Developing High-Resolution Ion Mobility Spectrometry-Charge Detection-Mass Spectrometry for Rapid Analysis in the Megadalton to Gigadalton Regime

项目摘要 该项目将推进一种新的质谱学技术--电荷检测质谱学 (CDMS)用于分析10到100 nm尺寸范围内的大的生物分子组件(即，具有分子 ~1MDA~1GDA的重量)。在这种大小范围内的物种，如病毒和脂蛋白，在 人类健康。然而，它们很难被发现和表征。传统的MS仪器可以 确定许多类型的生物分子的质量和碎裂模式，包括组件；但是， 测量仅限于~1丙二醛以下的物种。在CDMS中，单个离子的质量直接 通过同时测量每个离子的质荷比和电荷来确定。原则证明 使用我们的CDMS原型进行的研究表明，准确的质量可以被确定为丙二醛到GDA 政权。如本提案所示，这是一个有利的进步。然而，我们的原型设计有一个有限的 质量分辨率(我们测得的最大质量分辨率为m/Δm~330)。以及，测量所需的时间 完整的光谱使该仪器不适用于常规分析。该项目描述了进展情况 这将提高CDMS的分辨率和光谱获取速度-每个至少提高一个数量级 规模之大。这将允许对大物种的高分辨率质谱图进行常规记录 时间到了。我们还将开发用于CDMS的离子迁移率光谱(IMS)接口。IMS分离将 提高了整体峰容量，简化了非均相混合物的CDMS分析。在……里面 此外，离子的迁移率取决于其结构，因此提供了补充 来自CDMS的电荷和质量信息。组合的高分辨率IMS-CDMS仪器将提供 分析生物大分子的强有力的新方法。一旦校准和测试，IMS-CDMS 仪器将被用来调查许多对人类健康重要的问题。其中包括：病毒 组装和拆解，表征基因治疗载体，与病毒结合的药物和抗体， 以及脂蛋白亚类的鉴定。这样的测量具有变革的潜力。病毒基因 例如，治疗载体很难描述，因为它们很大，而且因为基因 物质被包含在病毒衣壳中。关键问题，例如衣壳是否包含完整的基因组， 必须解决部分基因组或没有基因组的问题。高分辨率IMS-CDMS测量将提供 关于微小结构缺陷的信息，这些缺陷很难(如果不是不可能)通过其他方法检测到。 同样，IMS-CDMS分析将影响其他领域，如心血管疾病--导致 美国的死亡事件。血浆脂蛋白在动脉粥样硬化的主要潜在原因中起着关键作用。然而， 主要的类别，高密度脂蛋白、低密度脂蛋白和极低密度脂蛋白，都以大小和组成的广泛分布存在。划定 子类的分类将使开发更可靠的诊断测试和更好的疗法成为可能。初步 结果表明，高分辨率IMS-CDMS将能够拆分关键的脂蛋白亚类。

Project Summary This project will advance a new mass spectrometry (MS) technology, charge detection mass spectrometry (CDMS) for analyzing large biomolecular assemblies in the 10 to 100 nm size range (i.e., having molecular weights of ~1 MDa to 1 GDa). Species in this size range, such as viruses and lipoproteins, play critical roles in human health. However, they are difficult to detect and characterize. Conventional MS instruments can determine masses and fragmentation patterns of many types of biomolecules, including assemblies; but, such measurements are limited to species below ~1 MDa. In CDMS, the masses of individual ions are directly determined from simultaneous measurement of each ion’s mass-to-charge ratio and charge. Proof-of-principle studies using our CDMS prototype, show that accurate masses can be determined into the MDa to GDa regime. As shown in this proposal, this is an enabling advance. However, our prototype design has a limited mass resolving power (the maximum we have measured is m/Δm ~ 330). And, the time required for measuring a complete spectrum makes this instrument impractical for routine analyses. This project describes advances that will improve both the CDMS resolving power and spectrum acquisition speed – each by at least an order of magnitude. This will allow high-resolution mass spectra for large species to be routinely recorded for the first time. We will also develop an ion mobility spectrometry (IMS) interface for CDMS. The IMS separation will improve the overall peak capacity and simplify the analysis of heterogeneous mixtures by CDMS analysis. In addition, the mobility of an ion depends on its structure and thus provides information that complements the charge and mass information from CDMS. The combined high-resolution IMS-CDMS instrument will provide a powerful new approach for the analysis of large biomolecules. Once calibrated and tested, the IMS-CDMS instrument will be used to investigate numerous problems important to human health. These include: virus assembly and disassembly, the characterization gene therapy vectors, drug and antibody binding to viruses, and the identification of lipoprotein subclasses. Such measurements have transformative potential. Viral gene therapy vectors, for example, are difficult to characterize because of their large sizes and because the genetic material is contained within virus capsids. Critical issues, such as whether capsids contain the full genome, partial genomes, or no genome, must be addressed. High-resolution IMS-CDMS measurements will provide information about small structural defects, which are difficult (if not impossible) to detect by other means. Similarly, IMS-CDMS analyses will impact other areas, such as cardiovascular diseases - the leading cause of death in the US. Plasma lipoproteins play a key role in the main underlying cause, atherosclerosis. However, the major classes, HDL, LDL, and VLDL, all exist as broad distributions of sizes and compositions. Delineation of subclasses will enable development of more reliable diagnostic tests and better therapeutics. Preliminary results indicate that high-resolution IMS-CDMS will be able to resolve key lipoprotein subclasses.