Gas-Phase Cross-Linking with Ion/Ion Chemistry Coupled to Ion Mobility/Mass Spectrometry

离子/离子化学气相交联与离子淌度/质谱联用

基本信息

批准号：
9807489
负责人：
Ian Webb
金额：
$ 20.7万
依托单位：
INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-08-01 至 2021-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9807489
关键词：
Address Amino Acids Amyloid Binding Sites Biological Process Biomedical Research Cell Mobility Cells Chemistry Clinical Research Clinical Treatment Complex Mixtures Coupled Coupling Cryoelectron Microscopy Crystallization Data Analyses Dimensions Disease Dissociation Drug Design Electrons Gases Generations Goals Health Heterogeneity Human Ions Knowledge Label Lead Location Malignant Neoplasms Mass Spectrum Analysis Measurement Measures Methods Mission Modification Molecular Conformation Molecular Weight Nature Nuclear Magnetic Resonance Organism Outcome Parkinson Disease Peptides Pharmaceutical Preparations Phase Phosphorylation Post-Translational Protein Processing Production Protein Analysis Protein Dynamics Proteins Public Health Quaternary Protein Structure Reaction Reaction Time Reagent Research Research Design Research Project Grants Resolution Roentgen Rays Sampling Shapes Signal Transduction Site Speed Structural Protein Structure System Techniques Technology United States National Institutes of Health alpha synuclein crosslink drug candidate drug development experimental study field study flexibility innovation ion mobility link protein mass spectrometer millisecond new technology novel preservation protein complex protein folding protein protein interaction protein structure protein structure function screening small molecule stoichiometry tandem mass spectrometry technology development therapy design tool transcription factor virtual

Address Amino Acids Amyloid Binding Sites Biological Process Biomedical Research Cell Mobility Cells Chemistry Clinical Research Clinical Treatment Complex Mixtures Coupled Coupling Cryoelectron Microscopy Crystallization Data Analyses Dimensions Disease Dissociation Drug Design Electrons Gases Generations Goals Health Heterogeneity Human Ions Knowledge Label Lead Location Malignant Neoplasms Mass Spectrum Analysis Measurement Measures Methods Mission Modification Molecular Conformation Molecular Weight Nature Nuclear Magnetic Resonance Organism Outcome Parkinson Disease Peptides Pharmaceutical Preparations Phase Phosphorylation Post-Translational Protein Processing Production Protein Analysis Protein Dynamics Proteins Public Health Quaternary Protein Structure Reaction Reaction Time Reagent Research Research Design Research Project Grants Resolution Roentgen Rays Sampling Shapes Signal Transduction Site Speed Structural Protein Structure System Techniques Technology United States National Institutes of Health alpha synuclein crosslink drug candidate drug development experimental study field study flexibility innovation ion mobility link protein mass spectrometer millisecond new technology novel preservation protein complex protein folding protein protein interaction protein structure protein structure function screening small molecule stoichiometry tandem mass spectrometry technology development therapy design tool transcription factor virtual

展开

项目摘要

PROJECT SUMMARY Although many technologies exist for studying protein structures, none possess a combination of speed, selectivity, accuracy, resolution, and flexibility. The long-term goal is to use a suite of gas-phase chemistries for ion/ion reactions coupled to ion mobility (IM) and tandem mass spectrometry (MS) measurements for high- throughput, virtually sample-prep free, protein structure measurements. The overall objective for this exploratory project, which is the next step toward attaining our long-term goal, is to implement rapid ion/ion cross-linking reactions on an IM/MS platform, using tandem mass spectrometry to identify cross-linked sites. The rationale for the development of this technology is to combine the information from native IM/MS with information obtained from cross-linking in an experimental method conducted on the sub-second timescale. The increase in throughput, information, and lack of sample prep (compared to, e.g., X-ray diffractometry, nuclear magnetic resonance, or cryo-electron microscopy) is expected to advance biomedical research determining protein structure and function to where protein structural determinations can become rapid and routine. The overall objective of this application will be reached through the following Specific Aims: 1. Combine gas-phase ion/ion cross-linking of intact proteins with IM/MS measurements; and 2. Use IM combined with tandem MS to determine cross-linking locations for intact proteins. For the first aim, a variety of monomeric and multimeric proteins will be cross-linked in the gas-phase. Changes in overall structure between cross-linked and unmodified proteins, as well as between solution and gas-phase cross-linked proteins, will be measured by IM. Under the second aim, a combination of collision induced dissociation (CID) and electron capture dissociation (ECD) will be used to determine cross-linked sites. The proposed technology is innovative because it represents a substantive departure from the status quo by coupling cross-linking and native IM/MS analysis into one gas-phase mass spectrometry experiment, allowing rapid cross-linking analysis and providing multiple complementary measures of gas-phase protein structure. This new technology is significant because it is expected to become a rapid tool for high-throughput characterization of primary, secondary, tertiary, and quaternary protein structure. When fully developed, the technology has the potential to be used for complex mixtures of intact proteins and for rapid screening of interactions with small molecules/drugs, creating new opportunities in clinical research, treatment, and drug design.

项目摘要尽管存在许多用于研究蛋白质结构的技术，但没有一个速度组合选择性，准确性，分辨率和灵活性。长期目标是使用一套气相化学套件进行离子/离子反应与离子迁移率（IM）和串联质谱法（MS）测量相关的离子/离子反应。吞吐量，几乎没有样品-PREP的蛋白质结构测量。探索性的总体目标项目是实现我们的长期目标的下一步，是实施快速离子/离子交联 IM/MS平台上的反应，使用串联质谱法识别交联位点。理由该技术的开发是将来自本机IM/MS的信息与获得的信息相结合从在亚秒时间尺度上进行的实验方法的交联。增加吞吐量，信息和样品准备不足（与X射线衍射法相比，核磁性共振或冷冻电子显微镜）有望推进生物医学研究确定蛋白质蛋白质结构测定可以成为迅速和常规的结构和功能。总体该应用的目的将通过以下特定目的达到：1。结合气相/离子离子与IM/MS测量值的完整蛋白质交联；和2。使用IM与串联MS合并来确定完整蛋白质的交联位置。对于第一个目标，各种单体和多聚蛋白将是在气相中交联。交联蛋白质和未修饰的蛋白质之间的总体结构变化，AS 以及在溶液和气相交联蛋白之间，将通过IM测量。在第二个目标下，碰撞诱导解离（CID）和电子捕获解离（ECD）的组合将用于确定交联位点。提出的技术具有创新性，因为它代表了实质性通过将交联和本机IM/MS分析耦合到一个气相质量，从现状偏离现状光谱实验，允许快速交联分析并提供多种互补措施气相蛋白结构。这项新技术很重要，因为它有望成为快速工具用于高通量，次级，第三和第四纪蛋白结构的高通量表征。完全该技术开发的潜力有可能用于完整蛋白质的复杂混合物，并用于快速筛选与小分子/药物的相互作用，在临床研究，治疗中创造了新的机会，和药物设计。