Kai C, Yoshikawa Y, Yamanouchi K, Okada H
J Immunol. 1983 Jun;130(6):2814-20.
The identification of the third component of complement (C3) of Japanese quails was attempted by using rabbit antiserum prepared against quail serum-treated zymosan (ZX) as an initial reagent. This antiserum (anti-ZX) had agglutinating activity on rabbit erythrocytes reacted with quail antibody and quail complement (EACq) but not on EAq, and developed two precipitin lines against quail serum at beta- and gamma-regions in crossed immunoelectrophoresis. Subsequently, monospecific antisera to each of these precipitin lines were prepared in rabbits, and quail serum proteins reactive with these antisera were purified by salt precipitation followed by Sephadex gel filtration and DEAE cellulose column chromatography. One protein with a m.w. of 184,000 (184K) resembled mammalian C3 in that: 1) monospecific antiserum (anti-184K protein serum) agglutinated EACq but not EAq; 2) treatment of fresh quail serum with either inulin or zymosan resulted in the conversion of the precipitin line developed against 184K protein from gamma to beta in crossed immunoelectrophoresis; 3) the 184K protein was shown to consist of two polypeptide chains of 110K and 73K linked by disulfide bonds. Furthermore, the 184K protein in serum was cleaved through the incubation with inulin to 174K and 140K proteins that might correspond to C3b and C3c of human complement; 4) the 184K protein bound to zymosan was eluted with hydrazine or methylamine but not with Nonidet P-40, indicating that 184K protein binds to zymosan by a covalent bond but not by a hydrophobic one; and 5) by treatment of fresh quail serum with methylamine, complement reactivity was reduced, although its activity was restored by the addition of purified 184K protein. These results suggest the 184K protein is the quail's equivalent to mammalian C3. When quail serum was reacted with cells that had complement-activating capacity, quail C3 deposited on their membrane as in mammalians; however, no conversion of quail C3 was noted by the reaction with CVF. Antibody to quail C3 failed to cross-react with that in mammals.
尝试以用抗鹌鹑血清处理的酵母聚糖（ZX）制备的兔抗血清作为初始试剂，来鉴定日本鹌鹑补体第三成分（C3）。这种抗血清（抗ZX）对与鹌鹑抗体和鹌鹑补体反应的兔红细胞（EACq）有凝集活性，但对EAq无凝集活性，并且在交叉免疫电泳中针对鹌鹑血清在β和γ区域产生两条沉淀线。随后，在兔中制备了针对每条沉淀线的单特异性抗血清，与这些抗血清反应的鹌鹑血清蛋白通过盐析，然后经Sephadex凝胶过滤和DEAE纤维素柱色谱法进行纯化。一种分子量为184,000（184K）的蛋白质在以下方面类似于哺乳动物C3：1）单特异性抗血清（抗184K蛋白血清）凝集EACq但不凝集EAq；2）用菊粉或酵母聚糖处理新鲜鹌鹑血清导致在交叉免疫电泳中针对184K蛋白产生的沉淀线从γ转变为β；3）184K蛋白显示由通过二硫键连接的110K和73K的两条多肽链组成。此外，血清中的184K蛋白通过与菊粉孵育被切割为174K和140K蛋白，它们可能对应于人补体的C3b和C3c；4）与酵母聚糖结合的184K蛋白用肼或甲胺洗脱，但不用Nonidet P - 40洗脱，表明184K蛋白通过共价键而非疏水键与酵母聚糖结合；5）用甲胺处理新鲜鹌鹑血清会降低补体反应性，尽管通过添加纯化的184K蛋白可恢复其活性。这些结果表明184K蛋白相当于鹌鹑的哺乳动物C3。当鹌鹑血清与具有补体激活能力的细胞反应时，鹌鹑C3如在哺乳动物中一样沉积在其膜上；然而，与眼镜蛇毒因子（CVF）反应未观察到鹌鹑C3的转化。抗鹌鹑C3抗体未能与哺乳动物的C3发生交叉反应。
