Wu Albert M, Wu June H, Singh Tanuja, Liu Jia-Hau, Herp Anthony
Glyco-Immunochemistry Research Laboratory, Institute of Molecular and Cellular Biology, Chang-Gung University, Kwei-san, Tao-yuan, 333, Taiwan.
Life Sci. 2004 Jul 16;75(9):1085-103. doi: 10.1016/j.lfs.2004.02.016.
Anguilla anguilla agglutinin (AAA) is a fucose-specific lectin found in the serum of the fresh water eel. It is suggested to be associated with innate immunity by recognizing disease-associated cell surface glycans, and has been widely used as a reagent in hematology and glycobiology. In order to gain a better understanding of AAA for further applications, it is necessary to elucidate its binding profile with mammalian glycotopes. We, therefore, analyzed the detailed carbohydrate specificity of AAA by enzyme-linked lectinosorbent assay (ELLSA) with our extended glycan/ligand collection and lectin-glycan inhibition assay. Among the glycans tested, AAA reacted well with nearly all human blood group Ah (GalNAcalpha1-->3[LFucalpha1-->2]Gal), Bh (Galalpha1-->3[LFucalpha1-->2]Gal), H LFucalpha1-->2Gal) and Leb (Fucalpha1-->2Galbeta1-->3[Fucalpha1-->4]GlcNAc) active glycoproteins (gps), but not with blood group Lea (Galbeta1-->3[Fucalpha1-->4]GlcNAc) substances, suggesting that residues and optimal density of alpha1-2 linked LFuc to Gal at the non-reducing end of glycoprotein ligands are essential for lectin-carbohydrate interactions. Blood group precursors, Galbeta1-3GalNAc (T), GalNAcalpha1-Ser/Thr (Tn) containing glycoproteins and N-linked plasma gps, gave only negligible affinity. Among the mammalian glycotopes tested, Ah, Bh and H determinants were the best, being about 5 to 6.7 times more active than LFuc, but were weaker than p-nitrophenylalphaFuc indicating that hydrophobic environment surrounding the LFuc moiety enhance the reactivity. The hierarchy of potency of oligo- and monosaccharides can be ranked as follows: p-nitrophenyl-alphaFuc > Ah, Bh and H > LFuc > LFucalpha1-->2Galbeta1-->4Glc (2'-FL) and Galbeta1-->4[LFucalpha1-->3]Glc (3'-FL), while LNDFH I (Leb hexa-), Lea, Lex (Galbeta1-->4[Fucalpha1-->3]GlcNAc), and LDFT (gluco-analogue of Ley) were inactive. From the present observations, it can be concluded that the combining site of AAA should be a small cavity-type capable of recognizing mainly H/crypto H and of binding to specific polyvalent ABH and Leb glycotopes.
欧洲鳗鲡凝集素（AAA）是一种在淡水鳗鱼血清中发现的岩藻糖特异性凝集素。它被认为通过识别与疾病相关的细胞表面聚糖与先天免疫相关，并已被广泛用作血液学和糖生物学中的试剂。为了更好地了解AAA以便进一步应用，有必要阐明其与哺乳动物糖表位的结合特征。因此，我们通过酶联凝集素吸附测定（ELLSA）以及扩展的聚糖/配体集合和凝集素 - 聚糖抑制测定，分析了AAA的详细碳水化合物特异性。在所测试的聚糖中，AAA与几乎所有人类血型A抗原（GalNAcα1→3[L - Fucα1→2]Gal）、B抗原（Galα1→3[L - Fucα1→2]Gal）、H抗原（L - Fucα1→2Gal）和Leb抗原（Fucα1→2Galβ1→3[Fucα1→4]GlcNAc）活性糖蛋白（gp）反应良好，但与血型Lea抗原（Galβ1→3[Fucα1→4]GlcNAc）物质不反应，这表明糖蛋白配体非还原端α1 - 2连接的L - Fuc与Gal的残基和最佳密度对于凝集素 - 碳水化合物相互作用至关重要。血型前体、含有Galβ1 - 3GalNAc（T抗原）、GalNAcα1 - Ser/Thr（Tn抗原）的糖蛋白以及N - 连接的血浆gp，亲和力可忽略不计。在所测试的哺乳动物糖表位中，A抗原、B抗原和H抗原决定簇是最佳的，活性比L - Fuc高约5至6.7倍，但比对硝基苯基α - Fuc弱，这表明L - Fuc部分周围的疏水环境增强了反应性。寡糖和单糖的活性等级可排列如下：对硝基苯基 - α - Fuc＞A抗原、B抗原和H抗原＞L - Fuc＞L - Fucα1→2Galβ1→4Glc（2'-FL）和Galβ1→4[L - Fucα1→3]Glc（3'-FL），而LNDFH I（Leb六糖）、Lea抗原、Lex抗原（Galβ1→4[Fucα1→3]GlcNAc）和LDFT（Ley的葡萄糖类似物）无活性。从目前的观察结果可以得出结论，AAA的结合位点应该是一种小腔型，主要能够识别H/隐H抗原，并与特定的多价ABH和Leb糖表位结合。
