Chemical Additives Enable Native Mass Spectrometry Measurement of Membrane Protein Oligomeric State within Intact Nanodiscs
Chemical Additives Enable Native Mass Spectrometry Measurement of Membrane Protein Oligomeric State within Intact Nanodiscs
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DOI：
10.1021/jacs.8b11529
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发表时间：
2019-01-16
影响因子：
15
通讯作者：
Marty, Michael T.
中科院分区：
文献类型：
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作者：
Keener, James E.;Zambrano, Dane Evan;Marty, Michael T.
膜蛋白发挥着关键的生化作用，但对其研究仍具有挑战性。近年来，天然或非变性质谱学在表征膜蛋白相互作用方面取得了长足的进步。然而，传统的天然多发性硬化症依赖于洗涤剂胶束，这可能会破坏自然相互作用。脂蛋白纳米盘为天然MS提供了一个在脂质双层环境中呈现膜蛋白的平台，但以往纳米盘中膜蛋白的天然MS受到纳米盘中等稳定性的限制。对于天然多发性硬化症来说，将膜蛋白从纳米盘中排出是困难的，同时也很难保留包含膜蛋白的完整的纳米盘复合体。在这里，我们使用化学试剂来调节在电喷雾电离(ESI)过程中获得的电荷。通过调节ESI条件，我们可以排出结合脂很少的膜蛋白复合体，或者捕获完整的膜蛋白纳米盘复合体-允许在完整的脂类双层环境中测量膜蛋白低聚状态。纳米盘在不同ESI条件下稳定性的显著差异为纳米盘的原生MS开辟了新的应用领域。
Membrane proteins play critical biochemical roles but remain challenging to study. Recently, native or nondenaturing mass spectrometry (MS) has made great strides in characterizing membrane protein interactions. However, conventional native MS relies on detergent micelles, which may disrupt natural interactions. Lipoprotein nanodiscs provide a platform to present membrane proteins for native MS within a lipid bilayer environment, but previous native MS of membrane proteins in nanodiscs has been limited by the intermediate stability of nanodiscs. It is difficult to eject membrane proteins from nanodiscs for native MS but also difficult to retain intact nanodisc complexes with membrane proteins inside. Here, we employed chemical reagents that modulate the charge acquired during electrospray ionization (ESI). By modulating ESI conditions, we could either eject the membrane protein complex with few bound lipids or capture the intact membrane protein nanodisc complex-allowing measurement of the membrane protein oligomeric state within an intact lipid bilayer environment. The dramatic differences in the stability of nanodiscs under different ESI conditions opens new applications for native MS of nanodiscs.