Development of a Novel Sulfoxide-Containing MS-Cleavable Homobifunctional Cysteine-Reactive Cross-Linker for Studying Protein-Protein Interactions
Development of a Novel Sulfoxide-Containing MS-Cleavable Homobifunctional Cysteine-Reactive Cross-Linker for Studying Protein-Protein Interactions
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DOI：
10.1021/acs.analchem.8b01287
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发表时间：
2018-06-19
影响因子：
7.4
通讯作者：
Huang, Lan
中科院分区：
文献类型：
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作者：
Gutierrez, Craig B.;Block, Sarah A.;Huang, Lan
交联质谱（XL-MS）已成为定义蛋白质-蛋白质相互作用（PPI）和阐明大蛋白质复合物结构的新兴技术。到目前为止，最广泛使用的交联剂靶向赖氨酸。虽然这些试剂已成功地应用于蛋白质组范围内的PPI图谱，但全面的PPI图谱分析将需要额外的交联化学。半胱氨酸是最具反应性的氨基酸之一，由于其在蛋白质结构中的独特作用，是交联的有吸引力的靶标。虽然巯基反应性交联剂是市售的，但它们在XL-MS研究中的应用仍然很少，可能是由于难以识别半胱氨酸交联肽。在此之前，我们开发了一类新的含亚砜的MS可裂解交联剂，以实现使用多级串联质谱法（MSn）快速准确地鉴定交联肽。在这里，我们提出了一种新的亚砜含MS-可切割的同型双功能半胱氨酸反应性交联剂，双马来酰亚胺亚砜（BMSO）的发展。我们表明，BMSO交联肽显示相同的特征碎片模式在碰撞诱导解离（CID）的其他亚砜含MS可裂解的交联肽，从而允许其简化的分析和明确的识别MSn。此外，我们表明，BMSO可以补充胺和酸性残基反应试剂映射蛋白质相互作用区域。总的来说，这项工作不仅扩大了具有不同化学性质的MS可裂解交联剂的工具箱，更重要的是扩大了我们研究PPI的能力。
Cross-linking mass spectrometry (XL-MS) has become an emerging technology for defining protein-protein interactions (PPIs) and elucidating architectures of large protein complexes. Up to now, the most widely used cross-linking reagents target lysines. Although such reagents have been successfully applied to map PPIs at the proteome-wide scale, comprehensive PPI profiling would require additional crosslinking chemistries. Cysteine is one of the most reactive amino acids and an attractive target for cross-linking owing to its unique role in protein structures. Although sulfhydryl-reactive cross-linkers are commercially available, their applications in XL-MS studies remain sparse, likely due to the difficulty in identifying cysteine cross-linked peptides. Previously, we developed a new class of sulfoxide-containing MS-cleavable cross-linkers to enable fast and accurate identification of cross-linked peptides using multistage tandem mass spectrometry (MSn). Here, we present the development of a new sulfoxide-containing MS-cleavable homobifunctional cysteine-reactive cross-linker, bismaleimide sulfoxide (BMSO). We demonstrate that BMSO-cross-linked peptides display the same characteristic fragmentation pattern during collision-induced dissociation (CID) as other sulfoxide-containing MS-cleavable cross-linked peptides, thus permitting their simplified analysis and unambiguous identification by MSn. Additionally, we show that BMSO can complement amine- and acidic-residue-reactive reagents for mapping protein-interaction regions. Collectively, this work not only enlarges the toolbox of MS-cleavable cross-linkers with diverse chemistries, but more importantly expands our capacity and capability of studying PPIs in general.