【摘要】 研究人皮肤成纤维细胞纤连蛋白及其片段 Ｎ－糖链结构。用明胶－Ｓｅｐｈａｒｏｓｅ ４Ｂ亲和柱纯化培养的人皮肤成纤维细胞所分泌的纤连蛋白（Ｆｎ），用胰糜蛋白酶限制性水解，获得明胶结合片段和肝素结合片段；用肼化的凝集素与经过碘酸氧化后的辣根过氧化物酶（ＨＲＰ）缩合的方法制备 ４种 ＨＲＰ－凝集素（Ｃｏｎ Ａ， ＤＳＡ，ＷＧＡ，ＬＣＡ）探针，用Ｗｅｓｔｅｒｎ Ｂｌｏｔ法检测明胶结合片段（Ｍｒ４３ × １０３）、肝素结合片段（Ｍｒ３０ ×１０３，２５ ×ｌ０３）在切除唾液酸及平分型 Ｎ－乙酰氨基葡萄糖（ＧＩｃＮＡｃ）前后的 Ｎ－糖链结构。结果： Ｆｎ的 Ｎ－糖链主要存在于 Ｍｒ分别为４３ × １０３、３０ ×１０３、２５ ×１０３的３个片段上，其结构为二、三（主要是Ｃ２Ｃ２６）或四天线的复杂型糖链；糖链末端连有唾液酸（ＳＡ）；都含有核心岩藻糖（ｃ－Ｆｕｃ；平分型 ＧｌｃＮＡｃ只存在于 ４３ × １０３片段所连的糖链中，而且很可能与 Ｃ—Ｆｕｃ共同存在于二天线或Ｃ２Ｃ２６三天线精链上；Ｆｎ的糖链中不含有Ｎ－乙酰乳糖胺重复顺序。结论：在Ｆｎ不同片段上均含有复杂型Ｎ糖链，并存在核心岩藻糖结构，平分型ＧｌｃＮＡｃ只存在于４３ ×１０３片段上。更多还原

【Abstract】 purpose To study the structure of N-glycan of fibronectin from human skin fibroblast and its fragments. METHODS The fibronectin (Fn) secreted by human skin fibroblast was purified from culture medium by gelatin affinity chromatographic column, and was restrictively hydrolyzed by chymotrypsin. Using western blot, we had analysed the structures of N-glycans in Fn fragments before or after exoglycosidases treatment with 4 HRP-lectins (Con A, WGA, DSA, LCA) conjugate probes. Abstract:RESULTS N-glycans of Fn were mainly located in 3 fragmentst agelatin binding fragment with M, (43-45) × 103, and two heparin binding fragments with M, 30 and 25 ×103. All of these 3 fragments contain bi-, tri-, (mainly C2C2,6) or tetraantennary N-glycan with terminal SA and core fucose (c-Fuc), but bisecting GlcNAc is only present in N-glycans of the fragment with M, 43 × 103, and possibly co-exsit with c-Fuc in bi- or tri-antennary C2C2,6 glycans. There is no repeat N-acetyllactosamine sequence in Fn glycans. CONCLUSIONS All of these 3 fragments contain complex N-glycan with terminal SA and Core fucose (c-Fuc), bisecting GlcNAc is only present in N-glycan of the fragment with M, 1 43 × 103.更多还原