Development of Single Particle Analyzer of Mass and Mobility (SPAMM)

单粒子质量和迁移率分析仪（SPAMM）的开发

• 批准号: 8026269
• 负责人: Evan R Williams
• 金额: $ 44.73万
• 依托单位: UNIVERSITY OF CALIFORNIA BERKELEY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-07-01 至 2015-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8026269
• 关键词: Buffers; Cell Mobility; Cell physiology; Charge; Chemistry; Complex; Coupled; Data; Detection; Development; Devices; Dissociation; Electrospray Ionization; Electrostatics; Elements; Fourier Transform; Gases; Helium; Heterogeneity; Individual; Ions; Lifting; Macromolecular Complexes; Mass Spectrum Analysis; Measurement; Measures; Methods; Modification; Molecular; Molecular Weight; Nitrogen; Particle Size; Phase; Photons; Play; Relative (related person); Reporting; Resolution; Role; Sampling; Services; Shapes; Solutions; Structure; System; Techniques; Therapeutic Agents; Time; Travel; Vacuum; austin; base; dalton; density; design; electric field; experience; improved; instrument; instrumentation; ion mobility; ionization; mass analyzer; mass spectrometer; molecular assembly/self assembly; molecular size; nanoscale; particle; pressure; programs; research study; stoichiometry; tandem mass spectrometry; theories

DESCRIPTION (provided by applicant): Mass spectrometry is rapidly becoming a powerful method for the structural elucidation of macromolecular complexes, and can provide complimentary information to conventional biophysical methods. Intact complexes can be lifted out of solution with electrospray ionization, which adds many charges to these molecular assemblies. Experiments have shown that in numerous cases, these gas-phase ions retain many elements of their solution-phase structure. A direct mass measurement provides a rapid measure of the stoichiometry of complexes, even for heterogeneous mixtures. Fragmentation of complexes can yield information about the connectivity of subunits within the complex. Additionally, ion mobility-based approaches are beginning to provide cross-sectional measurements of complexes, making it possible to obtain complementary shape information. A challenge in applying mass spectrometry to even larger complexes is sample heterogeneity, which can result in unresolved charge-state distributions. An additional challenge is that the sensitivity of many mass spectrometers decreases with increasing mass-to-charge ratio (m/z), which makes it more difficult to detect complexes with molecular weights of several mega Daltons and higher with high sensitivity. One aspect of electrospray ionization is that the degree of charging generally increases with molecular size. This makes charge detection mass spectrometry an attractive means of analyzing large macromolecular complexes, because it has the advantage that individual ions possessing multiple charges can be readily detected. Because charge will increase with size, the sensitivity of this technique actually improves with increasing molecular weight. The heterogeneity of the sample does not interfere with mass or mobility measurements, because each ion is analyzed individually. We plan to develop a Single Particle Analyzer of Mass and Mobility (SPAMM), which will incorporate m/z and charge detection measurements so that the masses of individual ions can be rapidly measured. Tandem mass spectrometry capabilities will be incorporated into this dual electrostatic ion trap instrument which will make it possible to measure a fragmentation spectrum of an individual ion of known mass (initial studies will focus on infrared multiple photon dissociation, although other methods could also be incorporated). Finally, ion mobility capabilities will be integrated into this device to make it possible to measure the mass of an ion, and subsequently obtain its absolute collision cross section, which provides information about the shape of the complex. Cross sections can also be obtained for fragments generated by the dissociation of a complex, providing information about the contribution of subunit connectivity to the overall structure of the complex. Because the induced current in this device is directly proportional to the number of charges on the macromolecular complex, sensitivity should increase with increasing particle size making this method applicable too much larger heterogeneous complexes than can be currently analyzed with commercially available mass spectrometers. PUBLIC HEALTH RELEVANCE: Macromolecular complexes play important roles in cellular function, but characterizing the structures of large complexes (tens of mega Daltons and higher) can be challenging owing to heterogeneity that can complicate many conventional methods of analysis. We aim to construct a new instrument that is capable of rapidly measuring the mass, absolute cross section and tandem mass spectra of individual ions of macromolecular complexes at a rate of tens to hundreds of molecules per minute. This would be a high-throuput method to structurally characterize macromolecular complexes, which should aid in understanding their function and make it possible to rapidly screen therapeutic agents that could regulate the activity of macromolecular complexes in a size range not accessible by other mass spectrometry methods.

描述（由申请人提供）：质谱法正在迅速成为阐明大分子复合物结构的有力方法，并且可以为传统生物物理方法提供补充信息。通过电喷雾电离可以将完整的复合物从溶液中分离出来，从而为这些分子组装体添加许多电荷。实验表明，在许多情况下，这些气相离子保留了其溶液相结构的许多元素。直接质量测量可以快速测量复合物的化学计量，甚至对于异质混合物也是如此。复合物的断裂可以产生有关复合物内亚基连接性的信息。此外，基于离子淌度的方法开始提供复合物的横截面测量，使得获得互补的形状信息成为可能。将质谱法应用于更大的复合物的一个挑战是样品异质性，这可能导致无法解析的电荷态分布。另一个挑战是，许多质谱仪的灵敏度随着质荷比 (m/z) 的增加而降低，这使得以高灵敏度检测分子量为数兆道尔顿及更高的复合物变得更加困难。电喷雾电离的一方面是带电程度通常随着分子尺寸的增加而增加。这使得电荷检测质谱成为分析大分子复合物的一种有吸引力的方法，因为它具有可以轻松检测具有多个电荷的单个离子的优点。由于电荷会随着尺寸的增加而增加，因此该技术的灵敏度实际上随着分子量的增加而提高。样品的异质性不会干扰质量或迁移率测量，因为每个离子都是单独分析的。我们计划开发单粒子质量和迁移率分析仪 (SPAMM)，它将结合 m/z 和电荷检测测量，以便可以快速测量单个离子的质量。串联质谱功能将被纳入该双静电离子阱仪器中，这将使得测量已知质量的单个离子的碎片光谱成为可能（初步研究将集中于红外多光子解离，尽管也可以采用其他方法） 。最后，离子淌度功能将集成到该设备中，以便测量离子的质量，并随后获得其绝对碰撞截面，从而提供有关复合物形状的信息。还可以获得复合物解离产生的片段的横截面，提供有关亚基连接对复合物整体结构的贡献的信息。由于该装置中的感应电流与大分子复合物上的电荷数成正比，因此灵敏度应随着颗粒尺寸的增加而增加，使得该方法适用于比目前商用质谱仪分析的大得多的异质复合物。 公共健康相关性：大分子复合物在细胞功能中发挥着重要作用，但表征大型复合物（数十兆道尔顿及更高）的结构可能具有挑战性，因为异质性可能使许多传统分析方法变得复杂。我们的目标是构建一种新仪器，能够以每分钟数十至数百个分子的速度快速测量大分子复合物的单个离子的质量、绝对截面和串联质谱。这将是一种对大分子复合物进行结构表征的高通量方法，这将有助于理解它们的功能，并使得快速筛选能够在其他质谱方法无法达到的尺寸范围内调节大分子复合物活性的治疗剂成为可能。