Functional analysis of the second and third complement component (C3) in carp using anti-peptide antibodies.

我们描述了从硬骨鱼鲤鱼（Cyprinus carpio）中分离的两个趋异的B/C2样cDNA克隆（B/C2-A和B/C2-B）。B/C2-A与目前报道的脊椎动物的B和C2具有相同的结构域结构，并且与斑马鱼B和青鳉鱼B/C2显示出密切的相似性。这些硬骨鱼类序列与高等脊椎动物的C2和B序列具有几乎相同的相似性。相比之下，B/C2-B具有新的结构特征，因为它包含四个SCR模块，并且在系统发育分析中没有近亲。北方杂交结果表明，B/C2-A和B/C2-B在鲤鱼肝胰腺中存在两个大小不同的转录本。本文还讨论了两个鲤鱼基因序列与哺乳动物C2和B基因序列之间可能的系统发育关系。我们还分离出8个不同的cDNA克隆编码C3从一个单一的鲤鱼（鲤鱼）。这8个序列被分为5种C3类型，命名为C3-H1、C-H2、C3-S、C3-Q1和C3-Q2。另一方面，从一条鲤鱼的血清中纯化了五种不同的C3形式，命名为C3-1至C3-5。N-末端测序和与[3 H]甘氨酸的共价结合鉴定C3- 1为C3-S的翻译产物，而C3-2为C3-H1的翻译产物，C3-5为C3-H2的翻译产物。C3-1的溶血活性比C3-2高3倍，而C3-5无活性，表明硫酯催化机制不是C3活性的必要决定因素，缺乏催化组氨酸的C3在鲤鱼和可能其他骨鱼的补体系统中起着重要作用。

We describe two diverged B/C2-like cDNA clones (B/C2-A and B/C2-B) isolated from a bony fish, common carp (Cyprinus carpio). B/C2-A shares the same domain structure with B and C2 of the vertebrate reported so far, and shows a close similarity to zebrafish B and medaka fish B/C2. These teleost sequences show almost the same similarity to C2 and B of higher vertebrates. In contrast, B/C2-B has a novel structural feature in that it contains four SCR modules and does not have a close relative upon phylogenetic analysis. Northern blotting revealed the presence of two transcripts with different size for each of B/C2-A and B/C2-B in the hepatopancreas of carp. A possible phylogenetic relationship between two carp sequences, and mammalian C2 and B is discussed. We have also isolated eight distinct cDNA clones encoding C3 from a single carp (Cyprinus carpio). The eight sequences were grouped into five C3 types, designated C3-H1, C-H2, C3-S, C3-Q1, and C3-Q2. On the other hand, five distinct C3 forms, named C3-1 to C3-5, were purified from serum of a single carp. N-terminal sequencing and covalent binding to [3H] glycine identified C3-1as the translated product of C3-S, while C3-2 was that of C3-H1, and C3-5 that of C3-H2. C3-1 showed a hemolytic activity three-fold higher than that of C3-2, whereas C3-5 was inactive, suggesting that the thioester-catalytic mechanism is not a necessary determinant for the C3 activity and that C3 lacking the catalytic histidine plays a significant role in the complement system of carp and probably other bone fish.