Merged Beam MALDI TOF-TOF Using Ion-Ion Reactions to Determine Structure of

本研究的最终目标是利用阳离子的电子转移解离(ETD)和负离子的质子转移解离(PTD)对具有生物意义的分子进行MALDI-TOF MS-MS检测的实用技术。这项技术将提供研究单一碰撞条件下离子-离子碰撞过程的能力，并直接测量电荷转移和碎裂的结果。将测量几种类型的离子-离子碰撞过程的竞争过程的反应截面。基础研究的重点是导致激发离子的反应，这些离子在碰撞后解离，产生结构信息丰富的碎片离子。单电荷前体可以通过采用双电荷转移来分析，首先产生有效碎片的激发中性分子，然后通过第二次碰撞使碎片电离，产生电荷与前体相反的碎片离子。除了基本知识外，这项研究还将提供确定MALDI产生的单电荷和双电荷分子离子的分子结构的实用技术。该方法应适用于多种分子，包括完整的蛋白质、多肽、寡核苷酸、寡糖、糖偶联物、脂类和代谢物，并可提供比电喷雾电离和离子陷阱更快速、灵敏和特异的未知识别和结构解释。该仪器的特点是在最近开发的高性能MALDI-TOF-TOF MS-MS系统中增加了第二个脉冲离子源。来自两个源的脉冲光束在碰撞池内合并，脉冲在时间上准确地同步，并且光束的相对速度被准确地确定并且非常接近于零。建议的装置将允许在以前无法获得的低平动能下灵敏地测量相反极性的离子之间的反应以及离子与激发的中性物质之间的反应。反应物的内部温度(或激发)可能相当高，但平移温度很低，导致这些过程的横截面非常高。约化质量为100Da、相对速度为1m/S的碰撞相当于5×10~(-7)eV的平动能和6×10~(-3)K的平移温度。 与公共卫生相关(由申请人提供)：生物应用质谱学的“圣杯”是一种仪器，它提供了关于复杂混合物中痕量蛋白质的准确的相对分子质量和完整、明确的序列，包括不寻常的氨基酸和翻译后修饰。它还应提供关于其他生物重要分子的质量和结构的类似信息，包括代谢物、脂类、低聚糖、糖共轭、寡核苷酸和多肽。可能就是这样了。

DESCRIPTION (provided by applicant): Merged Beam MALDI TOF-TOF Using Ion-Ion Reactions to Determine Structure of Biological Molecules Summary The ultimate goals of this research are practical techniques for MALDI-TOF MS-MS on biologically significant molecules using electron transfer dissociation (ETD) of positive ions and proton transfer dissociation (PTD) of negative ions to determine molecular structure. This technology will provide the ability to study ion-ion collision processes under single collision conditions with direct measurement of the results in terms of charge transfer and fragmentation. Reaction cross sections for competing processes will be measured for several types of ion-ion collision processes. The emphasis in the fundamental studies is on reactions that lead to excited ions that dissociate following collision to produce structurally informative fragment ions. Singly charged precursors may be analyzed by employing double charge transfer to first produce excited neutrals that fragment efficiently, and then ionize the fragments by a second collision to produce fragment ions with charge opposite that of the precursor. In addition to fundamental knowledge, this research will provide practical techniques for determining molecular structure of singly and doubly charged molecular ions produced by MALDI. The approach should be applicable to a wide variety of molecules including intact proteins, peptides, oligonucleotides, oligosaccharides, glycoconjugates, lipids, and metabolites, and may provide speed, sensitivity, and specificity for identification and structural elucidation of unknowns that surpasses that available with electrospray ionization and ion traps. The instrument features addition of a second pulsed ion source to a recently developed high- performance MALDI-TOF-TOF MS-MS system. Pulsed beams from the two sources are merged within a collision cell with the pulses accurately synchronized in time, and with the relative velocity of the beams accurately determined and very near zero. The proposed apparatus will allow sensitive measurements of reactions between ions of opposite polarity and between ions and excited neutrals at low translational energies not previously accessible. The internal temperature (or excitation) of the reactants may be quite high but the translational temperatures are very low leading to very high cross sections for these processes. Collisions with reduced mass of 100 Da and relative velocity of 1 m/s correspond to translational energy of 5x10-7 eV, and translational temperature of 6x10-3 K. We are not aware of any previous work on ion-ion collision processes under these conditions. PUBLIC HEALTH RELEVANCE (provided by applicant): The "Holy Grail" of mass spectrometry for biological applications is an instrument that provides accurate molecular weight and complete, unambiguous sequence, including unusual amino acids and post- translational modifications, on proteins present at trace levels in complex mixtures. It should also provide similar information on mass and structure of other biologically significant molecules, including metabolites, lipids, oligosaccharides, glycoconjugates, oligonucleotides, and peptides. This may be it.