Chan A C, Atkinson J P
J Clin Invest. 1983 Nov;72(5):1639-49. doi: 10.1172/JCI111123.
Immunoprecipitates of human C4 from EDTA-plasma were incubated with [14C]methylamine and analyzed by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis and fluorography. In addition to finding label in the alpha-chains of the secreted (C4s) and predominant plasma (C4p) forms of C4, two additional molecules with apparent molecular weights of approximately 168,000 (p168) and approximately 125,000 (p125) covalently incorporated methylamine, indicating the presence of an internal thioester bond. These two molecules were present at a concentration of approximately 5% of total plasma C4 and were not immunoprecipitated by antisera to C3 or alpha 2-macroglobulin. A human hepatoma-derived cell line (HepG2), in addition to synthesizing C4s and small quantities of the polypeptide precursor of C4 (pro-C4), was found to secrete p168 and p125 at concentrations of 14 +/- 4.8 and 21 +/- 9.2% (mean +/- SD), respectively, of total secreted C4. These molecules were not found intracellularly. Both molecules were present on reduced, but not nonreduced, SDS-polyacrylamide gels. Chido (C4B) and Rodgers' (C4A) alloantisera precipitated the C4A and C4B variants of pro-C4, p168, p125, and C4s. Both tryptic and Staphylococcus aureus V8 protease peptide analyses showed homology between p168 and the beta- and alpha-chains and between p125 and the alpha- and gamma-chains. Partial NH2-terminal sequencing revealed that the beta-chain was NH2-terminal in p168 and that the alpha-chain was NH2-terminal in p125. Taken together, these data indicate that p168 and p125 represent uncleaved beta-alpha- and alpha-gamma-fragments of pro-C4, respectively. Thus, in most individuals, plasma C4 consists of five structurally distinct molecules, the single polypeptide precursor (pro-C4), the three-subunit secreted (C4s) and predominant plasma (C4p) forms of C4, and two incompletely processed two-subunit molecules with uncleaved beta-alpha- (p168) or uncleaved alpha-gamma (p125)-subunits. In addition, all five molecules are observed for both C4A (Rodgers) and C4B (Chido) structural genes.
将来自乙二胺四乙酸（EDTA）血浆的人C4免疫沉淀物与[14C]甲胺一起孵育，然后通过十二烷基硫酸钠（SDS）-聚丙烯酰胺凝胶电泳和荧光自显影进行分析。除了在分泌型（C4s）和主要血浆型（C4p）C4的α链中发现放射性标记外，另外两个表观分子量约为168,000（p168）和约125,000（p125）的分子也共价结合了甲胺，这表明存在内部硫酯键。这两个分子的浓度约为血浆总C4的5%，并且不会被抗C3或α2-巨球蛋白的抗血清免疫沉淀。一种源自人肝癌的细胞系（HepG2），除了合成C4s和少量C4的多肽前体（pro-C4）外，还被发现分别以总分泌C4的14±4.8%和21±9.2%（平均值±标准差）的浓度分泌p168和p125。这些分子在细胞内未被发现。在还原型而非非还原型SDS-聚丙烯酰胺凝胶上都出现了这两种分子。奇多（C4B）和罗杰斯（C4A）同种抗血清沉淀了pro-C4、p168、p125和C4s的C4A和C4B变体。胰蛋白酶和金黄色葡萄球菌V8蛋白酶肽分析均显示p168与β链和α链之间以及p125与α链和γ链之间具有同源性。部分氨基末端测序显示，在p168中β链是氨基末端，在p125中α链是氨基末端。综合这些数据表明，p168和p125分别代表未切割的pro-C4的β-α片段和α-γ片段。因此，在大多数个体中血浆C4由五个结构不同的分子组成：单一的多肽前体（pro-C4）、分泌型（C4s）的三亚基和主要血浆型（C4p）的C4，以及两个加工不完全的二亚基分子，即未切割的β-α（p168）或未切割的α-γ（p125）亚基。此外，对于C4A（罗杰斯）和C4B（奇多）结构基因都观察到了所有这五个分子。
