9 cluster of the β(MT3)–β(MT3) mutant decreased significantly while the Cd3S9 cluster of the β(MT3)–α(MT1) mutant had a similar stability and solvent accessibility to that of hMT3. Interestingly, the bioassay results showed that the neuronal growth inhibitory activity of the β(MT3)–β(MT3) mutant decreased significantly, while the β(MT3)–α(MT1) mutant showed similar inhibitory activity to hMT3. Based on these results, we conclude that the α-domain is indispensable and plays an important role in modulating the stability of the metal cluster in the β-domain by domain–domain interactions, thus influencing the bioactivity of hMT3.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4

Similar content being viewed by others

References

  1. Kagi JHR, Schaffer A (1988) Biochemistry 27:8509–8511

    Article  PubMed  CAS  Google Scholar 

  2. Kagi JHR, Kojima Y (1987) Experientia 52:25–61

    CAS  Google Scholar 

  3. Margoshes M, Vallee BL (1957) J Am Chem Soc 79:4813–4814

    Article  CAS  Google Scholar 

  4. Uchida Y, Tomonaga M (1989) Brain Res 481:190–193

    Article  PubMed  CAS  Google Scholar 

  5. Uchida Y, Takio K, Titani K, Ihara Y, Tomonaga M (1991) Neuron 7:337–347

    Article  PubMed  CAS  Google Scholar 

  6. Palmiter RD, Findley SD, Whitemore TE, Durnam DM (1992) Proc Natl Acad Sci USA 89:6333–6337

    Article  PubMed  CAS  Google Scholar 

  7. Irie Y, Keung WM (2001) Biochem Biophys Res Commun 282:416–420

    Article  PubMed  CAS  Google Scholar 

  8. Irie Y, Keung WM (2003) Brain Res 960:228–234

    Article  PubMed  CAS  Google Scholar 

  9. Hasler DW, Faller P, Vasak M (1998) Biochemistry 37:14966–14973

    Article  PubMed  CAS  Google Scholar 

  10. Faller P, Vasak M (1997) Biochemistry 36:13341–13348

    Article  PubMed  CAS  Google Scholar 

  11. Sewell AK, Jensen LT, Erickson JC, Palmiter RD, Winge DR (1995) Biochemistry 34:4740–4747

    Article  PubMed  CAS  Google Scholar 

  12. Uchida Y, Ihara Y (1995) J Biol Chem 270:3365–3369

    Article  PubMed  CAS  Google Scholar 

  13. Romero-Isart N, Jensen LT, Zerbe O, Winge DR, Vasak M (2002) J Biol Chem 277:37023–37028

    Article  PubMed  CAS  Google Scholar 

  14. Hasler DW, Jensen LT, Zerbe O, Winge DR, Vasak M (2000) Biochemistry 39:14567–14575

    Article  PubMed  CAS  Google Scholar 

  15. Ni FY, Cai B, Ding ZC, Zheng F, Zhang MJ, Wu HM, Sun HZ, Huang ZX (2007) Proteins: Struct Funct Bioinf 68:255–266

    Google Scholar 

  16. Williamson MP (1994) Biochem J 297:249–260

    Google Scholar 

  17. Cai B, Zheng Q, Teng XC, Chen D, Wang Y, Wang KQ, Zhou GM, Xie Y, Zhang MJ, Sun HZ, Huang ZX (2006) J Biol Inorg Chem 11:476–482

    Article  PubMed  CAS  Google Scholar 

  18. Ding ZC, Teng XC, Cai B, Wang H, Zheng Q, Wang Y, Zhou GM, Zhang MJ, Wu HM, Sun HZ, Huang ZX (2006) Biochem Biophys Res Commun 349:674–682

