Comparative Fe and Zn K-edge X-ray absorption spectroscopic study of the ferroxidase centres of human H-chain ferritin and bacterioferritin from Desulfovibrio desulfuricans.

Iron uptake by the ubiquitous iron-storage
protein ferritin involves the oxidation of two Fe(II) ions
located at the highly conserved dinuclear ‘‘ferroxidase
centre’’ in individual subunits. We have measured X-ray
absorption spectra of four mutants (K86Q, K86Q/E27D,
K86Q/E107D, and K86Q/E27D/E107D, involving variations
of Glu to Asp on either or both sides of the dinuclear
ferroxidase site) of recombinant human H-chain ferritin
(rHuHF) in their complexes with reactive Fe(II) and redoxinactive
Zn(II). The results for Fe–rHuHf are compared
with those for recombinant Desulfovibrio desulfuricans
bacterioferritin (DdBfr) in three states: oxidised, reduced,
and oxidised/Chelex -treated. The X-ray absorption nearedge
region of the spectrum allows the oxidation state of
the iron ions to be assessed. Extended X-ray absorption fine
structure simulations have yielded accurate geometric
information that represents an important refinement of the
crystal structure of DdBfr; most metal–ligand bonds are
shortened and there is a decrease in ionic radius going from
the Fe(II) to the Fe(III) state. The Chelex -treated sample
is found to be partly mineralised, giving an indication of
the state of iron in the cycled-oxidised (reduced, then oxidised) form of DdBfr, where the crystal structure shows
the dinuclear site to be only half occupied. In the case of
rHuHF the complexes with Zn(II) reveal a surprising
similarity between the variants, indicating that the rHuHf
dinuclear site is rigid. In spite of this, the rHuHf complexes
with Fe(II) show a variation in reactivity that is reflected in
the iron oxidation states and coordination geometries.