Trapped Ion Mobility Spectrometry Reduces Spectral Complexity in Mass Spectrometry-Based Proteomics
Trapped Ion Mobility Spectrometry Reduces Spectral Complexity in Mass Spectrometry-Based Proteomics
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DOI：
10.1021/acs.analchem.1c01399
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发表时间：
2021-12-09
影响因子：
7.4
通讯作者：
Rost, Hannes L.
中科院分区：
文献类型：
--
作者：
Charkow, Joshua;Rost, Hannes L.
在自下而上的质谱蛋白质组学中，深度蛋白质组覆盖率受到高共片段化率的限制。当四极杆分离出一种以上的分析物时，发生共碎裂，随后的碎裂事件产生异质来源的碎片离子。降低共片段化速率的一种策略是通过有效的肽分离技术，例如色谱分离和最近流行的离子迁移率（IM）光谱法，其通过肽的碰撞横截面分离肽。在这里，我们使用一个计算模型来调查的能力，捕获的IM光谱（TIMS）设备在有效地分离肽离子和量化的TIMS设备的分离功率的上下文中的并行累积-串行碎片（PASEF）的工作流程。我们发现，TIMS分离增加了无干扰的MS 1肽特征的数量9.2倍，同时降低了前体光谱中的平均肽密度6.5倍。在数据依赖性采集PASEF工作流程中，IM分离将无共碎片的光谱数量增加了4.1倍，高质量光谱数量增加了17倍。使用分类模型，我们估计，这种观察到的光谱复杂性的降低导致肽光谱匹配的可能性增加，这可能会提高肽识别率。在数据独立采集工作流程的背景下，估计IM分离导致的光谱复杂性降低相当于隔离窗口宽度降低4倍（从25 Da降低到6.5 Da）。我们的研究表明，TIMS分离通过降低共片段化率来降低光谱复杂性，这表明TIMS分离可能有助于在PASEF工作流程中观察到的高识别率。
In bottom-up mass spectrometry-based proteomics, deep proteome coverage is limited by high cofragmentation rates. Cofragmentation occurs when more than one analyte is isolated by the quadrupole and the subsequent fragmentation event produces fragment ions of heterogeneous origin. One strategy to reduce cofragmentation rates is through effective peptide separation techniques such as chromatographic separation and, the more recently popularized, ion mobility (IM) spectrometry, which separates peptides by their collisional cross section. Here, we use a computational model to investigate the capability of the trapped IM spectrometry (TIMS) device at effectively separating peptide ions and quantify the separation power of the TIMS device in the context of a parallel accumulation-serial fragmentation (PASEF) workflow. We found that TIMS separation increases the number of interference-free MS1 peptide features 9.2-fold, while decreasing the average peptide density in precursor spectra 6.5-fold. In a data-dependent acquisition PASEF workflow, IM separation increases the number of spectra without cofragmentation by a factor of 4.1 and the number of high-quality spectra 17-fold. Using a categorical model, we estimate that this observed decrease in spectral complexity results in an increased likelihood for peptide spectral matches, which may improve peptide identification rates. In the context of a data-independent acquisition workflow, the reduction in spectral complexity resulting from IM separation is estimated to be equivalent to a 4-fold decrease in the isolation window width (from 25 to 6.5 Da). Our study demonstrates that TIMS separation decreases spectral complexity by reducing cofragmentation rates, suggesting that TIMS separation may contribute toward the high identification rates observed in PASEF workflows.