HUMAN SPLENIC GALAPTIN - CARBOHYDRATE-BINDING SPECIFICITY AND CHARACTERIZATION OF THE COMBINING SITE
HUMAN SPLENIC GALAPTIN - CARBOHYDRATE-BINDING SPECIFICITY AND CHARACTERIZATION OF THE COMBINING SITE
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DOI：
10.1021/bi00474a015
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发表时间：
1990-06-05
期刊：
影响因子：
2.9
通讯作者：
MATTA, KL
中科院分区：
文献类型：
--
作者：
AHMED, H;ALLEN, HJ;MATTA, KL
从人脾中分离出一种半乳糖结合凝集素（galaptin），经去唾液酸胎球蛋白-琼脂糖凝胶亲和层析纯化。半乳糖肽的碳水化合物结合特异性已经通过分析半乳糖肽与无唾液酸胎球蛋白在假定的抑制剂存在下的结合来研究。开发了一种酶联免疫吸附试验（ELISA），其中包括将去唾液酸胎球蛋白吸附到微量滴定板上。用多克隆兔抗半乳糖肽血清检测半乳糖肽与脱唾液酸胎球蛋白的结合，然后用山羊抗兔IgG-过氧化物酶缀合物检测。相对于对照，以图形方式确定半乳糖肽结合抑制50%的抑制剂浓度，并相对于半乳糖或乳糖进行标准化。这些分析表明，半乳糖肽具有至少与二糖一样大的结合位点。具有非还原末端β-的二糖与Glc或GlcNAc连接的半乳糖残基（1，3）、（1，4）和（1，6）是比游离Gal更好的抑制剂。游离或糖苷连接的GalNAc似乎对凝集素没有亲和力。硝基苯基半乳糖苷是比甲基半乳糖苷更好的抑制剂，表明疏水相互作用的发生。数据表明，在Gal β中，Gal的C-4和C-6处的OH基团和GlcNAc的C-3处的OH基团在Gal β中的C-4和C-6处的OH基团和GlcNAc的C-3处的OH基团在Gal β中的C-4和C-6处的OH基团和GlcNAc的C-3处的OH基团在Gal β中的C-4和C-6处的OH基团和GlcNAc的C-3处的OH基团在Gal β中的C-3处的OH基团在Gal β中的C-4和C-6处的OH基团和GlcNAc的C-3处的OH基团之间存在差异。（1，4）GlcNAc对于凝集素糖相互作用是重要的。我们的数据支持的假设，半乳糖肽的内源性受体最有可能是乳糖胺聚糖部分。
A galactose-binding lectin (galaptin) from human spleen has been purified to homogeneity by affinity chromatography on asialofetuin-Sepharose. The carbohydrate-binding specificity of galaptin has been investigated by analyzing the binding of galaptin to asialofetuin in the presence of putative inhibitors. An enzyme-linked immunosorbent assay (ELISA) was developed that involved adsorption of asialofetuin to microtiter plates. Galaptin bound to asialofetuin was detected with polyclonal rabbit anti-galaptin serum followed by goat anti-rabbit IgG-peroxidase conjugate. The concentrations of inhibitors giving 50% inhibition of galaptin binding relative to controls were graphically determined and normalized relative to galactose or lactose. These analyses revealed that galaptin has a combining site at least as large as a disaccharide. The disaccharides having non-reducing-terminal .beta.-galactosyl residues linked (1,3) (1,4), and (1,6) to Glc or GlcNAc are better inhibitors than free Gal. GalNAc, either free or glycosidically linked, appears to have no affinity for the lectin. The nitrophenyl galactosides are better inhibitors than methyl galactosides, indicating the occurrence of hydrophobic interactions. The data indicate that OH groups at C-4 and C-6 of Gal and the OH at C-3 of GlcNAc in Gal.beta.(1,4)GlcNAc are important for lectin sugar interaction. Our data support the hypothesis that endogenous receptors for galaptin are most likely lactosaminoglycan moieties.