El Ayeb M, Bahraoui E M, Granier C, Delori P, Van Rietschoten J, Rochat H
Mol Immunol. 1984 Mar;21(3):223-32. doi: 10.1016/0161-5890(84)90077-4.
We report the isolation and characterization of two IgG populations specific to two synthetic peptides corresponding to two antigenic sites of toxin II of the North African scorpion Androctonus australis Hector. Firstly, thanks to the use of: (1) antigenic homology studies between toxin II of A. australis Hector and toxin III of Buthus occitanus tunetanus, (2) chemical modification of toxin II of A. australis Hector, and (3) prediction of the localization of the four major antigenic sites of scorpion alpha-toxins by the method developed by Hopp and Woods [Proc. natn. Acad. Sci. U.S.A. 78, 3824-3828 (1981)], we have established that the region around the disulfide bridge between cysteines 12 and 63 as well as the stretch of residues 50-59 probably each enclosed an antigenic site. Secondly, the synthetic replicates of these regions linked to Sepharose allowed us to isolate, by immunoaffinity chromatography, two IgG populations from the whole anti-toxin II of A. australis Hector IgGs. Finally, each of these two IgG populations was shown to be specific to one antigenic site as evidenced by the multideterminant effect on the slopes of binding curves developed by Berzofsky et al. [Biochemistry 15, 2113-2121 (1976)]. Furthermore, these two IgG populations were found to be functionally independent and this could be related to the fact that the two regions carrying the two antigenic sites are not close to each other in space and that there is neither steric hindrance nor cooperative effects between them. The association constant of these site-specific IgG populations was calculated and found to be equal to 1.18-5.14 X 10(9) l/mole for IgG anti-site 1 and 1.16-5.62 X 10(9) l/mole for IgG anti-site 2 respectively by Sips [J. chem. Phys. 16, 490-495 (1948)], Scatchard [Am. N.Y. Acad. Sci. 51, 660-772 (1949)] and Steward and Petty [Immunology 23, 881-887 (1972)] representations. The index of heterogeneity of 0.9 for anti-P1 and anti-P2 indicates the purification of essentially homogeneous affinity IgG populations.
我们报道了针对与北非蝎子黄肥尾蝎（Androctonus australis Hector）毒素II的两个抗原位点相对应的两种合成肽的两个IgG群体的分离和表征。首先，由于使用了：（1）黄肥尾蝎毒素II与突尼斯杀牛蝎（Buthus occitanus tunetanus）毒素III之间的抗原同源性研究，（2）黄肥尾蝎毒素II的化学修饰，以及（3）通过霍普（Hopp）和伍兹（Woods）开发的方法[美国国家科学院院刊78, 3824 - 3828 (1981)]预测蝎子α-毒素的四个主要抗原位点的定位，我们确定了半胱氨酸12和63之间二硫键周围的区域以及50 - 59位残基片段可能各自包含一个抗原位点。其次，与琼脂糖凝胶连接的这些区域的合成复制品使我们能够通过免疫亲和色谱从黄肥尾蝎IgG的全抗毒素II中分离出两个IgG群体。最后，如贝佐夫斯基（Berzofsky）等人[生物化学15, 2113 - 2121 (1976)]所开发的结合曲线斜率上的多决定簇效应所证明，这两个IgG群体中的每一个都被证明对一个抗原位点具有特异性。此外，发现这两个IgG群体在功能上是独立的，这可能与携带两个抗原位点的两个区域在空间上彼此不接近且它们之间既没有空间位阻也没有协同效应这一事实有关。通过西普斯（Sips）[化学物理杂志16, 490 - 495 (1948)]、斯卡查德（Scatchard）[美国纽约科学院学报51, 660 - 772 (1949)]以及斯图尔特（Steward）和佩蒂（Petty）[免疫学23, 881 - 887 (1972)]的表示法分别计算出这些位点特异性IgG群体的缔合常数，发现抗位点1的IgG为1.18 - 5.14×10⁹ l/摩尔，抗位点2的IgG为1.16 - 5.62×10⁹ l/摩尔。抗P1和抗P2的异质性指数为0.9，表明纯化得到了基本上均一的亲和IgG群体。
