Improved Peptide Identification by Targeted Fragmentation Using CID, HCD and ETD on an LTQ-Orbitrap Velos
Improved Peptide Identification by Targeted Fragmentation Using CID, HCD and ETD on an LTQ-Orbitrap Velos
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DOI：
10.1021/pr1011729
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发表时间：
2011-05-01
影响因子：
4.4
通讯作者：
Mohammed, Shabaz
中科院分区：
文献类型：
--
作者：
Frese, Christian K.;Altelaar, A. F. Maarten;Mohammed, Shabaz
在过去的十年中，通过碰撞诱导解离（CID）进行肽测序已成为基于质谱的蛋白质组学的首选方法。电子转移解离（ETD）等替代裂解技术的发展扩展了串联质谱的可能性。仪器的最新进展允许以高速度和灵敏度检测肽碎片离子（例如，在2D或3D离子阱中）或以高分辨率和高质量精度（例如，Orbitrap或ToF）。在这里，我们描述了一个全面的实验比较使用ETD，离子阱CID，和光束型CID（HCD）结合线性离子阱或Orbitrap读出的胰蛋白酶肽的大规模分析。我们研究了片段化技术和质量分析仪的组合提供了最佳性能的分析不同的肽群体，如N-乙酰化，磷酸化和胰蛋白酶肽多达两个错过裂解。我们发现HCD比CID和ETD对双电荷肽提供更多的肽识别。在Mascot得分方面，ETD FT优于其他技术，用于电荷状态高于2的肽。我们的数据表明，使用轨道阱碎片离子检测时，光谱质量和速度之间有一个权衡。我们的结论是，决策树调节的高能碰撞解离（HCD）和ETD的组合可以提高平均Mascot分数。
Over the past decade peptide sequencing by collision induced dissociation (CID) has become the method of choice in mass spectrometry-based proteomics. The development of alternative fragmentation techniques such as electron transfer dissociation (ETD) has extended the possibilities within tandem mass spectrometry. Recent advances in instrumentation allow peptide fragment ions to be detected with high speed and sensitivity (e.g., in a 2D or 3D ion trap) or at high resolution and high mass accuracy (e.g., an Orbitrap or a ToF). Here, we describe a comprehensive experimental comparison of using ETD, ion-trap CID, and beam type CID (HCD) in combination with either linear ion trap or Orbitrap readout for the large-scale analysis of tryptic peptides. We investigate which combination of fragmentation technique and mass analyzer provides the best performance for the analysis of distinct peptide populations such as N-acetylated, phosphorylated, and tryptic peptides with up to two missed cleavages. We found that HCD provides more peptide identifications than CID and ETD for doubly charged peptides. In terms of Mascot score, ETD FT outperforms the other techniques for peptides with charge states higher than 2. Our data shows that there is a trade-off between spectral quality and speed when using the Orbitrap for fragment ion detection. We conclude that a decision-tree regulated combination of higher-energy collisional dissociation (HCD) and ETD can improve the average Mascot score.