Development of an apparatus for rapid release of oligosaccharides at the glycosaminoglycan-protein linkage region in chondroitin sulfate-type proteoglycans
Development of an apparatus for rapid release of oligosaccharides at the glycosaminoglycan-protein linkage region in chondroitin sulfate-type proteoglycans
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DOI：
10.1016/j.ab.2006.12.027
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发表时间：
2007-03-15
影响因子：
2.9
通讯作者：
Kakehi, Kazuaki
中科院分区：
文献类型：
--
作者：
Matsuno, Yu-ki;Yamada, Keita;Kakehi, Kazuaki
开发了一种装置AutoGlycoCutter（AGC），作为在碱性条件下快速释放O-连接型聚糖的工具。该系统允许在蛋白聚糖（PG）中的糖胺聚糖-蛋白质连接区域处快速释放寡糖。用软骨素酶ABC消化PGs后，AGC在3 min内成功地将连接区的寡糖从蛋白质核心中释放出来。用2-氨基苯甲酸标记释放的寡糖的还原端，并结合毛细管电泳（CE）和基质辅助激光解吸飞行时间质谱进行分析。此外，由软骨素酶ABC产生的不饱和二糖来自聚糖的外部部分被标记为2-氨基吖啶酮，并通过CE分析，以确定二糖组成。我们评估了AGC作为一些硫酸软骨素型PG（尿胰蛋白酶抑制剂，牛鼻软骨PG，牛聚集蛋白聚糖，牛核心蛋白聚糖和牛双糖蛋白聚糖）中糖胺聚糖结构分析的方法。对于本研究中检测的所有PG，释放寡糖的回收率为57-73%。特别是，我们强调，使用AGC实现了ca。1000-与常规方法相比，O-聚糖的快速释放增加了一倍。(c)2006年爱思唯尔公司All rights reserved.
An apparatus, AutoGlycoCutter (AGC), was developed as a tool for rapid release of O-linked-type glycans under alkaline conditions. This system allowed rapid release of oligosaccharides at the glycosaminoglycan-protein linkage region in proteoglycans (PGs). After digestion of PGs with chondroitinase ABC, the oligosaccharides at the linkage region were successfully released from the protein core by AGC within 3 min. The reducing ends of the released oligosaccharides were labeled with 2-aminobenzoic acid and analyzed by a combination of capillary electrophoresis (CE) and matrix-assisted laser desorption time-of-flight mass spectrometry. In addition, the unsaturated disaccharides produced by chondroitinase ABC derived from the outer parts of the glycans were labeled with 2-aminoacridone and analyzed by CE to determine the disaccharide compositions. We evaluated AGC as a method for structural analysis of glycosaminoglycans in some chondroitin-sulfate-type PGs (urinary trypsin inhibitor, bovine nasal cartilage PG, bovine aggrecan, bovine decorin, and bovine biglycan). Recoveries of the released oligosaccharides were 57-73% for all PGs tested in the present study. In particular, we emphasize that the use of AGC achieved ca. 1000-fold rapid release of O-glycans compared with the conventional method. (c) 2006 Elsevier Inc. All rights reserved.