Isolation and characterization of the third component of complement in the serum of the clawed frog, Xenopus laevis.
Isolation and characterization of the third component of complement in the serum of the clawed frog, Xenopus laevis.
复制标题
爪蛙（Xenopus laevis）血清中补体第三种成分的分离和表征。
DOI：
10.4049/jimmunol.133.3.1436
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发表时间：
1984
影响因子：
4.4
通讯作者：
C. Katagiri
中科院分区：
文献类型：
--
作者：
A. Sekizawa;T. Fujii;C. Katagiri
正常爪蟾血清对SRBC的溶血活性依赖于特异性抗体和Ca ~（++）、Mg ~（++），而对RRBC的溶血活性仅依赖于Mg ~（++）。这两种溶血活性消失后的血清与酵母聚糖或特异性兔抗血清对酵母聚糖结合蛋白的爪蟾血清。以该抗血清为探针，经聚乙二醇沉淀、DEAE-Sepharose CL-6 B、Sepharose CL-6 B和Sephadex G-200柱层析，从爪蟾血浆中纯化出单一的补体成分（XC）。在正常血清中含有2.3 mg/ml的XC，显示出β-球蛋白的电泳迁移率，其m.w.的204，000（204 K），包括125 K和85 K两个不同的亚基，它们通过二硫键彼此连接。204 K蛋白与爪蟾血清中的其它组分一起表现出强的溶血活性。胰蛋白酶消化204 K蛋白导致125 K亚基的特异性切割，并将其免疫电泳迁移率转换为阳极侧，使85 K亚基保持完整。用SDS和尿素处理XC导致125 K亚基分裂成78 K和40 K，但这种分裂在用甲胺预处理后被抑制，表明XC中存在硫酯键。XC的氨基酸组成与哺乳动物C3的氨基酸组成惊人相似。204 K蛋白（XC）在非洲爪蟾溶血过程中起着重要的作用，在经典溶血途径和旁路溶血途径中均起着重要作用。
The hemolytic activity against SRBC in the serum of normal Xenopus is dependent on specific antibody and both Ca++ and Mg++, whereas the activity against RRBC is dependent on Mg++ alone. Both of these hemolytic activities disappeared after treatment of the serum with zymosan or with the specific rabbit antiserum against one of the zymosan-binding proteins in Xenopus serum. By using this antiserum as a probe, a complement component (XC) was purified as a single entity from the Xenopus plasma after polyethylene glycol precipitation, DEAE-Sepharose CL-6B, Sepharose CL-6B, and Sephadex G-200 column chromatographies. The XC, contained at 2.3 mg/ml in normal serum, showed an electrophoretic mobility of beta-globulin, with a m.w. of 204,000 (204K) comprising two distinct subunits of 125K and 85K, which are linked with each other by disulfide bonds. The 204K protein exhibited a strong hemolytic activity in association with other components in Xenopus serum. Digestion of 204K protein by trypsin resulted in a specific cleavage of the 125K subunit and a conversion of its immunoelectrophoretic mobility to the anodal side, leaving the 85K subunits intact. The treatment of XC with SDS and urea resulted in the splitting of 125K subunits into 78K and 40K, but this splitting was inhibited upon pretreatment with methylamine, suggesting the presence of a thiol ester bond in the XC. The amino acid composition of the XC revealed a striking resemblance to that of mammalian C3. In all aspects, the 204K protein (XC) is regarded as representing the C3 of Xenopus laevis, which plays a key role in both the classical and alternative hemolytic pathways.