Expression of catabolic pathway of asparagine-linked oligosaccharides on plasma membrane and apoptosis

1. Endo HS去糖基化靶糖蛋白（Target glycoprotein，TGP）的验证和质量标准Endo HS去糖基化靶糖蛋白（Target glycoprotein，TGP）在人口腔附着上皮细胞中的检出率高于脱落上皮细胞，但在白细胞中未检出。用凝胶内去糖基化法鉴定贴壁细胞中分子量为55 K的蛋白质为TGP 55。分子量为20 K、50 K和105 ~ 110 K的蛋白质也分别命名为TGP 20、TGP 50和TGP 105 ~ 110。认为N-末端氨基酸被乙酰化。脱乙酰化TGP 55的N-末端氨基酸序列显示与人细胞角蛋白13的内部氨基酸序列高度同源，但是通过同源性搜索没有观察到具有相同N-末端氨基酸序列的蛋白质。TGP 50的N-末端氨基酸也被封闭。TGP 50的N端和内部氨基酸序列与人I型细胞角蛋白13的氨基酸序列具有高度的同源性，表明TGP 50是I型细胞角蛋白13或与之相似的蛋白质。Endo HS可能参与了上皮细胞的分化。3.抗TGP 55单克隆抗体分析TGP 55的免疫化学性质及细胞定位制备了3株抗TGP 55单克隆抗体。每种抗体均与贴壁和脱壁上皮细胞的TGP 55及其他蛋白质发生反应，且贴壁和脱壁上皮细胞的蛋白质种类不同。上皮细胞的边缘部分被抗体染色，表明TGP 55定位在上皮细胞的边缘部分周围，并且在上皮细胞剥离后定位发生变化。

1.Verification and specification of target glycoproteins deglycosylated by Endo HSTarget glycoprotein(TGP) deglycosylated by Endo HS was detected from attached epithelial cell in the human oral cavity more than detached cell, but not in leukocyte. The protein of molecular weight(MW) of 55K from the attached epithelial cell was specified as TGP55 by in-gel deglycosylation method. The proteins of MW of 20K, 50K and 105〜110K were also specified as TGP20, TGP50 and TGP105〜110.2.Identification of TGP55 and TGP50TGP55 and TGP50 were purified. The N-terminal amino acid was thought to be acetylated. The N-terminal amino acid sequence of deacetylated TGP55 showed high homology with the inner amino acid sequence of human cytokeratin 13, but no proteins having the same N-terminal amino acid sequence was not observed by homology search. The N-terminal amino acid of TGP50 was also blocked. The N-terminal and inner amino acid sequence of TGP50 showed high homology with those of human type I cytokerain 13, indicating that TGP50 is type I cytokeratin 13 or a similar protein to it. Human type I cytokeratin 13 has an asparagine of potential glycosylation site. Endo HS was thought to removing the asparagine-linked oligosaccharide at the site and taking part in the differentiation of the epithelial cell.3.Immunochemical properties and cellular localization of TGP55 analyzed using monoclonal antibodies against TGP55Three monoclonal antibodies against TGP55 were prepared. Each antibody reacted with TGP55 and other proteins of attached and detached epithelial cell, the kinds of proteins were different between attached and detached epithelial cell. The outskirts part of the epithelial cell was stained by the antibodies, indicating that TGP55 localizes around the outskirt part of the epithelial cell and the localization changes after peeling of the epithelial cell.