Defining metal ion inhibitor interactions with recombinant human H- and L-chain ferritins and site-directed variants: An isothermal titration calorimetry study

Fadi Bou-Abdallah, Paolo Arosio, Sonia Levi, Christine Janus-Chandler, N. Dennis Chasteen

Research output: Contribution to journalArticlepeer-review

Abstract

Zinc and terbium, inhibitors of iron incorporation in the ferritins, have been used for many years as probes of structure-function relationships in these proteins. Isothermal titration calorimetric and kinetic measurements of Zn(II) and Tb(III) binding and inhibition of Fe(II) oxidation were used to identify and characterize thermodynamically (n, K, ΔH°, ΔS°, and ΔG°) the functionally important binding sites for these metal ions in recombinant human H-chain, L-chain, and H-chain site-directed variant ferritins. The data reveal at least two classes of binding sites for both Zn(II) and Tb(III) in human H-chain ferritin: one strong, corresponding to binding of one metal ion in each of the eight three-fold channels, and the other weak, involving binding at the ferroxidase and nucleation sites of the protein as well as at other weak unidentified binding sites. Zn(II) and Th(III) binding to recombinant L-chain ferritin showed similar stoichiometries for the strong binding sites within the channels, but fewer weaker binding sites when compared to the H-chain protein. The kinetics and binding data indicate that the binding of Zn(II) and Tb(III) in the three-fold channels, which is the main pathway of iron(II) entry in ferritin, blocks the access of most of the iron to the ferroxidase sites on the interior of the protein, accounting for the strong inhibition by these metal ions of the oxidative deposition of iron in ferritin.

Original languageEnglish
Pages (from-to)489-497
Number of pages9
JournalJournal of Biological Inorganic Chemistry
Volume8
Issue number4
Publication statusPublished - Apr 2003

Keywords

  • Ferritin
  • Iron oxidation
  • Isothermal titration calorimetry
  • Terbium binding
  • Zinc binding

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Biochemistry

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