Huang He-Qing, Xiao Zhi-Qun, Chen Xu, Lin Qing-Mei, Cai Zong-Wei, Chen Ping
Department of Biology, School of Life Sciences, The Key Laboratory for Chemical Biology of Fujian Province, Xiamen University 361005, China.
Biophys Chem. 2004 Nov 1;111(3):213-22. doi: 10.1016/j.bpc.2004.06.002.
The ferritin consists of a protein shell constructed of 24 subunits and an iron core. The liver ferritin of Sphyrna zygaena (SZLF) purified by column chromatography is a protein composed of eight ferritins containing varying iron numbers ranging from 400+/-20 Fe3+/SZLF to 1890+/-20 Fe3+/SZLF within the protein shell. Nature SZLF (SZLFN) consisting of holoSZLF and SZLF with unsaturated iron (SZLFUI) to have been purified with polyacrylamide gel electrophoresis (PAGE) exhibited five ferritin bands with different pI values ranging from 4.0 to 7.0 in the gel slab of isoelectric focusing (IEF). HoloSZLF purified by PAGE (SZLFE) not only had 1890+/-20 Fe3+/SZLFE but also showed an identical size of iron core observed by transmission electron microscopy (TEM). Molecular weight of approximately 21 kDa for SZLFE subunit was determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Four peaks of molecular ions at mass/charge (m/z) ratios of 10611.07, 21066.52, 41993.16, and 63555.64 that come from the SZLFE were determined by matrix-assisted laser desorption ionization/time-of-flight mass spectrometry (MALDI-TOF MS), which were identified as molecular ions of the ferritin subunit (M+) and its polymers, namely, [M]2+, [M]+, [2M]+, and [3M]+, respectively. Both SZLFE and a crude extract from shark liver of S. zygaena showed similar kinetic characteristics of complete iron release with biphasic behavior. In addition, a combined technique of visible spectrometry and column chromatography was used for studying ratio of phosphate to Fe3+ within the SZLFE core. Interestingly, this ratio maintained invariable even after the iron release, which differed from that of other mammal ferritins.
铁蛋白由一个由24个亚基构成的蛋白质外壳和一个铁核心组成。通过柱色谱法纯化的皱唇鲨肝脏铁蛋白（SZLF）是一种蛋白质，由八个铁蛋白组成，其蛋白质外壳内的铁含量从400±20 Fe³⁺/SZLF到1890±20 Fe³⁺/SZLF不等。通过聚丙烯酰胺凝胶电泳（PAGE）纯化的天然SZLF（SZLFN），即由全铁SZLF和不饱和铁的SZLF（SZLFUI）组成，在等电聚焦（IEF）凝胶板上显示出五条不同pI值（范围从4.0至7.0）的铁蛋白条带。通过PAGE纯化的全铁SZLF（SZLFE）不仅含有1890±20 Fe³⁺/SZLFE，而且通过透射电子显微镜（TEM）观察到其铁核心大小相同。通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（SDS-PAGE）测定SZLFE亚基的分子量约为21 kDa。通过基质辅助激光解吸电离/飞行时间质谱（MALDI-TOF MS）测定了来自SZLFE的质荷比（m/z）为10611.07、21066.52、41993.16和63555.64的四个分子离子峰，它们分别被鉴定为铁蛋白亚基（M⁺）及其聚合物的分子离子，即[M]²⁺、[M]⁺、[2M]⁺和[3M]⁺。SZLFE和皱唇鲨肝脏粗提物均表现出具有双相行为的完全铁释放的相似动力学特征。此外，采用可见光谱法和柱色谱法相结合的技术研究SZLFE核心内磷酸盐与Fe³⁺的比例。有趣的是，即使在铁释放后该比例仍保持不变，这与其他哺乳动物铁蛋白不同。
