Gas-Phase Chemistry of Biological and Metal-Containing Ions

生物质谱（MS）的概念发展和技术突破使蛋白质组学的进步成为可能。我们的发现研究计划的长期目标将继续致力于更好地理解支撑肽和蛋白质鉴定和表征的科学的基本方面。在短期内，在这个资助周期内，我们的目标是探索(1)质子化肽（[M + H]+）及其关键片段的结构、形成、能量学和碎片化学；(2)多肽的分子自由基阳离子（M•+）；(3)金属化肽配合物。这些问题将采用实验和理论相结合的方法来解决。实验技术将包括：(i)串联质谱与同位素标记13C， 2H， 18O和15N；（ii）阈值碰撞诱导解离（CID）在配有环形离子导q2的改良质谱仪上；（iii）在FELIX设施中进行的IRMPD光谱分析。在此，我们建议继续提高我们对质子化肽及其关键片段的结构和解离化学的理解。我们将从选定的a(n)产物离子和它们的下一代碎片开始，这些碎片是在失去一个小的中性产物后形成的，比如氨产生a(n)*，水产生a(n)0离子。我们将研究侧链官能团对a(2)异构体的相对能量学和稳定性的影响，特别是新发现的亚胺-酰胺结构（L2）和环结构（C1），通过进行包括碱性和酸性残基的系统研究。对a(n)*和a(n)0离子的结构所知相对较少。初步结果表明，五丙氨酸（A5）的a(3)*和a(4)*离子具有亚胺-恶唑酮结构，具有非常规结构。令人惊讶的是，五甘氨酸（G5）的a(4)*离子不是亚胺-恶唑酮，而是质子化咪唑酮，这是我们首先提出的将G4脱水质子化的一类新产物，名义上是b4离子。此外，G5的a(4)*离子的IRMPD谱与a(4)0的IRMPD谱非常相似，这表明这两种结构都是质子化的咪唑酮。我们将充分探索和确定导致不同异构体产物的机制和能量学。我们将继续提高我们对决定肽自由基离子结构稳定性的因素及其断裂机制的理解。在非末端位置含有单个酪氨酸(Y)或色氨酸(W)残基的甘氨酸寡肽中，M•+的CID通常显示出最突出的产物来自于Y或W的N-C （α）键断裂后形成的质子结合的“二聚体”。令人惊讶的是，含有单个Y残基的低聚丙氨酸的CID的初步数据揭示了额外的和令人困惑的断裂通道。我们计划通过沿着低聚丙氨酸框架“移动”Y，并将Y作为第三、第四和后续残基，来充分研究这些迷人的碎片化通道。此外，我们还将检查含有单个W残基的低聚丙氨酸的M•+的CID结果。我们计划继续研究金属化肽复合物的多阶段质谱，以产生不寻常的和迷人的肽离子，包括散正质子化的a3和a2离子，它们由于体积小而具有高电荷密度。我们将LaIII产生这些离子的有效性归因于它的高电荷和它的电荷离域到配体上的倾向，以及La对O的高亲和力（从而促进COO-的裂解）。我们计划将研究范围扩展到使用Ce•3+和其他镧系离子作为金属化中心。

Advances in proteomics have been made possible by conceptual developments and technological breakthroughs in biological mass spectrometry (MS). The long-term objective of our Discovery research program will continue to aim at better understanding fundamental aspects of the science that underpins peptide and protein identification and characterization. In the shorter term, and within the duration of this funding cycle, our objectives are to explore the structure, formation, energetics and fragmentation chemistry of (1) protonated peptides ([M + H]+) and their key fragments; (2) molecular radical cations of peptides (M•+); and (3) metalated peptide complexes. These will be tackled using a combination of experimental and theoretical approaches. Experimental techniques will include: (i) tandem MS with isotopic labeling with 13C, 2H, 18O and 15N; (ii) threshold collision-induced dissociation (CID) on a modified MS instrument fitted with a ring ion guide as q2; and (iii) IRMPD spectroscopy performed in the FELIX facility.We propose herein to continually improve our understanding of the structures and dissociation chemistry of protonated peptides and their key fragments. We will start with selected a(n) product ions and their next-generation fragments formed after losing a small neutral product, such as ammonia giving a(n)*, and water giving a(n)0 ions. We will examine the effects of side-chain functional groups on the relative energetics and stability of a(2) isomers, especially those of a newly discovered imine-amide structure (L2) and a cyclic structure (C1), by performing a systematic study including basic and acidic residues. Relatively little is known about the structures of a(n)* and a(n)0 ions. Our preliminary results strongly suggest that the a(3)* and a(4)* ions of pentaalanine (A5) have unconventional structures in that they have imine-oxazolone structures. Surprisingly, the a(4)* ion of pentaglycine (G5) is not an imine-oxazolone, but a protonated imidazolone, a new class of products first proposed by us for dehydrated protonated G4, a nominal b4 ion. In addition, the IRMPD spectrum of the a(4)* ion of G5 closely resembles that of the a(4)0, thereby suggesting both structures are protonated imidazolones. We will fully explore and determine the mechanisms and energetics leading to the different isomeric products.We will continue improving our understanding of factors that determine structural stability in peptide radical ions and their fragmentation mechanisms. CID of M•+ from oligopeptides of glycine containing a single tyrosine (Y) or tryptophan (W) residue in a non-terminal position typically shows most-prominent products that originate from a proton-bound ‘dimer’ formed after cleavage of the N–C(alpha) bond of Y or W. Surprisingly, preliminary data on the CID of oligoalanines containing a single Y residue reveal additional and puzzling fragmentation channels. We plan to fully investigate these fascinating fragmentation channels by ‘moving’ Y along the oligoalanine framework and having Y as the third, fourth and subsequent residues. In addition, we will also examine CID results of the M•+ from oligoalanines containing a single W residue. We plan to continue examining multi-stage MS of metalated peptide complexes to generate unusual and fascinating peptide ions, including dispositive protonated a3 and a2 ions which have high charge-density due to their small size. We attribute the effectiveness of LaIII in generating these ions to its high charge and its propensity to delocalize its charge onto the ligand, as well as the high affinity of La for O (thereby facilitating the cleavage of COO-). We plan to extend the line of study to using Ce•3+ and other lanthanide ions as the metalation center.