Abstract：
Mass-spectrometry based assays in structural biology studies measure either intact or digested proteins. Typically, different mass spectrometers are dedicated for such measurements: those optimized for rapid analysis of peptides or those designed for high molecular weight analysis. A commercial trapped ion mobility-quadrupole-time-of-flight (TIMS-Q-TOF) platform is widely utilized for proteomics and metabolomics, with ion mobility providing a separation dimension in addition to liquid chromatography. The ability to perform high-quality native mass spectrometry of protein complexes, however, remains largely uninvestigated. Here, we evaluate a commercial TIMS-Q-TOF platform for analyzing noncovalent protein complexes by utilizing the instrument\'s full range of ion mobility, MS, and MS/MS (both in-source activation and collision cell CID) capabilities. The TIMS analyzer is able to be tuned gently to yield collision cross sections of native-like complexes comparable to those previously reported on various instrument platforms. In-source activation and collision cell CID were robust for both small and large complexes. TIMS-CID was performed on protein complexes streptavidin (53 kDa), avidin (68 kDa), and cholera toxin B (CTB, 58 kDa). Complexes pyruvate kinase (237 kDa) and GroEL (801 kDa) were beyond the trapping capabilities of the commercial TIMS analyzer, but TOF mass spectra could be acquired. The presented results indicate that the commercial TIMS-Q-TOF platform can be used for both omics and native mass spectrometry applications; however, modifications to the commercial RF drivers for both the TIMS analyzer and quadrupole (currently limited to m/z 3000) are necessary to mobility analyze protein complexes greater than about 60 kDa.
摘要：
结构生物学研究中基于质谱的测定测量完整或消化的蛋白质。通常,不同的质谱仪专用于这样的测量：那些优化的快速分析的肽或那些设计用于高分子量分析。商业捕获离子迁移率-四极杆飞行时间（TIMS-Q-TOF）平台广泛用于蛋白质组学和代谢组学，离子迁移率提供分离尺寸除了液相色谱。能够对蛋白质复合物进行高质量的天然质谱分析，然而,基本上没有被调查。这里,我们评估了一个商业的TIMS-Q-TOF平台，用于分析非共价蛋白质复合物通过利用仪器的全范围的离子迁移率，MS,和MS/MS(源中激活和碰撞小区CID)功能。TIMS分析仪能够轻轻调整，以产生与以前在各种仪器平台上报道的类似天然复合物的碰撞截面。源中激活和碰撞细胞CID对于小型和大型复合物都是稳健的。TIMS-CID在蛋白复合物链霉亲和素（53kDa）上进行，抗生物素蛋白(68kDa),和霍乱毒素B（CTB，58kDa）。配合物丙酮酸激酶（237kDa）和GroEL（801kDa）超出了商业TIMS分析仪的捕获能力，但是可以获得TOF质谱。所呈现的结果表明，商业TIMS-Q-TOF平台可用于组学和天然质谱应用；然而，对TIMS分析仪和四极杆（目前限制为m/z3000）的商用RF驱动器的修改对于迁移率分析大于约60kDa的蛋白质复合物是必要的。
