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JBIC Journal of Biological Inorganic Chemistry

, Volume 4, Issue 5, pp 621–631 | Cite as

Identification of platination sites on human serum transferrin using 13C and 15N NMR spectroscopy

  • Mark C. Cox
  • Kevin J. Barnham
  • Tom A. Frenkiel
  • James D. Hoeschele
  • Anne B. Mason
  • Qing-Yu He
  • Robert C. Woodworth
  • P. J. Sadler
ORIGINAL ARTICLE

Abstract

 Reactions between various apo and metal-bound forms of human serum transferrin (80 kDa) and the recombinant N-lobe (40 kDa) with [Pt(en)Cl2] or cis-[PtCl2(NH3)2] have been investigated in solution via observation of [1H,15N] NMR resonances of the Pt complexes, [1H,13C] resonances of the eCH3 groups of the protein methionine residues, and by chromatographic analysis of single-site methionine mutants. For the whole protein, the preferred Pt binding site appears to be Met256. Additional binding occurs at the other surface-exposed methionine (Met499), which is platinated at a slower rate than Met256. In contrast, binding of similar Pt compounds to the N-lobe of the protein occurs at Met313, rather than Met256. Met313 is buried in the interlobe contact region of intact transferrin. After loss of one chloride ligand from Pt and binding to methionine sulfur of the N-lobe, chelate-ring closure appears to occur with binding to a deprotonated backbone amide nitrogen, and the loss of the other chloride ligand. Such chelate-ring closure was not observed during reactions of the whole protein, even after several days.

Key words Transferrin Platinum Anticancer complexes NMR spectroscopy Methionine 

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Copyright information

© Society of Biological Inorganic Chemistry 1999

Authors and Affiliations

  • Mark C. Cox
    • 1
  • Kevin J. Barnham
    • 1
  • Tom A. Frenkiel
    • 2
  • James D. Hoeschele
    • 3
  • Anne B. Mason
    • 4
  • Qing-Yu He
    • 4
  • Robert C. Woodworth
    • 4
  • P. J. Sadler
    • 1
  1. 1.Department of Chemistry, University of Edinburgh West Mains Road, Edinburgh EH9 3JJ, UK e-mail: p.j.sadler@ed.ac.uk Tel.: +44-131-6504729 Fax: +44-131-6506452GB
  2. 2.Biomedical NMR Centre, National Institute for Medical Research, The Ridgeway, Mill Hill, London NW7 1AA, UKGB
  3. 3.Department of Chemistry, Michigan State University East Lansing, MI 48824, USAUS
  4. 4.Department of Biochemistry, University of Vermont College of Medicine, Burlington, VT 05405, USAUS

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