Signature-Ion-Triggered Mass Spectrometry Approach Enabled Discovery of N- and O-Linked Glycosylated Neuropeptides in the Crustacean Nervous System
Signature-Ion-Triggered Mass Spectrometry Approach Enabled Discovery of N- and O-Linked Glycosylated Neuropeptides in the Crustacean Nervous System
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DOI：
10.1021/acs.jproteome.9b00525
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发表时间：
2020-02-01
影响因子：
4.4
通讯作者：
Li, Lingjun
中科院分区：
文献类型：
--
作者：
Cao, Qinjingwen;Yu, Qing;Li, Lingjun
甲壳动物是研究神经调节的常用模式生物。尽管甲壳类神经肽家族及其功能已有众多报道，但关于神经肽糖基化的研究尚未见报道。这在一定程度上是因为缺乏能够破译这种复杂的低丰度翻译后修饰的敏感方法，尽管糖基化已被证明在神经调节中发挥重要作用。在这里，我们描述了利用高能碰撞解离(HCD)MS/MS光谱中产生的特征氧离子的无浓缩方法发现糖基化神经肽。在HCD扫描中检测到的氧离子表明，多糖与多肽相连，允许进行电子转移/高能碰撞解离(EThcD)，以选择性地阐明埋藏在非糖基化多肽中的糖基化神经肽的结构信息。总体而言，在甲壳类神经系统中首次发现了4个N-连接和14个O-连接的糖基化神经肽。此外，基于收集的HCD扫描，还发现了91个新的推定神经肽。这种混合方法结合了神经肽发现的鸟枪法和糖基化鉴定的靶向策略，使甲壳类中糖基化神经肽的第一次报告成为可能，同时发现了更多的神经肽。新的糖基化神经肽的阐明为甲壳动物多肽组的研究提供了新的线索，并为未来的神经肽功能研究提供了新的见解。
Crustaceans are commonly used model organisms to study neuromodulation. Despite numerous reported crustacean neuropeptide families and their functions, there has been no report on neuropeptide glycosylation. This is in part due to a lack of sensitive methods that enable deciphering this intricate low-abundance post-translational modification, even though glycosylation has been shown to play an important role in neuromodulation. Here, we describe the discovery of glycosylated neuropeptides with an enrichment-free approach, taking advantage of signature oxonium ions produced in higher-energy collision dissociation (HCD) MS/MS spectra. The detection of the oxonium ions in the HCD scans suggests glycan attachment to peptides, allowing electron-transfer/higher-energy collision dissociation (EThcD) to be performed to selectively elucidate structural information of glycosylated neuropeptides that are buried in nonglycosylated peptides. Overall, 4 N-linked and 14 O-linked glycosylated neuropeptides have been identified for the first time in the crustacean nervous system. In addition, 91 novel putative neuropeptides have been discovered based on the collected HCD scans. This hybrid approach, coupling a shotgun method for neuropeptide discovery and targeted strategy for glycosylation characterization, enables the first report on glycosylated neuropeptides in crustaceans and the discovery of additional neuropeptides simultaneously. The elucidation of novel glycosylated neuropeptides sheds light on the crustacean peptidome and offers novel insights into future neuropeptide functional studies.