Studies on the glycoproteins expressing neural specific carbohydrate

随着免疫学技术和组织化学方法的进步，神经元特异性糖基化部分被发现。这些携带神经特异性碳水化合物的糖复合物主要在细胞表面观察到，并在神经系统发育过程中以区域或阶段特异性的方式表达。因此，这些神经元特异性碳水化合物部分可能在神经细胞的识别和/或迁移中起重要作用。辣根过氧化物酶（horseradish peroxidase，HRP）抗血清特异性标记蝗虫等昆虫的神经系统，并能识别神经元表面表达的糖蛋白，为了确定糖蛋白表位的结构，我们采用凝集素层析法从蝗虫神经系统中分离表达神经元特异性糖蛋白抗原的糖蛋白。还原条件下SDS-PAGE显示该糖蛋白的表观分子量为35 kDa。通过用α-甘露糖苷酶消化除去α-连接的甘露糖残基完全破坏了35 kDa糖蛋白与抗HRP抗体的结合活性。使用具有脉冲安培检测器的HPLC系统分析35 kDa糖蛋白的碳水化合物组成。35 kDa糖蛋白由Fuc、Man、Gal、Glc、GalNH 2和GlcNH 2组成，分子比分别为1.0、1.4、0.4、1.5、0.4和1.5。

With progress in immunological techniques and histochemical methods, neuron-specific carbohydrate moieties have been discovered. These glycoconjugates carrying the neural specific carbohydrates are mainly observed on the cell surface and expressed in region- or stage-specific manner during the development of the nervous system. Therefore, these neuron- specific carbohydrate moieties may play important roles in recognition and/or migration of neural cells. Antiserum raised against horseradish peroxidase(HRP)specifically labels the nervous system of grasshopper and other insects and has been shown to recognize carbohydrate moiety that is expressed on the surface of neurons.In order to determine the structure of the carbohydrate epitope, we isolated the glycoprotein which express the neuron- specific carbohydrate antigen, from grasshopper nervous system using lectin chromatography. The glycoprotein showed apparent molecular weight of 35 kDa on SDS-PAGE under reducing condition. Removal of alpha-linked mannose residues by digestion with alpha-mannosidase completely destroyed the binding activity of the 35 kDa glycoprotein to the anti-HRP antibody. The carbohydrate composition of the 35 kDa glycoprotein was analyzed using a HPLC system with a pulsed amperometry detector. The 35 kDa glycoprotein was composed of Fuc, Man, Gal, Glc, GalNH2, and GlcNH2 with a molecular ratio of 1.0,1.4, 0.4, 1.5, 0.4, and 1.5, respectively.