Isolation and characterization of the Glc3Man9GlcNAc2 from lipid-linked oligosaccharides of plants.
Isolation and characterization of the Glc3Man9GlcNAc2 from lipid-linked oligosaccharides of plants.
复制标题
从植物脂质连接寡糖中分离和表征 Glc3Man9GlcNAc2。
DOI：
10.1016/0003-9861(82)90273-9
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发表时间：
1982
影响因子：
3.9
通讯作者：
Elbein,AD
中科院分区：
文献类型：
--
作者：
Hori,H;JamesJr,DW;Elbein,AD
从在 [2-3H] 甘露糖或 [2-3H] 葡萄糖存在下孵育的悬浮培养大豆细胞中分离出脂质连接的寡糖。在 DEAE-纤维素上纯化脂质连接的寡糖后，通过弱酸水解释放寡糖，并通过 Bio-Gel P-4 柱上的凝胶过滤分离。用[3H]甘露糖或[3H]葡萄糖标记的主要寡糖与Glc3Man9GlcNAc2的真实样品共迁移，通过在Bio-Gel P-4上重复层析将其纯化至均质。该寡糖的特点是对多种酶处理（即氨基葡萄糖苷酶 H、α-甘露糖苷酶、含有加工葡萄糖苷酶的肝膜制剂）和所得产品的分析敏感。它还通过甲基化分析和所得甲基化糖的鉴定来表征。因此，[3H]甘露糖标记的寡糖的甲基化产生2,3,4,6-四甲基甘露糖、3,4,6-三甲基甘露糖和2,4-二甲基甘露糖，以及痕量的2,4,6-三甲基甘露糖。 [3H]葡萄糖标记的寡糖的甲基化产生几乎等量的2,3,4,6-四甲基葡萄糖、3,4,6-三甲基葡萄糖和2,4,6-三甲基葡萄糖。这些数据表明植物 Glc3Man9GlcNAc2 可能与动物 Glc3Man9GlcNAc2 相似（即使不相同）。当孵育混合物中包含衣霉素时，该寡糖的生物合成受到抑制。
Lipid-linked oligosaccharides were isolated from suspension-cultured soybean cells incubated in the presence of [2-3H]mannose or [2-3H]glucose. After purification of the lipid-linked oligosaccharides on DEAE-cellulose, the oligosaccharides were released by mild acid hydrolysis and were isolated by gel filtration on columns of Bio-Gel P-4. The major oligosaccharide, labeled with either [3H]mannose or [3H]glucose, that comigrated with an authentic sample of Glc3Man9GlcNAc2was purified to homogeneity by repeated chromatography on Bio-Gel P-4. This oligosaccharide was characterized by its susceptibility to a variety of enzymatic treatments (i.e., endoglucosaminidase H, α-mannosidase, a liver membrane preparation containing the processing glucosidases) and analysis of the resulting products. It was also characterized by methylation analysis and identification of the resulting methylated sugars. Thus, methylation of the [3H]mannose-labeled oligosaccharide gave rise to 2,3,4,6-tetramethylmannose, 3,4,6-trimethylmannose, and 2,4-dimethylmannose, as well as a trace amount of 2,4,6-trimethylmannose. Methylation of the [3H]glucose-labeled oligosaccharide yielded 2,3,4,6-tetramethylglucose, 3,4,6-trimethylglucose, and 2,4,6-trimethylglucose in almost equal amounts. These data suggest that the plant Glc3Man9GlcNAc2is probably similar, if not identical, to the animal Glc3Man9GlcNAc2. The biosynthesis of this oligosaccharide was inhibited when tunicamycin was included in the incubation mixtures.