Electron transfer dissociation of N-glycopeptides:: loss of the entire N-glycosylated asparagine side chain
Electron transfer dissociation of N-glycopeptides:: loss of the entire N-glycosylated asparagine side chain
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DOI：
10.1002/rcm.2929
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发表时间：
2007-01-01
影响因子：
2
通讯作者：
Wuhrer, Manfred
中科院分区：
文献类型：
--
作者：
Catalina, M. Isabel;Koeleman, Carolien A. M.;Wuhrer, Manfred
最近引入的电子转移解离（ETD）技术在糖肽水平上为糖蛋白的结构表征开辟了新的可能性。在这份报告中，我们研究了模型糖蛋白辣根过氧化物酶（HRP）的胰蛋白酶N-糖肽的ETD质谱。对通过电喷雾电离获得的多重质子化的N-糖肽进行ETD。通过碰撞诱导解离（CID）（MS 3）进一步分析ETD获得的碎片离子，以确定其明确的结构归属。揭示了以下片段化特征：（1）观察到c型和z型肽骨架裂解，保留了完整聚糖部分，揭示了肽序列、聚糖连接位点和聚糖质量;（2）在较小程度上，糖苷键裂解记录了保留的完整肽序列;（3）确实发生了一系列氨基酸侧链损失。值得注意的是，观察到完整的N-糖基化天冬酰胺侧链的损失。聚糖修饰侧链的这种损失有助于糖肽的结构表征，因为它允许轻松推导和验证聚糖质量和聚糖连接位点处氨基酸残基的性质。重要的是，在本研究中通过在线耦合到射频（rf）四极杆离子阱质谱仪的反相纳米液相色谱（LC）获得了信息性ETD光谱，其中交替采集来自自动选择的前体组的CID和ETD质谱（数据依赖模式）。因此，我们的研究将具有CID和ETD的nano-LC/QIT-MS作为糖肽水平糖蛋白质组学的强大技术。版权所有（c）2007约翰威利父子有限公司。
The recently introduced electron transfer dissociation (ETD) technique opens new possibilities for the structural characterization of glycoproteins at the glycopeptide level. In this report, we investigate the ETD mass spectra of tryptic N-glycopeptides of the model glycoprotein horseradish peroxidase (HRP). Multiply protonated N-glycopeptides obtained by electrospray ionization were subjected to ETD. Fragment ions obtained by ETD were further analyzed by collision-induced dissociation (CID) (MS3) for their unambiguous structural assignment. The following fragmentation features were revealed: (1) c- and z-type peptide backbone cleavages were observed with retention of the intact glycan moiety revealing peptide sequence, glycan attachment site, and glycan mass; (2) to a lesser extent, glycosidic bond cleavages were registered with retention of the intact peptide sequence; and (3) a range of amino acid side chain losses did occur. Remarkably, the loss of the complete N-glycosylated asparagine side chain was observed. This loss of the glycan-modified side chain helps with the structural characterization of glycopeptides by allowing the facile deduction and verification of the glycan mass and the nature of the amino acid residue at the glycan attachment site. Importantly, informative ETD spectra were obtained in this study by reversed-phase nano-liquid chromatography (LC) coupled online to a radio-frequency (rf) quadrupole ion trap QIT) mass spectrometer with alternating acquisition of CID and ETD mass spectra from an automatically selected set of precursors (data-dependent mode). Thus, our study brings nano-LC/QIT-MS" with CID and ETD to the fore as a powerful technique for glycoproteornics at the glycopeptide level. Copyright (c) 2007 John Wiley & Sons, Ltd.