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Biochemical and spectroscopic studies of human melanotransferrin (MTf): electron-paramagnetic resonance evidence for a difference between the iron-binding site of MTf and other transferrins

  • Sebastien Farnaud
    ,
  • Maryam Amini
    ,
  • Chiara Rapisarda
    ,
  • Richard Cammack
    ,
  • Tam Bui
    ,
  • Alex F. Drake
Research Output: Contribution to journal Article Peer-review

Abstract

Melanotransferrin (MTf) is a member of the transferrin (Tf) family of iron (Fe)-binding proteins that was first identified as a cell-surface marker of melanoma. Although MTf has a high-affinity Fe-binding site that is practically identical to that of serum Tf, the protein does not play an essential role in Fe homeostasis and its precise molecular function remains unclear. A Zn(II)-binding motif, distinct from the Fe-binding site, has been proposed in human MTf based on computer modelling studies. However, little is known concerning the interaction of its proposed binding site(s) with metals and the consequences in terms of MTf conformation. For the first time, biochemical and spectroscopic techniques have been used in this study to characterise metal ion-binding to recombinant MTf. Initially, the binding of Fe to MTf was examined using 6 M urea gel electrophoresis. Although four different iron-loaded forms were observed with serum Tf, only two forms were found with MTf, the apo-form and the N-monoferric holo-protein, suggesting a single high-affinity site. The presence of a single Fe(III)-binding site was also supported by EPR results which indicated that the Fe(III)-binding characteristics of MTf were unique, but somewhat comparable to the N-lobes of human serum Tf and chicken ovo-Tf. Circular dichroism (CD) analysis indicated that, as for Tf, no changes in secondary structure could be observed upon Fe(III)-binding. The ability of MTf to bind Zn(II) was also investigated using CD which demonstrated that the single high-affinity Fe-binding site was distinct from a potential Zn(II)-binding site.

Publication Information

Output type

Research Output: Contribution to journal Article Peer-review

Original language

English

Pages from-to (Number of pages)

Pages 2739

Journal (Volume, Issue Number)

International Journal of Biochemistry and Cell Biology (Volume 40, Issue 12)

Publication milestones

  • Published - 01/01/2008

Publication status

Published - 01/01/2008

ISSN

1357-2725

External Publication IDs

  • handle.net: 10547/593439
  • Scopus: 51249103806

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