Meldrum F C, Douglas T, Levi S, Arosio P, Mann S
School of Chemistry, University of Bath, UK.
J Inorg Biochem. 1995 Apr;58(1):59-68. doi: 10.1016/0162-0134(94)00037-b.
The formation of Mn(III) oxyhydroxide (MnOOH) cores within the nanoscale cavity of the iron storage protein ferritin has been investigated by electron microscopy and visible absorption spectroscopy. At pH 8.9, discrete amorphous MnOOH cores were formed within horse spleen apoferritin at a range of metal:protein ratios, as well as in ferritin molecules seeded with a small ferrihydrite nucleus. Analysis of the resultant core size distributions showed that the reconstitution of horse spleen apoferritin with Mn(II) was similar to that observed previously for Fe(II) reconstitution in recombinant human L-chain ferritin, suggesting that horse spleen apoferritin does not exhibit Mn(II) oxidase activity at pH 8.9. Reconstitution with MnOOH shows essentially "all-or-nothing" behavior in which many protein molecules remain unmineralized whilst others are loaded to maximum capacity. Kinetic studies showed no significant differences between horse spleen ferritin, recombinant H- and L-chain homopolymers, and H-chain variants containing site-directed modifications at the ferroxidase and putative Fe nucleation centers. Our results indicate that the reconstitution of ferritin with MnOOH cores proceeds by a nonspecific pathway. We propose that the outer surface of the protein inhibits the development of MnOOH nuclei in bulk solution whereas the inner surface is inactive, enabling nucleation and growth to proceed unperturbed within the cavity. One possibility is that differences in the general polyelectrolyte properties of these two surfaces, rather than site-specific charges, account for the "Janus" behavior of the molecule. A similar mechanism might also increase the specificity of iron oxide mineralization in ferritins that lack ferroxidase centers.
通过电子显微镜和可见吸收光谱法研究了铁储存蛋白铁蛋白纳米级腔内氢氧化锰（MnOOH）核的形成。在pH 8.9时，在一系列金属与蛋白质比例下，马脾脱铁铁蛋白内形成了离散的无定形MnOOH核，在接种有小水铁矿核的铁蛋白分子中也是如此。对所得核尺寸分布的分析表明，马脾脱铁铁蛋白与Mn（II）的重构与之前在重组人L链铁蛋白中观察到的Fe（II）重构相似，这表明马脾脱铁铁蛋白在pH 8.9时不表现出Mn（II）氧化酶活性。用MnOOH重构显示出基本上“全有或全无”的行为，其中许多蛋白质分子未矿化，而其他分子则被加载到最大容量。动力学研究表明，马脾铁蛋白、重组H链和L链同聚物以及在铁氧化酶和假定的铁成核中心含有定点修饰的H链变体之间没有显著差异。我们的结果表明，用MnOOH核重构铁蛋白是通过非特异性途径进行的。我们提出，蛋白质的外表面抑制了本体溶液中MnOOH核的形成，而内表面则无活性，从而使成核和生长在腔内不受干扰地进行。一种可能性是，这两个表面的一般聚电解质性质的差异，而不是位点特异性电荷，解释了分子的“两面神”行为。类似的机制也可能增加缺乏铁氧化酶中心的铁蛋白中铁氧化物矿化的特异性。
