Ferritin reactions: Direct identification of the site for the diferric peroxide reaction intermediate
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- Xiaofeng Liu
- Center for BioIron at Children's Hospital Oakland Research Institute, 5700 Martin Luther King Way, Oakland, CA 94609
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- Elizabeth C. Theil
- Center for BioIron at Children's Hospital Oakland Research Institute, 5700 Martin Luther King Way, Oakland, CA 94609
抄録
<jats:p> Ferritins managing iron–oxygen biochemistry in animals, plants, and microorganisms belong to the diiron carboxylate protein family and concentrate iron as ferric oxide ≈10 <jats:sup>14</jats:sup> times above the ferric <jats:italic>K</jats:italic> <jats:sub>s</jats:sub> . Ferritin iron (up to 4,500 atoms), used for iron cofactors and heme, or to trap DNA-damaging oxidants in microorganisms, is concentrated in the protein nanocage cavity (5–8 nm) formed during assembly of polypeptide subunits, 24 in maxiferritins and 12 in miniferritins/DNA protection during starvation proteins. Direct identification of ferritin ferroxidase (F <jats:sub>ox</jats:sub> ) sites, complicated by multiple types of iron–ferritin interactions, is now achieved with chimeric proteins where putative F <jats:sub>ox</jats:sub> site residues were introduced singly and cumulatively into an inactive host, an L maxiferritin. A dimagnesium ferritin cocrystal model guided site design and the diferric peroxo F <jats:sub>ox</jats:sub> intermediates ( <jats:italic>A</jats:italic> at 650 nm) monitored activity. Diferric peroxo formation in chimeric and WT proteins had similar <jats:italic>K</jats:italic> <jats:sub>app</jats:sub> values and Hill coefficients. Catalytic activity required cooperative ferrous substrate binding to two sites A (E, EXXH) and B (E, QXXD). The weaker B sites in ferritin contrast with stronger B sites (E, EXXH) in diiron carboxylate oxygenases, explaining diferric oxo/hydroxo product release in ferritin vs. diiron cofactor retention in oxygenases. Codons for Q/H and D/E differ by single nucleotides, suggesting simple DNA mutations relate site B diiron substrate sites and diiron cofactor sites in proteins. The smaller <jats:italic>k</jats:italic> <jats:sub>cat</jats:sub> values in chimeras indicate the absence of second-shell residues important for ferritin substrate–product channeling that, when identified, will outline the entire iron path from ferritin pores through the F <jats:sub>ox</jats:sub> site to the mineral cavity. </jats:p>
収録刊行物
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- Proceedings of the National Academy of Sciences
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Proceedings of the National Academy of Sciences 101 (23), 8557-8562, 2004-05-27
Proceedings of the National Academy of Sciences
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詳細情報 詳細情報について
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- CRID
- 1363670318308409088
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- NII論文ID
- 30016240915
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- ISSN
- 10916490
- 00278424
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- データソース種別
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