Enabling electron-induced fragmentation in tandem mass spectrometry

在串联质谱分析中实现电子诱导碎裂

• 批准号: 9346138
• 负责人: Valery G. Voinov
• 金额: $ 22.5万
• 依托单位: E-MSION, INC.
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-09-01 至 2018-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9346138
• 关键词: Acetylation; Address; Adopted; Adoption; Affect; Amides; Biological; Carbohydrates; Cardiovascular Diseases; Cells; Charge; Cleaved cell; Complex; Computer Simulation; Detection; Deuterium; Diagnostic; Disease; Dissociation; Electrons; Electrostatics; Filament; Fourier transform ion cyclotron resonance; Goals; High Pressure Liquid Chromatography; Hydrogen; Industry; Inflammation; Ions; Isotope Labeling; Isotopes; Lipids; Malignant Neoplasms; Manufacturer Name; Mass Spectrum Analysis; Measures; Medical; Methodology; Methods; Modernization; Modification; Molecular; Movement; Nerve Degeneration; Optics; Pattern; Peptide Fragments; Peptides; Pharmaceutical Preparations; Phase; Phosphopeptides; Phosphorylation; Physiologic pulse; Polysaccharides; Post-Translational Protein Processing; Power Sources; Process; Promega; Proteins; Proteomics; Radiation; Reaction Time; Research; Research Personnel; Resolution; Sampling; Small Business Innovation Research Grant; Speed; TRAP Peptide; Technology; Therapeutic Intervention; Time; Tissues; Trypsin; United States National Institutes of Health; Work; base; chemical bond; density; design; disease diagnosis; electron energy; experimental study; improved; instrument; ion mobility; lens; magnetic field; mass spectrometer; millisecond; next generation; programs; research and development; success; tandem mass spectrometry; therapeutic biomarker; tool; transmission process

Summary: The speed, resolution and high mass accuracy of modern mass spectrometers have revolutionized proteomics, but the accurate identification and quantitation of post-translational modifications (PTMs) remain a major challenge—a key limitation for many important medical applications. A key weakness with current mass spectrometry for proteomics lies in the methods used to induce fragmentation, because PTMs such as phosphorylation are among the most labile chemical bonds in proteins and are lost in complex ways by current collision-based fragmentation approaches. An alternative fragmentation methodology called electron capture dissociation (ECD) is well established to produce exceptionally clean spectra that preserve PTMs, but is currently feasible only in expensive FTICR mass spectrometers. The fundamental limitation to ECD is the difficulty of providing enough low-energy electrons to efficiently fragment peptides. We have discovered how to use carefully sculpted magnetic fields with a hot electron-producing filament to restrain large numbers of electrons in the flight path of ions. This can be adapted in any common tandem mass spectrometer without changing the existing ion optics, but our best designs can only fragment 3-5% of doubly charged trypsin-digested peptides—the most common workflow used in mass spectrometry. This low fragmentation efficiency limits sensitivity, which has proved to be the major barrier to adopting this powerful methodology by the mass spectrometry industry. The key focus of this Phase I SBIR project is determining how to increase the interaction time of ions with electrons confined to a narrow beam by the magnetic fields to prove this concept feasible. The reaction time currently is 1-2 microseconds. Our Phase I feasibility question is whether fragmentation can be effectively increased at least two-fold by transiently stopping peptide ions in the ECD cell without significant loss due to electrostatic scattering. In addition, the design must retain the sub-millisecond speed necessary to be compatible for current front-end HPLC and ion mobility separations used with mass spectrometers for complex samples. Rigorous computer simulations show these objectives can be accomplished by carefully cooling precursor ions and then transiently stopping their flight with carefully timed electrical pulses to electrostatic lenses. Proof of feasibility and validated concept demonstration (Phase II) are essential in engaging the major instrument manufacturers to further develop and commercialize our ECD technology for use in their mass spectrometer products. Success will also show how our technology can produce better fragmentation of the most challenging analytes analyzed by mass spectrometry, including lipids, glycans, and other difficult-to- fragment drugs/metabolites. The adoption of our technology will accelerate the ability of many NIH investigators to probe disease mechanisms and identify diagnostic/therapeutic biomarkers with increased accuracy and greater speed, while making fewer mistaken identifications in complex biological samples.

摘要：现代质谱仪的速度，分辨率和高质量精度具有 革命性的蛋白质组学，但对翻译后的准确识别和定量 修改（PTM）仍然是一个主要挑战，这是许多重要的医疗应用的关键限制。 当前质谱蛋白质组学的关键弱点在于用于诱导的方法 碎片化，因为PTM（例如磷酸化）是蛋白质中最不稳定的化学键之一 并通过当前基于碰撞的分裂方法以复杂的方式丢失。另一种 碎裂方法称为电子捕获解离（ECD），可以很好地确定生产 保留PTM的非常干净的光谱，但目前仅在昂贵的FTICR质量中可行 光谱仪。 ECD的基本限制是提供足够低能电子设备的困难 有效地碎片肽。我们发现了如何使用热雕刻的磁场使用 产生电子丝以限制离子飞行路径中的大量电子。这可以 适用于任何常见的串联质谱仪，而无需更改现有的离子光学元件，但我们最好 设计只能片段碎片3-5％的双重胰蛋白酶消化肽，这是最常见的工作流程 用于质谱法。这种低的破碎效率限制了灵敏度，已被证明是 通过质谱行业采用这种强大方法的主要障碍。关键重点 该阶段I SBIR项目正在确定如何增加离子与受限制的电子的相互作用时间 通过磁场到狭窄的光束，以证明这一概念可行。当前的反应时间是1-2 微秒。我们的阶段I可行性问题是至少至少可以有效地增加破碎 通过瞬时停止ECD细胞中的肽离子而没有静电造成的显着损失的两倍 散射。此外，设计必须保留所需的子毫米速度以兼容 当前的前端HPLC和离子迁移率分离与质谱仪一起用于复杂样品。 严格的计算机模拟显示这些目标可以通过仔细冷却前体来实现 离子，然后用精心定时的电脉冲到静电镜头瞬时停止飞行。 可行性证明和经过验证的概念演示（第二阶段）对于参与主要 仪器制造商将进一步开发和商业化我们的ECD技术以供其质量 光谱仪产品。成功还将表明我们的技术如何产生更好的分裂 大多数通过质谱分析的挑战分析物，包括脂质，聚糖和其他难以 - 碎片药物/代谢产物。我们技术的采用将加速许多NIH的能力 研究人员探测疾病机制并鉴定诊断/治疗生物标志物随增加 准确性和更高的速度，同时在复杂的生物样品中识别错误。

项目成果

Enhanced Top-Down Protein Characterization with Electron Capture Dissociation and Cyclic Ion Mobility Spectrometry.

• DOI: 10.1021/acs.analchem.1c04870
• 发表时间: 2022-03-08
• 期刊: Analytical chemistry
• 影响因子: 7.4
• 作者: Shaw JB;Cooper-Shepherd DA;Hewitt D;Wildgoose JL;Beckman JS;Langridge JI;Voinov VG
• 通讯作者: Voinov VG