Structural characterization of the epitopes of the monoclonal antibodies 473HD, CS-56, and MO-225 specific for chondroitin sulfate D-type using the oligosaccharide library
Structural characterization of the epitopes of the monoclonal antibodies 473HD, CS-56, and MO-225 specific for chondroitin sulfate D-type using the oligosaccharide library
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DOI：
10.1093/glycob/cwi036
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发表时间：
2005-06-01
期刊：
影响因子：
4.3
通讯作者：
Sugahara, K
中科院分区：
文献类型：
--
作者：
Ito, Y;Hikino, M;Sugahara, K
在脊椎动物中，硫酸软骨素(CS)/硫酸皮肤素(DS)链的硫酸盐化变化调节中枢神经系统的发育。值得注意的是，二硫化二糖D-单位GlcUA(2-O-硫酸盐)-GalNAc(6-O-硫酸盐)通过嵌入蛋白多糖DSD-1-PG/磷酸聚糖CS部分的DSD-1表位促进轴突生长。单抗473HD抑制DSD-1依赖的神经发生，并识别鲨鱼软骨CS-D，其特征是突出的D-单位，也被另外两个单抗CS-56和MO-225识别。我们用ELISA法和低聚糖芯片技术研究了这些CS-D反应性单抗的寡糖表位结构。CS-56和MO-225识别8-和更大的低聚糖，但后者也结合了一个独特的六糖D-A-D，其中A表示二糖A-单位GlcUA-GalNAc(4-O-硫酸盐)。与CS-56和MO-225反应的八糖共享一个核心A-D四糖，而位于A-D还原和/或非还原侧的相邻结构元件对每个抗体的识别序列都有不同的偏好。相反，473HD与多个六糖或更大的寡糖反应，这些寡糖也含有A-D或D-A四糖序列。与CS-56和MO-225相比，473HD具有明显的特异性，免疫组织学显示473HD表位在小鼠外周器官中表现出不同的表达模式，扩展了先前报道的中枢神经系统限制性表达。经软骨素酶B消化后，DSD-1-PG中的473HD表位被去除，而CS-56和MO-225的表位不被去掉，提示L-艾杜糖酸与473HD表位有密切的结合关系。尽管有这样的补充信息，但完整表位仍有待分离以进行鉴定和全面的分析表征。从而获得了这三种单抗表位中含有A-D四糖核心的糖序列的新信息。
The variation in the sulfation profile of chondroitin sulfate (CS)/dermatan sulfate (DS) chains regulates central nervous system development in vertebrates. Notably, the disulfated disaccharide D-unit, GlcUA(2-O-sulfate)-GalNAc(6-O-sulfate), correlates with the promotion of neurite outgrowth through the DSD-1 epitope that is embedded in the CS moiety of the proteoglycan DSD-1-PG/phosphacan. Monoclonal antibody (mAb) 473HD inhibits the DSD-1-dependent neuritogenesis and also recognizes shark cartilage CS-D, which is characterized by the prominent D-unit and is also recognized by two other mAbs, CS-56 and MO-225. We investigate the oligosaccharide epitope structures of these CS-D-reactive mAbs by ELISA and oligosaccharide microarrays using lipid-derivatized CS oligosaccharides. CS-56 and MO-225 recognized the octa- and larger oligosaccharides, though the latter also bound one unique hexasaccharide D-A-D, where A denotes the disaccharide A-unit GlcUA-GalNAc(4-O-sulfate). The octasaccharides reactive with CS-56 and MO-225 shared a core A-D tetrasaccharide, whereas the neighboring structural elements located on the reducing and/or nonreducing sides of the A-D gave a differential preference additionally to the recognition sequence for each antibody. In contrast, 473HD reacted with multiple hexa- and larger oligosaccharides, which also contained A-D or D-A tetrasaccharide sequences. Consistent with the distinct specificity of 473HD as compared with CS-56 and MO-225, the 473HD epitope displayed a different expression pattern in peripheral mouse organs as revealed by immunohistology, extending the previously reported CNS-restricted expression. The epitope of 473HD, but not of CS-56 or MO-225, was eliminated from DSD-1-PG by digestion with chondroitinase B, suggesting the close association of L-iduronic acid with the 473HD epitope. Despite such supplemental information, the integral epitope remains to be isolated for identification and comprehensive analytical characterisation. Thus novel information on the sugar sequences containing the A-D tetrasaccharide core was obtained for the epitopes of these three useful mAbs.