title: Defining metal ion inhibitor interactions with recombinant human H- and L-chain ferritins and site-directed variants: an isothermal titration calorimetry study.
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, Delta H degrees, Delta S degrees, and Delta G degrees ) 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 Tb(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.
authors.affiliation: Department of Chemistry, University of New Hampshire, Durham, NH 03824, USA.
authors.forename: Fadi
authors.initials: F
authors.lastname: Bou-Abdallah
authors.forename: Paolo
authors.initials: P
authors.lastname: Arosio
authors.forename: Sonia
authors.initials: S
authors.lastname: Levi
authors.forename: Christine
authors.initials: C
authors.lastname: Janus-Chandler
authors.forename: N Dennis
authors.initials: ND
authors.lastname: Chasteen
mesh_terms: D001052:Apoferritins; D002151:Calorimetry / Q000379:methods; D005293:Ferritins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D008670:Metals / Q000737:chemistry / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism*
chemical_list: D008670:Metals; D011994:Recombinant Proteins; D005293:Ferritins; D001052:Apoferritins