Duk M, Mitra D, Lisowska E, Kabat E A, Sharon N, Lis H
Ludwik Hirszfeld Institute of Immunology and Experimental Therapy, Polish Academy of Sciences, Wroclaw.
Carbohydr Res. 1992 Dec 15;236:245-58. doi: 10.1016/0008-6215(92)85019-v.
The specificity of the anti A+N lectin of Moluccella laevis (MLL) was examined by hemagglutination experiments with enzyme-modified human erythrocytes and by inhibition of hemagglutination. In addition, binding to various glycoproteins and inhibition by different sugars and glycoproteins were examined by enzyme immunoassay with antibodies to the lectin. Treatment of AMM erythrocytes with proteolytic enzymes increased their agglutinability by MLL 4-16-fold; similar treatment of ONN cells decreased their agglutinability 8-16-fold. This is in line with the known location and enzyme sensitivity of A and N specificity determinants. Treatment of the erythrocytes with sialidase increased their agglutinability and abolished the distinction between N and M cells. Hapten inhibition of hemagglutination of AMM and ONN erythrocytes by the lectin, and its binding to glycoproteins measured by enzyme immunoassay, confirmed the high specificity of MLL for N-acetyl-D-galactosamine (200-500 times more than for D-galactose) and suggested the presence of hydrophobic interactions around HO-2 of the D-galactose unit. The methyl alpha-glycosides of D-galactose and of N-acetyl-D-galactosamine were better inhibitors than the corresponding beta-glycosides; this preference was abolished, and sometimes reversed, when the p-nitrophenyl glycosides of the same monosaccharides were tested, stressing again the importance of hydrophobic interactions in the binding of carbohydrates to MLL. The lectin reacted well with ONN substance and with glycophorin A of the N phenotype (GPAN), but did not react with OMM substance or GPAM. The strongest inhibitor was asialo ovine submaxillary mucin, which contains many unsubstituted alpha-D-GalpNAc-(1-->3)-Ser/Thr residues; calculated per N-acetyl-D-galactosamine residue, it was 1500 stronger than free N-acetyl-D-galactosamine. In accordance with this result, it was found that the lectin strongly agglutinates Tn cells. The specificity of MLL can, thus, be defined as anti-Tn, crossreactive with blood types A and N, and with sialosyl-Tn. The N-specificity can best be explained by assuming that GPAN contains a small number of unsubstituted or partially sialylated alpha-D-GalpNAc-(1-->3)-Ser/Thr residues, which are present in smaller proportions, if at all, in GPAM.
通过用酶修饰的人红细胞进行血凝实验以及血凝抑制实验，检测了光滑野芝麻（Moluccella laevis，MLL）抗A + N凝集素的特异性。此外，通过用针对该凝集素的抗体进行酶免疫测定，检测了其与各种糖蛋白的结合以及不同糖类和糖蛋白的抑制作用。用蛋白水解酶处理AMM红细胞可使其对MLL的凝集性增加4至16倍；对ONN细胞进行类似处理则使其凝集性降低8至16倍。这与已知的A和N特异性决定簇的位置及酶敏感性一致。用唾液酸酶处理红细胞可增加其凝集性，并消除N和M细胞之间的区别。凝集素对AMM和ONN红细胞血凝的半抗原抑制作用，以及通过酶免疫测定法测定的其与糖蛋白的结合，证实了MLL对N - 乙酰 - D - 半乳糖胺具有高特异性（比对D - 半乳糖高200至500倍），并表明在D - 半乳糖单元的HO - 2周围存在疏水相互作用。D - 半乳糖和N - 乙酰 - D - 半乳糖胺的α - 甲基糖苷比相应的β - 糖苷是更好的抑制剂；当测试相同单糖的对硝基苯基糖苷时，这种偏好被消除，有时甚至逆转，这再次强调了疏水相互作用在碳水化合物与MLL结合中的重要性。该凝集素与ONN物质和N表型的血型糖蛋白A（GPAN）反应良好，但不与OMM物质或GPAM反应。最强的抑制剂是去唾液酸羊下颌粘蛋白，其含有许多未取代的α - D - GalpNAc - (1→3)-Ser/Thr残基；按每个N - 乙酰 - D - 半乳糖胺残基计算，它比游离的N - 乙酰 - D - 半乳糖胺强1500倍。根据这一结果，发现该凝集素能强烈凝集Tn细胞。因此，MLL的特异性可定义为抗Tn，与A和N血型以及唾液酸化Tn交叉反应。N特异性最好通过假设GPAN含有少量未取代或部分唾液酸化的α - D - GalpNAc - (1→3)-Ser/Thr残基来解释，而这些残基在GPAM中即使存在，比例也较小。
