【摘要】 提纯人血浆运铁蛋白（Tf）,经链霉蛋白酶水解,再经肼解法制备Tf中的二天线N糖链,后者经还原末端的氨基吡啶（PA）化进行荧光标记,再切除外链的唾液酸和半乳糖残基,获得Gn₂Man₃Gn₂-PA荧光标记糖链,以此制备的糖链为受体底物,UDP-GlcNAc为供体底物,用反相HPLC分离底物和产物,建立了N乙耽氨基葡萄糖转移酶V（GnT-V）的测定法。用GnT-V作为肿瘤生化标志物,观察到在二乙基亚硝胺诱发大鼠肝癌的过程中,此酶在诱癌第4周轻度上升,第6周恢复,第10周后持续升高,至18周达正常鼠肝的20倍以上,与病理学上的癌变期相一致。更多还原

【Abstract】 Human transferrin （Tf） was purified from plasms, successively subjected to pronase digestion and hydrazinolysis to release its N-linked sugar chans, which were labeled with fluoresence at the reducing ends by using 2-aminopyridine （PA）. followed by removing the sialic acids and galactoses of the outer chains.The prepared fluorescence labeled sugar chain, Gn₂Man₃Gn₂-PA, formed a single peak on high performance liquid chromatography（HPLC）with a BONDAPAK C18 column and its retention time was equal to the standard sugar chain identified by ¹H-NMR. Using this sugar chain as the acceptor, substrste and UDP-GIcNAc as the donor substrate, an assay for the determination of N-acetylglucosaminyltransferase V（GnTV） was established by means of reverse phase HPLC method to separate substrata and product, which was also identified by the standard sugar chain. GnT-V catalyses the addition of N-acetylglucosamine （GIcNAc） through a β 1-6 bond to α 1-6 linked mannose of the pentasaccharide core and to increase the number of antennary of acceptor N-linked sugar chain. In the course of N nitro-sodiethylamine induced rat hepatocarcinoma, the liver GnT-V activity was slightly increased in 4th week from the beginning of induced carcinogenesis,compatible with the proliferative change in morphology,then decreased to normal level in 6tb week. After 10th week, GnT-V was progressively elevated, reached a level which was higher than 20 times above normal in 18-th week and was consistant with the cancerous change of pathological appearance.更多还原