Rapid and simple preparation of N-linked oligosaccharides by cellulose-column chromatography
Rapid and simple preparation of N-linked oligosaccharides by cellulose-column chromatography
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DOI：
10.1016/s0008-6215(01)00113-6
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发表时间：
2001-06-15
影响因子：
3.1
通讯作者：
Mizuochi, T
中科院分区：
文献类型：
--
作者：
Shimizu, Y;Nakata, M;Mizuochi, T
作为一种从糖蛋白的肼解产物中快速、简便地制备N-连接寡糖的方法，开发了一种使用纤维素柱层析的方法。将含有一系列双触角复合型寡糖的人IgG肼解物上样至用（4：1：1，v/v）1-丁醇-乙醇-水平衡的纤维素柱。用（1：1，v/v）乙醇-水洗脱N-连接寡糖，并通过NPLC结合连续糖苷酶消化进行分析。在用（1：1，v/v）乙醇-水洗脱的馏分中定量回收了含或不含唾液酸的寡糖，而没有来自蛋白质或纤维素的杂质的UV可检测污染。通过用（1：1，v/v）乙醇-水洗脱纤维素柱，也从肼解物中定量制备了其他类型的N-连接寡糖，如铝酸糖蛋白（四触角复合型）、卵清蛋白（混合型）和核糖核酸酶（B大腿甘露糖型），这些寡糖的质量与常规纸色谱法制备的寡糖的质量一样高。(C)2001爱思唯尔科技有限公司版权所有。
As a means of preparing N-linked oligosaccharides from hydrazinolysates of glycoproteins in a rapid and simple manner, a method has been developed using cellulose-column chromatography. Hydrazinolysates of human IgG, containing a series of biantennary complex type oligosaccharides, were applied to a cellulose column equilibrated with (4:1:1, v/v) 1-butanol-ethanol-water. The N-linked oligosaccharides were eluted with (1:1, v/v) ethanol-water, and analyzed by NPLC in combination with sequential glycosidase digestion. The oligosaccharides, with or without sialic acid, were quantitatively recovered in the fraction eluted with (1:1, v/v) ethanol-water without UV-detectable contamination by impurities derived from protein or the cellulose. Other types of N-linked oligosaccharides of al-acid glycoprotein (tetraantennary complex-type), ovalbumin (hybrid-type), and ribonuclease B thigh mannose-type) were also quantitatively prepared from the hydrazinolysates by elution of the cellulose column with (1:1, v/v) ethanol-water and these had as high a quality as those prepared by conventional paper chromatography. (C) 2001 Elsevier Science Ltd. All rights reserved.