    Article  PubMed  CAS  Google Scholar 

  19. Higucchi R, Krummel B, Saiki RK (1988) Nucleic Acids Res 16:7351–7367

    Google Scholar 

  20. Vasak M (1991) Methods Enzymol 205:452–458

    PubMed  CAS  Google Scholar 

  21. Grassetti DR, Murray JF (1967) Arch Biochem Biophys 119:41–49

    Article  PubMed  CAS  Google Scholar 

  22. Winge DR, Miklossy KA (1982) J Biol Chem 257:3471–3476

    PubMed  CAS  Google Scholar 

  23. Vasak M, Kagi JHR (1983) In: Sigel H (ed) Metal ions in biological systems. Marcel Dekker Inc., New York, pp 213–273

    Google Scholar 

  24. Kagi JHR, Vallee B (1961) J Biol Chem 236:2435–2442

    PubMed  CAS  Google Scholar 

  25. Wang Y, Mackay EA, Kurasaki M, Kagi JHR (1994) Eur J Biochem 225:449–457

    Article  PubMed  CAS  Google Scholar 

  26. Li TY, Kraker AJ, Show CF III, Petering DH (1980) Proc Natl Acad Sci USA 77:6334–6338

    Google Scholar 

  27. Shaw CF III, Savas MM, Petering DH (1991) Methods Enzymol 205:401–414

    Article  PubMed  CAS  Google Scholar 

  28. Uchida Y, Ihara Y, Tomonaga M (1988) Biochem Biophys Res Commun 150:1263–1267

    Article  PubMed  CAS  Google Scholar 

  29. Chung RS, Vickers JC, Chuan MI, Eckhardt BL, West AK (2002) Exp Neurol 178:1–12

    Article  PubMed  CAS  Google Scholar 

  30. Erickson JC, Sewell AK, Jensen LT, Winge DR, Palmiter RD (1994) Brain Res 649:297–304

    Article  PubMed  CAS  Google Scholar 

  31. Xiong Y, Ru BG (1997) J Biochem 121:1102–1106

    PubMed  CAS  Google Scholar 

  32. Vasak M (1998) Biodegradation 9:501–512

    Article  PubMed  CAS  Google Scholar 

  33. Oz G, Pountney DL, Armitage IM (1998) Biochem Cell Biol 76:223–234

    Article  PubMed  CAS  Google Scholar 

  34. Vasak M, Kagi JHR, Hill HAO (1981) Biochemistry 20:2852–2856

    Article  PubMed  CAS  Google Scholar 

  35. Stillman MJ, Cai WH, Zelazowski AJ (1987) J Biol Chem 262:4538–4558

    PubMed  CAS  Google Scholar 

  36. Zheng Q, Yang WM, Yu WH, Cai B, Teng XC, Xie Y, Sun HZ, Zhang MJ, Huang ZX (2003) Protein Eng 16:865–870

    Article  PubMed  CAS  Google Scholar 

  37. Cismowski MJ, Huang PC (1991) Biochemistry 30:6626–6632

    Article  PubMed  CAS  Google Scholar 

  38. Chang CC, Liao WF, Huang PC (1998) Protein Eng 11:41–46

    Article  PubMed  CAS  Google Scholar 

  39. Jiang LJ, Vasak M, Vallee BL, Maret W (2000) Proc Natl Acad Sci USA 97:2503–2508

    Article  PubMed  CAS  Google Scholar 

  40. Faller P, Hasler DW, Zerbe O, Klauser S, Winge DR, Vasak M (1999) Biochemistry 38:10158–10167

    Article  PubMed  CAS  Google Scholar 

  41. Erickson JC, Hollopeter G, Thomas SA, Froelick GJ, Palmiter RD (1997) J Neurosci 17:1271–1281

    PubMed  CAS  Google Scholar 

  42. Palmiter RD (1995) Toxicol Appl Pharmacol 135:139–146

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

This project was supported by the National Science Foundation of China.

Author information

Authors and Affiliations

  1. Chemical Biology Lab, Department of Chemistry, Fudan University, Shanghai, 200433, China

    Zhi-Chun Ding, Qi Zheng, Bin Cai, Wen-Hao Yu, Xin-Chen Teng & Zhong-Xian Huang

  2. Department of Anatomy, Histology and Embryology, Medical School, Fudan University, Shanghai, 200032, China

    Yang Wang & Guo-Ming Zhou

  3. State Key Lab of Bio-organic and Natural Products, Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, Shanghai, 200032, China

    Hou-Ming Wu

  4. Department of Chemistry, The University of Hong Kong, Pokfulam Road, Hong Kong, China

    Hong-Zhe Sun

  5. Department of Biochemistry, Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong, China

    Ming-Jie Zhang

Authors
  1. Zhi-Chun Ding
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Qi Zheng
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Bin Cai
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Wen-Hao Yu
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Xin-Chen Teng
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Yang Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Guo-Ming Zhou
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Hou-Ming Wu
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Hong-Zhe Sun
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Ming-Jie Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Zhong-Xian Huang
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Zhong-Xian Huang.

Additional information

Z.-C. Ding and Q. Zheng contributed equally to this work.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Ding, ZC., Zheng, Q., Cai, B. et al. Effect of α-domain substitution on the structure, property and function of human neuronal growth inhibitory factor. J Biol Inorg Chem 12, 1173–1179 (2007). https://doi.org/10.1007/s00775-007-0287-x

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00775-007-0287-x

Keywords

Search

Navigation