Ihara K, Maeguchi K, Young C T, Theil E C
J Biol Chem. 1984 Jan 10;259(1):278-83.
Cell-specific differences occur in the primary structure of ferritin. For example, red cell and liver ferritin from bullfrog tadpoles differ by 1.5 times in serine content. To determine if cell-specific differences in ferritin primary structure are expressed in the tetraeicosomer, which thus might distinguish the proteins in a functional state, phosphorylation in vitro was employed as a probe using [gamma-32P]ATP and the catalytic subunit from the cAMP-dependent protein kinase of bovine skeletal muscle. Subunits of both proteins in the tetraeicosomers were phosphorylated. Based on tryptic peptide maps, five regions common to both red cell and liver apoferritin were phosphorylated, as confirmed for two peptides by amino acid analyses. [32P]Apoferritin from red cells yielded an additional four 32P-fragments by mapping, at least three of which were unique by amino acid analysis and, in one case, might represent a 32P-Fe complex bound by a fragment of the iron-binding site. One peptide appeared to be unique to liver apoferritin. High concentrations of ATP yielded one additional peptide common to liver and red cell and one red cell-specific peptide in the tryptic peptide maps. The maximum moles of 32P/molecule were 13 +/- 4 and 6 +/- 2, respectively, for red cell and liver apoferritin, which corresponded within experimental error to the number of 32P-tryptic peptides. The level of phosphorylation was, on the average, not more than one site/subunit. Furthermore, above certain levels of phosphorylation, some subunits in the assemblage of 24 appeared to be unavailable as substrates, possibly because of charge repulsion or conformational changes. The possibility that post-translational modifications of ferritin which amplify cell-specific structural features occur in vivo with cytoplasmic components, e.g. protein kinases, is considered in terms of the physiological availability of iron from different iron storage cells and developmental changes in iron storage.
铁蛋白的一级结构存在细胞特异性差异。例如，牛蛙蝌蚪的红细胞和肝脏铁蛋白在丝氨酸含量上相差1.5倍。为了确定铁蛋白一级结构中的细胞特异性差异是否在二十四聚体中表现出来，从而可能在功能状态下区分这些蛋白质，体外磷酸化被用作探针，使用[γ-32P]ATP和牛骨骼肌cAMP依赖性蛋白激酶的催化亚基。二十四聚体中两种蛋白质的亚基都被磷酸化。根据胰蛋白酶肽图，红细胞和肝脏脱铁铁蛋白共有的五个区域被磷酸化，通过氨基酸分析对两个肽段进行了确认。红细胞来源的[32P]脱铁铁蛋白经图谱分析产生了另外四个32P片段，其中至少三个通过氨基酸分析是独特的，在一种情况下，可能代表一个与铁结合位点片段结合的32P-Fe复合物。有一个肽段似乎是肝脏脱铁铁蛋白特有的。高浓度的ATP在胰蛋白酶肽图中产生了一个肝脏和红细胞共有的额外肽段以及一个红细胞特异性肽段。红细胞和肝脏脱铁铁蛋白每分子的最大32P摩尔数分别为13±4和6±2，在实验误差范围内与32P-胰蛋白酶肽的数量相对应。平均磷酸化水平不超过每个亚基一个位点。此外，在高于一定磷酸化水平时，24聚体组合中的一些亚基似乎不能作为底物，可能是由于电荷排斥或构象变化。考虑到来自不同铁储存细胞的铁的生理可用性以及铁储存中的发育变化，探讨了铁蛋白的翻译后修饰在体内与细胞质成分（如蛋白激酶）一起放大细胞特异性结构特征的可能性。
