Iron Deposition and Mobilization in Ferritin

描述（由申请人提供）：从微生物到人类，铁蛋白在铁的生物管理中起着核心作用。铁蛋白通过将铁作为水合氧化铁矿物沉积在其壳状结构内而起到铁储存和解毒蛋白的作用。这种铁随后可以被动员用于血红素的合成。虽然来自各种生物体的铁蛋白具有许多共同的结构特征，是12或24个亚基蛋白，但它们在铁沉积的化学性质上明显不同。所有已知的铁蛋白都含有亚铁氧化酶位点，可以通过分子氧或过氧化氢催化铁（II）的氧化;然而，它们以显着不同的方式这样做。本论文主要研究了重组铁蛋白的铁沉积机制，包括人H-和L-链铁蛋白、新发现的人线粒体铁蛋白、含血红素的E. coli细菌铁蛋白（EcBFR）和E.北方螺旋细菌铁蛋白（EcFtnA）。还将对12聚体蛋白：无害李斯特菌铁蛋白和DNA结合蛋白，来自大肠杆菌的Dps进行研究。杆菌重要的问题，有关分子氧结合，铁蛋白铁蛋白协会在铁氧化，瞬态自由基和铁中间体和化学计量方程的铁氧化和水解使用分子氧和过氧化氢作为氧化剂将解决不同的蛋白质。将通过定点突变结合X射线结构数据、等温滴定量热法、紫外可见停流动力学、快速冷冻猝灭穆斯堡尔谱和EPR谱、自旋捕获、光散射、血氧测定和pH统计来阐明各种铁蛋白的铁沉积机制的特征。广泛的研究建议应导致详细了解各种铁蛋白如何作为可逆的铁储存蛋白，帮助细胞科普氧化应激，并进一步了解铁生物矿化过程的化学和生物化学一般。

DESCRIPTION (provided by applicant): From microorganisms to mankind, ferritin plays a central role in the biological management of iron. The ferritins function as iron storage and detoxification proteins by depositing iron as a hydrous ferric oxide mineral within their shell-like structures. This iron can be subsequently mobilized for the synthesis of heme. While ferritins from various organisms share many common structural features, being either 12 or 24 subunit proteins, they differ markedly in their chemistries of iron deposition. All known ferritins contain ferroxidase sites that catalyze iron(II) oxidation by either molecular oxygen or hydrogen peroxide; however they do so in significantly different ways. This proposal focuses on the mechanisms of iron deposition in a variety of recombinant 24mer ferritins that include human H- and L-chain ferritins, a newly discovered human mitochondrial ferritin, the heme-containing E. coli bacterioferritin (EcBFR) and the E. coil northeme bacterial ferritin (EcFtnA). Studies will also be conducted with the l2mer proteins: Listeria innocua ferritin and the DNA binding protein, Dps from E. coli. Important questions relating to dioxygen binding, ferritin-ferritin association during iron oxidation, transient radical and iron intermediates and the stoichiometric equations for iron oxidation and hydrolysis using dioxygen and hydrogen peroxide as oxidants will be addressed for the different proteins. Features of the mechanisms of iron deposition of the various ferritins will be elucidated through a combination of site-directed mutagenesis in conjunction with x-ray structure data, isothermal titration calorimetry, UV-visible stopped-flow kinetics, rapid-freeze quench Mossbauer and EPR spectroscopies, spin trapping, light scattering, oximetry and pH stat. The extensive studies proposed should lead to a detailed understanding of how various ferritins function as reversible iron storage proteins, assisting the cell cope with oxidative stress, and further our knowledge of the chemistry and biochemistry of iron biomineralization processes in general.