IDBR: Type A. High-Performance Surface-induced Dissociation MS/MS

俄亥俄州立大学被授予开发和仪器，通过改进质谱仪中蛋白质复合体的解离来表征蛋白质复合体。生命系统中的蛋白质可以扮演结构性角色，比如皮肤和头发中的蛋白质；它们也可以扮演活跃的角色，比如帮助消化的酶，或者携带氧气和二氧化碳通过身体的血红蛋白。新陈代谢、细胞信号、基因表达、运输、细胞周期调控和亚细胞结构的形成等细胞过程依赖于蛋白质复合体/组合体的形成和动态相互作用。这些蛋白质复合体也可能与其他生物分子(DNA/RNA)结合，以特定的方式相互作用。由于蛋白质结构与功能密切相关，因此需要有效的工具来表征蛋白质-蛋白质和蛋白质-配体复合体。这项拟议的研究将导致将表面诱导解离装置集成到多种类型的质谱仪中，这些质谱仪将提供生物学上重要的蛋白质复合体的结构信息。该项目的发现将在大学课程和国际、国家、区域和地方关于质谱学和生物分子结构的研讨会/会议上展示，并在同行评议的文献中展示。本科生和研究生以及博士后助理将接受仪器开发方面的培训，还将通过与私营部门合作伙伴以及当地电子和机械车间的互动获得合作技能。早期的SID采用者将作为修改成功的直接探索者。该项目是供应商和平台中立的，直接包括多个私营部门合作伙伴，Ardara Technologies、Waters(Q-IM-TOF，四极离子迁移率飞行时间)、Thermo(Orbitrap)和Bruker(ICR，离子回旋共振)，为项目团队提供了在公司用户会议上发言的机会，并为这些合作伙伴评估仪器改进的商业价值提供了机会。这项研究将开发高/超高分辨率质谱仪中的表面诱导解离(SID)，并改进其在离子迁移率(IM)质谱仪中的应用，该质谱仪的目的是表征蛋白质络合物。所提出的研究将有益于许多不同领域的研究人员，他们对蛋白质复合体的拓扑结构特征感兴趣。对Thermo和Bruker的计划合作访问是研究的必要部分，以熟悉所需的仪器平台，计划修改，并允许供应商评估进展。最终的仪器配置将结合IM或高/超高分辨率与高效的SID碎片和增强的产品离子收集。建议的组合目前在任何商业仪器上都不可用，预计将提高分析质量，扩大可用质谱学(MS)有效表征的生物样品的范围。计划中的SID改进利用了碰撞后离开表面的离子的RF聚焦，而不是过去一直使用的纯DC聚焦。将SID纳入目前的商业超高分辨率平台尚未完成，预计将极大地扩大生物系统分析的范围和深度。在布鲁克仪器中，SID的改进将允许探索SID，不仅作为一种可能的额外激活方法，而且作为碰撞诱导解离(CID)的替代品。该项目的总体智力优势是采用高性能质谱仪，包括一种用于耐火分析物碎裂的更高性能的激活方法，现有方法无法提供直接的亚结构信息。

An award is made to Ohio State University to develop and instrument that can characterize protein complexes through improved dissociation of the complexes in mass spectrometers. Proteins in living systems can play structural roles, such as those in skin and hair; they can also play active roles such as enzymes that help with digestion, or hemoglobin that carries oxygen and carbon dioxide through the body. Cellular processes such as metabolism, cell signaling, gene expression, trafficking, cell cycle regulation, and the formation of subcellular structures are contingent upon the formation and dynamic interactions of complexes/assemblies of proteins. These protein complexes may also be bound to other biomolecules (DNA/RNA), interacting in specific ways. Because protein structure is tied to function, effective tools are needed to characterize protein-protein and protein-ligand complexes. The proposed research will result in integration of surface-induced dissociation devices into multiple types of mass spectrometers that will provide information on the structures of protein complexes important in biology. Discoveries from the project will be presented in university courses and at international, national, regional, and local workshops/ conferences on mass spectrometry and biomolecule structure and in the peer-reviewed literature. The undergraduate and graduate students and the postdoctoral associate will be trained in instrument development and will also gain collaboration skills through interactions with the private sector partners and local electronics and machine shops. Early SID adopters will serve as a direct sounding board for the success of modifications. The project is vendor- and platform-neutral, with direct inclusion of multiple private sector partners, Ardara Technologies, Waters (Q-IM-TOF, quadrupole ion mobility time of flight), Thermo (Orbitrap), and Bruker (ICR, ion cyclotron resonance), providing the project team an opportunity to present at company user meetings and an opportunity for those partners to assess the commercial value of the instrument modification.This research will develop surface-induced dissociation (SID) in high/ultra-high resolution mass spectrometers and refine its use in an ion mobility (IM) mass spectrometer whose purpose is to characterize protein complexes. The proposed research will benefit researchers from many different fields interested in the characterization of the topology of protein complexes. Planned, collaborative visits to Thermo and Bruker are a necessary part of the research, to gain familiarity with needed instrument platforms, to plan modifications, and to allow the vendors to assess progress. The final instrument configurations will combine either IM or high/ultrahigh resolving power with efficient SID fragmentation and enhanced collection of product ions. The proposed combinations are currently unavailable on any commercial instrument and are expected to enhance the quality of analysis and expand the range of biological samples that can be effectively characterized by mass spectrometry (MS). Planned improvements to SID utilize RF focusing of the ions departing the surface post collision, in contrast with the DC-only focusing that has been used in the past. Inclusion of SID in current commercial ultra-high resolution platforms has not yet been accomplished and is expected to dramatically enhance both the scope and depth of analysis on biological systems. In the Bruker instrument, the modification will allow SID to be explored, not only as a possible additional activation method, but as a replacement for collision-induced dissociation (CID). The overall intellectual merit of the project is adaptation of high performance mass spectrometers to include a higher performance activation method for fragmentation of refractory analytes, for which existing methods fail to provide direct substructure information.