Department of Chemistry and Biochemistry, The Ohio State University, Columbus, Ohio 43210, United States.
Resource for Native MS Guided Structural Biology, The Ohio State University, Columbus, Ohio 43210, United States.
Anal Chem. 2021 Apr 6;93(13):5513-5520. doi: 10.1021/acs.analchem.0c05373. Epub 2021 Mar 22.
Native mass spectrometry (nMS), particularly in conjunction with gas-phase ion mobility spectrometry measurements, has proven useful as a structural biology tool for evaluating the stoichiometry, conformation, and topology of protein complexes. Here, we demonstrate the combination of trapped ion mobility spectrometry (TIMS) and surface-induced dissociation (SID) on a Bruker SolariX XR 15 T FT-ICR mass spectrometer for the structural analysis of protein complexes. We successfully performed SID on mobility-selected protein complexes, including the streptavidin tetramer and cholera toxin B with bound ligands. Additionally, TIMS-SID was employed on a mixture of the peptides desArg1 and desArg9 bradykinin to mobility-separate and identify the individual peptides. Importantly, results show that native-like conformations can be maintained throughout the TIMS analysis. The TIMS-SID spectra are analogous to SID spectra acquired using quadrupole mass selection, indicating little measurable, if any, structural rearrangement during mobility selection. Mobility parking was used on the ion or mobility of interest and 50-200 SID mass spectra were averaged. High-quality TIMS-SID spectra were acquired over a period of 2-10 min, comparable to or slightly longer than SID coupled with ion mobility on various instrument platforms in our laboratory. The ultrahigh resolving power of the 15 T FT-ICR allowed for the identification and relative quantification of overlapping SID fragments with the same nominal / based on isotope patterns, and it shows promise as a platform to probe small mass differences, such as protein/ligand binding or post-translational modifications. These results represent the potential of TIMS-SID-MS for the analysis of both protein complexes and peptides.
天然质谱（nMS），特别是与气相离子淌度谱测量相结合，已被证明是一种有用的结构生物学工具，可用于评估蛋白质复合物的化学计量、构象和拓扑结构。在这里，我们展示了在 Bruker SolariX XR 15 T FT-ICR 傅里叶变换离子回旋共振质谱仪上结合离子淌度谱（TIMS）和表面诱导解离（SID）用于蛋白质复合物结构分析。我们成功地对迁移率选择的蛋白质复合物进行了 SID，包括四聚体链霉亲和素和结合配体的霍乱毒素 B。此外，TIMS-SID 还用于混合肽 desArg1 和 desArg9 缓激肽，以迁移率分离和鉴定各个肽。重要的是，结果表明，在整个 TIMS 分析过程中可以保持天然样构象。TIMS-SID 谱类似于使用四极杆质量选择获得的 SID 谱，表明在迁移率选择过程中几乎没有可测量的（如果有的话）结构重排。在感兴趣的离子或迁移率上使用迁移率停车，并平均 50-200 个 SID 质谱。在 2-10 分钟的时间内获得了高质量的 TIMS-SID 谱，与我们实验室中各种仪器平台上结合离子淌度的 SID 相当或稍长。15 T FT-ICR 的超高分辨率允许根据同位素模式识别和相对定量重叠的 SID 片段，并且有望成为探测小质量差异（如蛋白质/配体结合或翻译后修饰）的平台。这些结果代表了 TIMS-SID-MS 用于分析蛋白质复合物和肽的潜力。
