Ion Mobility-Mass Spectrometry (IM-MS) for Top-Down Proteomics: Increased Dynamic Range Affords Increased Sequence Coverage
Ion Mobility-Mass Spectrometry (IM-MS) for Top-Down Proteomics: Increased Dynamic Range Affords Increased Sequence Coverage
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DOI：
10.1021/ac300193s
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发表时间：
2012-04-03
影响因子：
7.4
通讯作者：
Russell, David H.
中科院分区：
文献类型：
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作者：
Zinnel, Nathanael F.;Pai, Pei-Jing;Russell, David H.
描述了一种结合了质谱学(MS)、碰撞诱导解离(CID)、离子迁移率(IM)和MS的自上而下蛋白质组学的通用方法，称为MS-CID-IM-MS。利用这种方法，CID产物离子被分散在两个维度上，特别是大小与电荷比(IM)和质量与电荷比(MS)，由此产生的2D数据显示极大地促进了肽/蛋白质质量图谱、氨基酸序列分析和特定部位蛋白质修饰的确定。此外，这种方法缓解了自顶向下蛋白质组学的一些固有限制，即由于大离子系统可用碎片通道的数量以及由此导致的光谱拥堵，片段离子丰度动态范围的限制。对于像蜂毒素(2845 Da)这样的大分子多肽，[M+~(3+)](3+)、[M+4H](4+)和[M+5H](5+)离子的CID得到的氨基酸序列覆盖率分别为42.3%、38.5%和7.7%，而混合MS-CID-IM-MS方法的氨基酸序列覆盖率分别为84.6%、65.4%和69.2%。对于生物大分子，如泛素(8565Da)，氨基酸序列覆盖率从39%增加到76%。MS-CID-IM-MS自上而下的方法允许指定和研究内部碎片离子，从而允许更深入的信息。最后，对泛素的甲酯化和人铁硫簇U(HISCU，14.3 kDa)的单点突变的分析表明，MS-CID-IM-MS能够快速识别修饰的存在和位置。
A general approach that combines mass spectrometry (MS), collision-induced dissociation (CID), ion mobility (IM), and MS for top-down proteomics is described, denoted as MS-CID-IM-MS. Using this approach, CID product ions are dispersed in two dimensions, specifically size-to-charge (IM) and mass-to-charge (MS), and the resulting 2D data display greatly facilitates peptide/protein mass mapping, amino acid sequence analysis, and determination of site-specific protein modifications. Also, this approach alleviates some of the inherent limitations of top-down proteomics, viz, the limitations in dynamic range for fragment ion abundances owing to the number of fragmentation channels available to large ionic systems as well as the resulting spectral congestion. For large peptides such as melittin (2845 Da), CID of the [M + 3H](3+), [M + 4H](4+), and [M + 5H](5+) ions yields amino acid sequence coverage of 42.3%, 38.5%, and 7.7%, respectively, whereas the hybrid MS-CID-IM-MS approach yields amino acid sequence coverages of 84.6%, 65.4%, and 69.2%, respectively. For large biomolecules such as ubiquitin (8565 Da), the amino acid sequence coverage increases from 39% to 76%. The MS-CID-IM-MS top-down approach allows for greater depth of information by allowing the assignment and study of internal fragment ions. Lastly, analysis of the methyl esterification of ubiquitin and single point mutation of human iron sulfur cluster U (HISCU, 14.3 kDa) demonstrates the ability of MS-CID-IM-MS to rapidly identify the presence and sites of modifications.