Constant features of iron mineralization and release

Iron minerals are nucleated on the inner surface of ferritin, at clusters of conserved carboxylate residues [45]. Substitution of Glu residues with Ala in L-type ferritin slows the rate of mineralization and decreases the pattern of bound water, which is highly conserved [46]. The nucleation sites are constant in all known ferritin sequences, which contrasts with the presence of the ferroxidase site only in H-type ferritins [1, 2, 47]. 

The iron mineral is stable inside ferritin until the addition of reductant and/or chelators [48]. The rate at which the iron is released by reductive chelation (FMN/ NADH -I- bipyridyl) is relatively constant for recombinant H- and L-type ferritins [49] and for natural mixtures of subunits from different tissues, except when post-translationally modified [50, 51]. The site for iron release is unknown, but rates of release are similar in recombinant ferritins composed of all the ferritin subunit types currently known. 

A mutation that substituted proline for a highly conserved ligand that does not bind iron, the leucine residue at position 134 which is not at the ferroxidase or nucleation sites, changed the rate of reductive iron release dramatically [49]. Initial rates of reductive iron release were increased four-fold in the pro/leu protein and were monophasic compared to wild-type proteins, which have a biphasic rate of iron release [52]. As a result, the time required to release all of the iron from the mineral (480 Fe/24-mer) was greatly decreased 5 minutes compared to 150 minutes for leu/pro compared to a biphasic release rate in wild-type protein (Table 12-1). 

Since leucine will not bind Fe directly, the effect of the pro/leu substitution must be on the higher order protein structure of ferritin. X-ray crystallography analysis of protein crystals showed that the pro/leu substitution disrupted the C/D helices near the turn. The effect of the single amino acid change was amplified by proximity to the junction of three-subunits [49]. The pro/leu protein assembled normally, but there were eight regions of localized disorder distributed around the molecule, at the 

Ferritin subunit type Initial rate (s ) % Fe released at 5 minutes 

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