Direct observation of the iron binding sites in a ferritin

Hempstead, Paul D.; Hudson, Aaron J.; Artymiuk, Peter J.; Andrews, Simon C.; Banfield, Mark J.; Guest, John R.; Harrison, Pauline M.

doi:10.1016/0014-5793(94)00781-0

Cited by 97 publications

(80 citation statements)

References 29 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Moreover, the subsequent increase of the fluorescence signal ( t l l Z -450 s) must be related to factors other than iron oxidation, namely to the migration of Fe(II1) from the ferroxidase center to the nucleation sites in the internal cavity, a process that cannot be singled out by optical absorption which monitors Fe(II1) formation. This interpretation is in good agreement with the model of iron incorporation proposed on the basis of other spectroscopic experiments (Bauminger et al, 1989;Treffry et al, 1996) and of crystallographic data (Hempstead et al, 1994). It is also consistent with the fluorescence changes observed upon stepwise addition of iron (Fig.…”

Section: Discussionsupporting

confidence: 92%

Formation and movement of Fe(III) in horse spleen, H‐ and L‐recombinant ferritins. A fluorescence study

et al. 1998

View full text Add to dashboard Cite

Iron oxidation and incorporation into apofemtins of different subunit composition, namely the recombinant H and L homopolymers and the natural horse spleen heteropolymer (10-15% H), have been followed by steady-state and time-resolved fluorescence. After aerobic addition of 100 Fe(I1) atoms/polymer, markedly different kinetic profiles are observed. In the rL-homopolymer a slow monotonic fluorescence quenching is observed which reflects binding, slow oxidation at the threefold apoferritin channels, and diffusion into the protein cavity. In the rH-homopolymer a fast fluorescence quenching is followed by a partial, slow recovery. The two processes have been attributed to Fe(I1) binding and oxidation at the ferroxidase centers and to Fe(II1) released into the cavity, respectively. The fluorescence kinetics of horse spleen apoferritin is dominated by the H chain contribution and resembles that of the H homopolymer. It brings out clearly that the rate of the overall process is limited by the rate at which Fe(II1) leaves the ferroxidase centers of the H chains where binding of incoming Fe(I1) and its oxidation take place. The data obtained upon stepwise addition of iron and the results of optical absorption measurements confirm this picture. The correspondence between steady-state and time-resolved data is remarkably good; this is manifest when the latter are used to calculate the change in fluorescence intensity as apparent in the steady-state measurements.

show abstract

Section: Discussionsupporting

confidence: 92%

Formation and movement of Fe(III) in horse spleen, H‐ and L‐recombinant ferritins. A fluorescence study

et al. 1998

View full text Add to dashboard Cite

show abstract

“…This distance could be too long for direct ligand exchange to take place, but taken into account the low resolution of the structure and the transient character of the complex, closer interactions between the two sites, which would include frataxin His-83, and ferrochelatase Asp-320 and possibly His-317 (previously shown to bind cadmium) are feasible in solution. Although in solution, Fe 2ϩ is normally coordinated by six water molecules, Fe 2ϩ at the di-iron ferroxidation site of frataxin, similarly to ferritin, may be coordinated to a histidine and acidic amino acid residues as well as to water (66,67). The water molecule could take part in ligand exchange by being initially replaced by one of the acidic residues of the -helix of ferrochelatase.…”

Section: Discussionmentioning

confidence: 99%

The Structure of the Complex between Yeast Frataxin and Ferrochelatase

Söderberg

Gillam²,

Ahlgren

et al. 2016

Journal of Biological Chemistry

View full text Add to dashboard Cite

Frataxin is a mitochondrial iron-binding protein involved in iron storage, detoxification, and delivery for iron sulfur-cluster assembly and heme biosynthesis. The ability of frataxin from different organisms to populate multiple oligomeric states in the presence of metal ions, e.g. Fe 2؉ and Co 2؉ , led to the suggestion that different oligomers contribute to the functions of frataxin. Here we report on the complex between yeast frataxin and ferrochelatase, the terminal enzyme of heme biosynthesis. Protein-protein docking and cross-linking in combination with mass spectroscopic analysis and single-particle reconstruction from negatively stained electron microscopic images were used to verify the Yfh1-ferrochelatase interactions. The model of the complex indicates that at the 2:1 Fe 2؉ -to-protein ratio, when Yfh1 populates a trimeric state, there are two interaction interfaces between frataxin and the ferrochelatase dimer. Each interaction site involves one ferrochelatase monomer and one frataxin trimer, with conserved polar and charged amino acids of the two proteins positioned at hydrogen-bonding distances from each other. One of the subunits of the Yfh1 trimer interacts extensively with one subunit of the ferrochelatase dimer, contributing to the stability of the complex, whereas another trimer subunit is positioned for Fe 2؉ delivery. Single-turnover stopped-flow kinetics experiments demonstrate that increased rates of heme production result from monomers, dimers, and trimers, indicating that these forms are most efficient in delivering Fe 2؉ to ferrochelatase and sustaining porphyrin metalation. Furthermore, they support the proposal that frataxin-mediated delivery of this potentially toxic substrate overcomes formation of reactive oxygen species.In an oxidative environment, like that of the mitochondrial matrix (1), free Fe 2ϩ is rapidly oxidized to Fe 3ϩ with the subsequent formation of insoluble Fe(OH) 3 . This type of Fe 2ϩ oxidation generally produces oxygen radicals. In addition, through the Fenton reaction in which Fe 2ϩ reacts with hydrogen peroxide, highly toxic hydroxyl radicals are produced. Organisms have evolved mechanisms for the control of iron uptake, delivery, storage, and detoxification, including those for Fe 2ϩ handling within mitochondria. Frataxin, a major mitochondrial protein player in this process, has a central role in iron detoxification and iron delivery in heme and iron-sulfur cluster (ISC) 5 synthesis (2-5) as well as in aconitase repair (6). Low levels of frataxin in humans are responsible for the progressive neurodegenerative disease Friedreich's ataxia, caused by trinucleotide repeat expansions in the first intron of the frataxin gene and the consequent gene silencing. Frataxin deficiency results in aberrations in cellular iron homeostasis, progressive accumulation of iron in mitochondria, high levels of oxidative stress, and deficiency in heme and ISC biosynthesis (4, 7-9).Numerous studies focusing on human and Saccharomyces cerevisiae (Yfh1) frataxin (FXN) and their Esch...

show abstract

“…This site is reminiscent of the ferroxidation site of E. coli ferritin (Fig. 13D) (57); and interestingly, in Yfh1, residues His-74, Asp-78, Asp-79, Asp-82, and His-83 compose the ferroxidation site, and residues Asp-101 and Glu-103 are part of the mineralization site of the protein (28,58). Alignment of cobalt-bound CyaY monomer (PDB code 2EFF) identified a fourth potential iron-binding site formed by residue Asp-104 (Asp-82 in Yfh1) from trimer 1 and Asp-124 (Asp-101 in Yfh1) from trimer 2.…”

Section: Sub-complexes (I)mentioning

confidence: 97%