Jiao Jenny, Zhang Hailong, Reinhold Vernon N
The Glycomics Center, Department of Chemistry, University of New Hampshire, Durham, NH 03824.
Int J Mass Spectrom. 2011 Jun 1;303(2-3):109-117. doi: 10.1016/j.ijms.2011.01.016.
This report outlines and applies a high performance sequencing technology to evaluate the glycome of a common model glycoprotein, ovalbumin. The targets were the N-linked glycans enzymatically released from the protein, the N-glycoproteome. These product glycans were reduced, methylated and directly infused into the MS using a chip-based nanoelectrospray with the ions structurally characterized by sequential disassembly. Ten major ions were selected for detailed analysis. Isomer topologies (glycan connectivity) were determined from ion pathways of disassembly. Linkage information was revealed by specific cross-ring cleavage fragments within smaller oligomers. Both connectivity and linkage features were assisted with described bioinformatic tools and details confirmed with a standards library of fragments. The number of isomeric structures found within these 10 parent ions were 37, more than double earlier reports, and setting a new goal for developing technology. In this non-chromatographic, high performance spatial approach, the focus has been patterned to be comprehensive, and stay within the bounds of a plausible high throughput strategy consistent with automation. Selective structures are described in the text to appraise readers of the general approach; a more comprehensive coverage has been included in supplemental material.
本报告概述并应用了一种高性能测序技术来评估一种常见的模型糖蛋白——卵清蛋白的聚糖组。研究对象是从该蛋白上酶解释放出的N-连接聚糖，即N-糖蛋白组。这些产物聚糖经过还原、甲基化处理，然后使用基于芯片的纳米电喷雾直接注入质谱仪，通过顺序拆解对离子进行结构表征。选择了10个主要离子进行详细分析。从拆解的离子途径确定异构体拓扑结构（聚糖连接性）。通过较小寡聚体中的特定交叉环裂解片段揭示连接信息。连接性和连接特征均借助所述生物信息学工具辅助分析，并用片段标准库确认细节。在这10个母离子中发现的异构体结构数量为37个，比早期报告多出一倍以上，为技术发展设定了新目标。在这种非色谱的高性能空间方法中，重点已设定为全面性，并保持在与自动化一致的合理高通量策略范围内。文中描述了选择性结构，以使读者了解一般方法；补充材料中包含了更全面的内容。
