The N-terminal carbohydrate recognition domain of galectin-8 recognizes specific glycosphingolipids with high affinity
The N-terminal carbohydrate recognition domain of galectin-8 recognizes specific glycosphingolipids with high affinity
复制标题
DOI：
10.1093/glycob/cwg094
复制
发表时间：
2003-10-01
期刊：
影响因子：
4.3
通讯作者：
Yamashita, K
中科院分区：
文献类型：
--
作者：
Ideo, H;Seko, A;Yamashita, K
半乳糖凝集素-8是半乳糖凝集素家族的成员，具有两个串联重复的碳水化合物识别结构域（CRD）。我们使用ELISA和表面等离子体共振测定确定了半乳糖凝集素-8及其两个CRD对寡糖和鞘糖脂的结合特异性。半乳糖凝集素-8对3 '-O-硫酸化或3'-O-唾液酸化乳糖和含刘易斯x聚糖的亲和力远高于对终止于Galbeta 1->3/4GlcNAc的寡糖的亲和力。这种特异性主要归因于N-末端CRD（N-结构域），而C-末端CRD（C-结构域）仅对A血型聚糖具有弱亲和力。N-结构域不仅与寡糖结合，还与鞘糖脂结合，包括硫苷脂（SM 4 s）、SM 3、唾液酸化Lc 4Cer、SB 1a、GD 1a、GM 3和唾液酸化nLc 4Cer，表明N-结构域识别3-O-硫酸化或3-O-唾液酸化Gal残基。用硫酸盐取代乳糖或N-乙酰乳糖胺中Gal残基的C-3比用唾液酸取代更有效地增加了半乳糖凝集素-8的识别程度。这是首次证明半乳糖凝集素-8以高亲和力（K（D）类似于10（-8）-10（-9）M）与特异性硫酸化或唾液酸化鞘糖脂结合。当N-结构域的Gln（47）残基转化为Ala（47）时，对硫酸化或唾液酸化聚糖的特异性亲和力选择性丧失，表明该Gln（47）在与Neu 5Acalpha 2->3Gal或SO 3->3Gal残基结合中起重要作用。使用主要表达GM 3的CHO细胞证实了半乳糖凝集素-8与膜结合的GM 3的结合能力。CHO细胞与突变蛋白的结合显著低于与N-结构域的结合。
Galectin-8 is a member of the galectin family and has two tandem repeated carbohydrate recognition domains (CRDs). We determined the binding specificities of galectin-8 and its two CRDs for oligosaccharides and glycosphingolipids using ELISA and surface plasmon resonance assays. Galectin-8 had much higher affinity for 3'-O-sulfated or 3'-O-sialylated lactose and a Lewis x-containing glycan than for oligosaccharides terminating in Galbeta1-->3/4GlcNAc. This specificity was mainly attributed to the N-terminal CRD (N-domain), whereas the C-terminal CRD (C-domain) had only weak affinity for a blood group A glycan. The N-domain bound not only to oligosaccharides but also to glycosphingolipids including sulfatide (SM4 s), SM3, sialyl Lc4Cer, SB1a, GD1a, GM3, and sialyl nLc4Cer, suggesting that the N-domain recognizes a 3-O-sulfated or 3-O-sialylated Gal residue. The substitution of the C-3 of the Gal residue in lactose or N-acetyllactosamine with sulfate increased the degree of recognition by galectin-8 more potently than substitution with sialic acid. This is the first demonstration that galectin-8 binds to specific sulfated or sialylated glycosphingolipids with high affinity (K(D)similar to10(-8)-10(-9) M). When the Gln(47) residue of the N-domain was converted to Ala(47), the specific affinity for sulfated or sialylated glycans was selectively lost, indicating that this Gln(47) plays important roles for binding to Neu5Acalpha2-->3Gal or SO3--->3Gal residues. The binding ability of galectin-8 to membrane-associated GM3 was confirmed using CHO cells, which predominantly express GM3. Binding of CHO cells to the mutein was significantly lower than to the N-domain.