Levi S, Yewdall S J, Harrison P M, Santambrogio P, Cozzi A, Rovida E, Albertini A, Arosio P
Department of Biomedical Science and Technology, San Raffaele Institute, University of Milano, Italy.
Biochem J. 1992 Dec 1;288 ( Pt 2)(Pt 2):591-6. doi: 10.1042/bj2880591.
The ability to incorporate iron in vitro was studied in homopolymers of human ferritin L-chain, human ferritin H-chain and its variants and in homopolymer mixtures. The H-chain variants carried amino acid substitutions in the ferroxidase centre and/or in carboxy residues on the cavity surface. Iron incorporation was examined by gel electrophoresis of the reaction products by staining for iron and protein. It was found that inactivation of the ferroxidase centre combined with the substitution of four carboxy groups on the cavity abolished the ability of H-chain ferritin to incorporate iron. Competition experiments with limited amounts of iron showed that, at neutral pH, L-chain ferritin is more efficient in forming iron cores than the H-chain variants altered at the ferroxidase activity or in the cavity. Competition experiments at pH 5.5 demonstrated that L-chain apoferritin is able to incorporate iron only when in the presence of H-chain variants with ferroxidase activity. The results indicate that L-chain apoferritin has a higher capacity than the H-chain apoferritin to induce iron-core nucleation, whereas H-chain ferritin is superior in promoting Fe(II) oxidation. The finding of cooperative roles of the H- and L-chains in ferritin iron uptake provides a clue to understanding the biological function of isoferritins.
研究了人铁蛋白L链、人铁蛋白H链及其变体的同聚物以及同聚物混合物在体外结合铁的能力。H链变体在铁氧化酶中心和/或腔表面的羧基残基上有氨基酸取代。通过对反应产物进行凝胶电泳，并用铁和蛋白质染色来检测铁的掺入情况。结果发现，铁氧化酶中心失活并结合腔上四个羧基的取代会消除H链铁蛋白结合铁的能力。用有限量铁进行的竞争实验表明，在中性pH值下，L链铁蛋白比在铁氧化酶活性或腔内发生改变的H链变体更有效地形成铁芯。在pH 5.5下的竞争实验表明，L链脱铁铁蛋白只有在存在具有铁氧化酶活性的H链变体时才能结合铁。结果表明，L链脱铁铁蛋白比H链脱铁铁蛋白具有更高的诱导铁芯成核的能力，而H链铁蛋白在促进Fe(II)氧化方面更具优势。H链和L链在铁蛋白铁摄取中的协同作用的发现为理解异铁蛋白的生物学功能提供了线索。
