CAREER: Developing hybrid IMS/OMS-MS/MS techniques for protein complex structure characterization

职业：开发用于蛋白质复合物结构表征的混合 IMS/OMS-MS/MS 技术

• 批准号: 1553021
• 负责人: Stephen Valentine
• 金额: $ 40万
• 依托单位: West Virginia University Research Corporation
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-04-15 至 2022-03-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1553021&HistoricalAwards=false
• 关键词: CAREER; Developing; hybrid; IMS; OMS

With support from the Chemical Measurement and Imaging Program in the Division of Chemistry and the Instrument Development for Biological Research (IDBR) Program in the Division of Biological Infrastructure, Professor Valentine at West Virginia University and his group are developing scientific instrumentation and techniques for studying the three-dimensional shapes of complex, biological molecules. This developmental work is aimed at the study of protein complexes. Proteins play a crucial role in life processes. When such molecules do not form the correct three dimensional shapes, disease often ensues and therefore it becomes important to be able to determine the structures of these biological molecules and how their components come to form critical shapes. However, because of their relatively large sizes and other factors, it is difficult to determine accurate structures for many molecules. The new instrumentation and techniques developed by Professor Valentine are important ways for determining the structures of such proteins. Professor Valentine is also engaged in educational outreach efforts to improve the training of Middle- and High-School teachers in the science, technology, engineering and mathematical fields thereby benefitting many students. The characterization of protein complex structures as well as the structures of coexisting protein species presents one challenge to modern analytical techniques. Professor Valentine is developing novel gas-phase separation instrumentation and techniques to study these problems. This work has three major tasks. First, a hybrid drift tube that combines traditional ion mobility spectrometry (IMS) separations with overtone mobility spectrometry (OMS) analysis is constructed and coupled with an existing linear ion trap mass spectrometer. The approach may allow the probing of minor conformational changes (~4%) occurring on very short timescales (microsecond and lower), enhancing current methods for structural assessments. Second, the novel technique is used to characterize a protein system associated with Huntington's Disease. Multimeric peptide ions formed by portions of the huntingtin (Htt) protein are subjected to OMS-hydrogen deuterium exchange (HDX) analysis with tandem mass spectrometry (MS). The results are compared with those obtained from IMS-OMS-MS analysis to provide a means for testing the new instrumentation and techniques on a real-world sample as well as provide new information in the form of species that are undergoing structural transformations during the course of the measurement. The final task is characterizing these same protein systems using solution-phase HDX combined with OMS-MS/MS studies to unravel linking of solution- and gas-phase structures as well as to provide information regarding conformation establishment in the gas-phase. This research may provide important clues about Huntington's Disease progression. The educational component establishes strategies for improving the efficacy of middle- and high-school science instruction, develops undergraduate curricula, introduces undergraduate students to research, and provides unique research and instructional opportunities for graduate students.

在化学测量和成像计划中的化学分区和生物学研究仪器开发（IDBR）计划的支持下，西弗吉尼亚大学瓦伦丁教授及其小组正在开发科学仪器和研究技术，用于研究复杂分子的三维形状。这项发育工作旨在研究蛋白质复合物。蛋白质在生活过程中起着至关重要的作用。当这样的分子不形成正确的三维形状时，疾病通常会随之而来，因此能够确定这些生物分子的结构以及它们的成分如何形成关键形状变得很重要。但是，由于它们的大小和其他因素相对较大，因此很难确定许多分子的准确结构。 Valentine教授开发的新仪器和技术是确定此类蛋白质结构的重要方法。 Valentine教授还从事教育外展工作，以改善科学，技术，工程和数学领域的中学和高中老师的培训，从而使许多学生受益。蛋白质复合物结构的表征以及共存蛋白质物种的结构对现代分析技术提出了一个挑战。 Valentine教授正在开发新型的气相分离仪器和技术来研究这些问题。这项工作有三个主要任务。首先，构建并与现有的线性离子陷阱质谱仪结合了传统离子迁移率（IMS）分离（OMS）分析（OMS）分析的混合漂移试管。该方法可以允许在非常短的时间尺度（微秒及较低）上探测发生的较小构象变化（约4％），从而增强了当前的结构评估方法。其次，这种新技术用于表征与亨廷顿氏病有关的蛋白质系统。由Huntingtin（HTT）蛋白部分形成的多聚体肽离子通过串联质谱法（MS）进行OMS-HYDREDERED氘交换（HDX）分析。将结果与从IMS-OMS-MS分析获得的结果进行了比较，以提供一种在现实世界样本上测试新仪器和技术的方法，并以在测量过程中正在进行结构转化的物种形式提供新信息。最终任务是使用溶液相HDX与OMS-MS/MS研究结合来揭示溶液和气相结构的联系，并提供有关气相中构象建立的信息。 这项研究可能会提供有关亨廷顿疾病进展的重要线索。 教育组成部分建立了提高中学和高中科学教学功效，开发本科课程，介绍本科生进行研究的策略，并为研究生提供独特的研究和教学机会。