Expanding the Depth and Sensitivity of Cross-Link Identification by Differential Ion Mobility Using High-Field Asymmetric Waveform Ion Mobility Spectrometry.,Analytical Chemistry

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Expanding the Depth and Sensitivity of Cross-Link Identification by Differential Ion Mobility Using High-Field Asymmetric Waveform Ion Mobility Spectrometry.
Analytical Chemistry ( IF 6.7 ) Pub Date : 2020-07-09 , DOI: 10.1021/acs.analchem.0c01273
Lennart Schnirch ₁ , Michal Nadler-Holly ₁ , Siang-Wun Siao _{1,

2} , Christian K Frese ₂ , Rosa Viner ₃ , Fan Liu ₁

Affiliation

In cross-linking mass spectrometry (XL-MS), the depth and sensitivity of cross-link detection is often limited by the low abundance of cross-links compared to non-cross-linked peptides in the digestion mixture. To improve the identification efficiency of cross-links, here, we present a gas-phase separation strategy using high-field asymmetric waveform ion mobility spectrometry (FAIMS) coupled to the Orbitrap Tribrid mass spectrometers. By enabling an additional peptide separation step in the gas phase using the FAIMS device, we increase the number of cross-link identifications by 22% for a medium complex sample and 59% for strong cation exchange-fractionated HEK293T cell lysate in XL-MS experiments using disuccinimidyl sulfoxide (DSSO) cross-linker. When disuccinimidyl suberate (DSS) cross-linker is in use, we are able to boost cross-link identification by 89% for the medium and 100% for the high complex sample compared to the analyses without FAIMS. Furthermore, we show that, for medium complex samples, FAIMS enables the collection of single-shot XL-MS data with a comparable depth to the corresponding sample fractionated by chromatography-based approaches. Altogether, we demonstrate FAIMS is highly beneficial for XL-MS studies by expanding the proteome coverage of cross-links while improving the efficiency and confidence of cross-link identification.

中文翻译：

使用高场非对称波形离子淌度谱仪通过差分离子淌度扩展交联识别的深度和灵敏度。

在交联质谱法（XL-MS）中，与消解混合物中的非交联肽相比，交联检测的深度和灵敏度通常受交联量低的限制。为了提高交叉链接的识别效率，在这里，我们提出一种气相分离策略，该方法使用高场非对称波形离子迁移谱仪（FAIMS）结合Orbitrap Tribrid质谱仪。通过使用FAIMS设备在气相中进行额外的肽分离步骤，我们在XL-MS实验中将中等复杂样品的交联鉴定数量增加了22％，将强阳离子交换级分的HEK293T细胞裂解物的交联鉴定数量提高了59％使用二琥珀酰亚胺基亚砜（DSSO）交联剂。当使用辛二酸二琥珀酰亚胺酯（DSS）交联剂时，与不使用FAIMS的分析相比，我们对中型样品和高复杂样品的交叉链接识别能力提高了89％。此外，我们表明，对于中等复杂的样品，FAIMS可以收集单次XL-MS数据，其深度与通过基于色谱的方法分离出的相应样品的深度相当。总之，我们证明了FAIMS通过扩大交叉链接的蛋白质组覆盖率，同时提高交叉链接识别的效率和置信度，对XL-MS研究非常有利。FAIMS能够收集单次XL-MS数据，其深度与通过基于色谱的方法分离出的相应样品相当。总之，我们证明了FAIMS通过扩大交叉链接的蛋白质组覆盖率，同时提高交叉链接识别的效率和置信度，对XL-MS研究非常有利。FAIMS可以收集单次XL-MS数据，其深度与通过基于色谱的方法分离出的相应样品相当。总之，我们证明了FAIMS通过扩大交叉链接的蛋白质组覆盖率，同时提高交叉链接识别的效率和置信度，对XL-MS研究非常有利。

更新日期：2020-08-04

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