Kirkeby S, Moe D
Department of Oral Function and Physiology, The Panum Institute, Nørre Allé 20, Copenhagen 2200 N, Denmark.
Immunol Cell Biol. 2001 Apr;79(2):121-7. doi: 10.1046/j.1440-1711.2001.00992.x.
Glycoconjugates with terminal Galalpha1-3Galbeta1-4GlcNAc sequences (alpha-galactosyl epitopes, natural xenoreactive antigens) are present on various tissues in pigs and are recognized by human anti-alphagalactosyl (alphaGal) antibodies1. Hence xenotransplantation (pig-to-human) would trigger immune reactions involving complement activation and lead to the hyperactute rejection of the graft. Xenoreactive antigens are often studied by using the lectin Griffonia simplicifolia 1 isolectin B4 (GS1 B4), which shows high affinity to galactose. We here estimate the specificity of GS1 B4 for detecting various galactosyl epitopes by measuring lectin binding to neoglycoproteins, thyroglobulin and pig skeletal muscle. Enzyme linked lectin assays confirmed that GS1 B4 was highly specific to alpha-galactosylated neoglycoproteins while the lectin did not detect a beta-galactosylated ligand. The length of the sugar chains influenced the lectin-carbohydrate interaction. A monosaccharide linked to serum albumin showed higher lectin affinity than did neoglycoproteins with di- and tri-alpha-galactosyl epitopes. When the carbohydrate was extended, as in the xenoreactive pentasaccharide (Galalpha1-3Galbeta1-4GlcNAcbeta1-3Galbeta1-4Glc), the carbohydrate- lectin interaction was meagre. Not only the terminal, but also the subterminal sugar affected the lectin binding because the GS1 B4 affinity to Galalpha1-3Gal was much stronger than to Galalpha1-3GalNAc. In bovine and porcine thyroglobulin most alphaGal epitopes appear to be cryptic, but are unmasked by a heat denaturation. In pig skeletal muscle there was lectin reaction not only in the muscle capillaries, but also in the connective tissue and intracellularly in muscle fibres. In Western blots of isolated proteins from pig muscle at least three bands were strongly stained after incubation with lectin.
带有末端Galα1-3Galβ1-4GlcNAc序列的糖缀合物（α-半乳糖基表位，天然异种反应性抗原）存在于猪的各种组织中，并可被人类抗α-半乳糖基（αGal）抗体识别。因此，异种移植（猪到人的移植）会引发涉及补体激活的免疫反应，并导致移植物的超急性排斥反应。异种反应性抗原通常通过使用凝集素单叶豆凝集素1异凝集素B4（GS1 B4）进行研究，该凝集素对半乳糖具有高亲和力。我们在此通过测量凝集素与新糖蛋白、甲状腺球蛋白和猪骨骼肌的结合，评估GS1 B4检测各种半乳糖基表位的特异性。酶联凝集素测定证实，GS1 B4对α-半乳糖基化新糖蛋白具有高度特异性，而该凝集素未检测到β-半乳糖基化配体。糖链的长度影响凝集素与碳水化合物的相互作用。与血清白蛋白相连的单糖比具有二-α-半乳糖基表位和三-α-半乳糖基表位的新糖蛋白表现出更高的凝集素亲和力。当碳水化合物延长时，如在异种反应性五糖（Galα1-3Galβ1-4GlcNAcβ1-3Galβ1-4Glc）中，碳水化合物与凝集素的相互作用很微弱。不仅末端糖，亚末端糖也影响凝集素结合，因为GS1 B4对Galα1-3Gal的亲和力比对Galα1-3GalNAc的亲和力强得多。在牛和猪的甲状腺球蛋白中，大多数αGal表位似乎是隐蔽的，但可通过热变性暴露出来。在猪骨骼肌中，不仅在肌毛细血管中有凝集素反应，在结缔组织和肌纤维细胞内也有凝集素反应。在用凝集素孵育后，猪肌肉分离蛋白的Western印迹中至少有三条带被强烈染色。
