Advancing Native Mass Spectrometry for Probing Protein Equilibria and Dynamics

在化学系化学测量成像项目的支持下，以及在分子和细胞生物科学系分子生物物理项目的部分共同资助下，华盛顿大学的布什教授和他的小组正在开发新的方法来表征蛋白质及其复合物的平衡和动力学。&在布什实验室开发的最先进的技术使新的测量的相互作用和生物分子在溶液中的结构。这些测量解决了对生物分子和生物材料的快速表征的未满足的需求，包括它们的组装、异质性、质量和相似性。这些测量将有助于回答有关蛋白质和蛋白质复合物的结构和功能的重要问题，这对于理解人类疾病的分子基础和开发治疗方法至关重要。在布什实验室工作的本科生和研究生接受测量科学、数据科学和技术交流方面的培训，为他们在各种各样的职业生涯中取得成功做好准备，从而提高美国的经济竞争力。对于探测溶液中蛋白质平衡和动力学的测量，特别是快速、灵敏和容忍样品异质性的方法，存在未满足的需求。自然离子迁移率（IM）质谱（MS）具有许多属性，使其能够很好地满足这些未满足的需求。然而，对使用溶液和气相测量确定的平衡性质之间的差异的担忧降低了本地IM-MS数据的置信度，并抑制了这些技术的更广泛采用。本研究通过以下方式解决了这些未满足的需求：（1）使用、表征和控制电喷雾电离伴随的电化学诱导样品变化;（2）促进负离子电喷雾电离更广泛地用于天然IM-MS;（3）使用变温电喷雾电离快速表征热曲线和蛋白质折叠可逆的程度。这反过来将使原生IM-MS在生物物理学和结构生物学中发挥更大的作用，这将有助于对生物过程和人类疾病治疗的新知识。本科生研究人员正在开发和制造低成本的仪器，以扩大化学测量和扩大参与研究。该奖项反映了NSF的法定使命，并已被认为是值得通过使用基金会的智力价值和更广泛的影响审查标准进行评估的支持。

With support from the Chemical Measurement & Imaging Program in the Division of Chemistry and partial co-funding from the Molecular Biophysics Program in the Division of Molecular and Cellular Biosciences, Professor Bush and his group at the University of Washington are developing new methods to characterize the equilibria and dynamics of proteins and their complexes. State-of-the-art techniques developed in the Bush lab enable new measurements of the interactions and structures of biological molecules in solution. These measurements address unmet needs for the rapid characterization of biological molecules and biological materials, including their assembly, heterogeneity, quality, and similarity. These measurements will help answer important questions about the structure and function of proteins and protein complexes, important for understanding the molecular basis for human diseases and developing therapeutics. Undergraduate and graduate students working in the Bush lab are trained in measurement science, data science, and technical communication, which prepares them for success in a wide variety of careers that increase the economic competitiveness of the United States.There are unmet needs for measurements that probe protein equilibria and dynamics in solution, especially methods that are fast, sensitive, and tolerant of sample heterogeneity. Native ion mobility (IM) mass spectrometry (MS) has many attributes that make it well positioned to fulfill these unmet needs. However, concerns about differences between equilibrium properties determined using solution- and gas-phase measurements reduces the confidence in native IM-MS data and inhibits the broader adoption of these technologies. This research addresses these unmet needs by (1) using, characterizing, and controlling the electrochemically-induced changes to samples concomitant with electrospray ionization; (2) facilitating the broader adoption of negative ion electrospray ionization for native IM-MS; and (3) using variable-temperature electrospray ionization to rapidly characterize thermal profiles and the extent to which protein folding is reversible. This in turn will enable native IM-MS to play a greater role in biophysics and structural biology, which will contribute to new knowledge of biological processes and cures for human disease. Undergraduate student researchers are developing and fabricating low-cost instruments to expand access to chemical measurements and broaden participation in research.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.