Qualitative and Quantitative Analysis of Carbohydrate Modification on Glycoproteins from Seeds of Ginkgo biloba
Qualitative and Quantitative Analysis of Carbohydrate Modification on Glycoproteins from Seeds of Ginkgo biloba
复制标题
银杏种子糖蛋白碳水化合物修饰的定性和定量分析。
DOI：
10.1021/acs.jafc.7b01690
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发表时间：
2017-09-06
影响因子：
6.1
通讯作者：
Liu, Li
中科院分区：
文献类型：
--
作者：
Wang, Ting;Hu, Xiao-Chun;Liu, Li
最近的进展，在碳水化合物的交叉反应决定簇（CCD）和过敏反应之间的关系突出了碳水化合物部分在先天免疫系统中的重要性。先前的研究指出，银杏种子中的蛋白质过敏原是糖基化的，与这些蛋白质结合的寡糖也可能导致过敏。本研究的目的是分析来自不同地方的银杏种子糖蛋白的碳水化合物部分，特别是N-连接聚糖的详细结构，以便进一步研究N-聚糖在银杏引起的过敏中所起的作用。单糖组成和免疫印迹分析结果表明存在N-聚糖。通过亲水相互作用超高效液相色谱法（HLLIC-UPLC）和基质辅助激光解吸/电离飞行时间质谱法（MALDI-TOF-MS）进一步对N-聚糖进行详细结构解析。结果表明，在UPLC检测到的16个结构中，有14个结构得到了MALDI-TOF-MS和串联质谱的确证，其中含有刘易斯A决定簇的复合型N-聚糖和高甘露糖型N-聚糖为首次从银杏种子中鉴定。通过使用外标物对N-聚糖进行精确定量，各N-聚糖的绝对量和不同类型N-聚糖的百分比均显示样品间存在显著差异，无任何地理变异模式。
Recent progress in the relationship between carbohydrate cross-reactive determinants (CCDs) and allergic response highlights the importance of carbohydrate moieties in the innate immune system. Previous research pointed out that the protein allergen in Ginkgo biloba seeds is glycosylated, and the oligosaccharides conjugated to these proteins might also contribute to the allergy. The aim of this study was to analyze carbohydrate moieties, especially N-linked glycans, of glycoproteins from Ginkgo seeds originating from different places for detailed structures, to enable further research on the role played by N-glycans in Ginkgo-caused allergy. Results of monosaccharide composition and immunoblotting assays indicated the existence of N-glycans. Detailed structural elucidation of the N-glycans was further carried out by means of hydrophilic interaction ultraperformance liquid chromatography (HLLIC-UPLC) and matrix-assisted laser desorption/ionization time-of flight mass spectrometry (MALDI-TOF-MS). In total, 14 out of 16 structures detected by UPLC were confirmed by MALDI-TOF-MS and tandem mass spectrometry, among which complex-type N-glycans bearing Lewis A determinants and high-mannose-type N-glycans were identified from Ginkgo seeds for the first time. Precise quantification of N-glycans was performed by use of an external standard, and both the absolute amount of each N-glycan and the percentage of different types of N-glycan showed significant diversity among the samples without any pattern of geographic variation.