Chanthamontri Chamnongsak, Liu Jian, McLuckey Scott A
Department of Chemistry, Purdue University, West Lafayette, Indiana 47907-2084.
Int J Mass Spectrom. 2009 Jun 1;283(1-3):9-16. doi: 10.1016/j.ijms.2008.12.007.
Ions derived from nano-electrospray ionization (nano-ESI) of α-synuclein, a 14.5 kDa, 140 amino acid residue protein that is a major component of the Lewy bodies associated with Parkinson's disease, have been subjected to ion trap and beam-type collisional activation. The former samples products from fragmentation at rates generally lower than 100 s(-1) whereas the latter samples products from fragmentation at rates generally greater than 10(3) s(-1). A wide range of protein charge states spanning from as high as M+17H to as low as M+4H have been formed either directly from nano-ESI or via ion/ion proton transfer reactions involving the initially formed protein cations and have been subjected to both forms of collision-induced dissociation (CID). The extent of sequence information (i.e., number of distinct amide bond cleavages) available from either CID method was found to be highly sensitive to protein precursor ion charge state. Furthermore, the relative contributions of the various competing dissociation channels were also dependent upon precursor ion charge state. The qualitative trends in the changes in extent of amide bond cleavages and identities of bonds cleaved with precursor ion charge state were similar for two forms of CID. However, for every charge state examined, roughly twice the primary sequence information resulted from beam-type CID relative to ion trap CID. For example, evidence for cleavage of 86% of the protein amide bonds was observed for the M+9H precursor ion using beam-type CID whereas 41% of the bonds were cleaved for the same precursor ion using ion trap CID. The higher energies required to drive fragmentation reactions at rates necessary to observe products in the beam experiment access more of the structurally informative fragmentation channels, which has important implications for whole protein tandem mass spectrometry.
α-突触核蛋白是一种14.5 kDa、含140个氨基酸残基的蛋白质，是帕金森病相关路易小体的主要成分。对其经纳米电喷雾电离（nano-ESI）产生的离子进行了离子阱和束型碰撞活化。前者采集的是碎裂速率通常低于100 s⁻¹的产物，而后者采集的是碎裂速率通常大于10³ s⁻¹的产物。通过纳米电喷雾直接形成或通过涉及最初形成的蛋白质阳离子的离子/离子质子转移反应，形成了范围广泛的蛋白质电荷态，从高达[M + 17H]¹⁷⁺到低至[M + 4H]⁴⁺，并对这两种形式的碰撞诱导解离（CID）进行了研究。发现从任何一种CID方法获得的序列信息程度（即不同酰胺键断裂的数量）对蛋白质前体离子电荷态高度敏感。此外，各种竞争解离通道的相对贡献也取决于前体离子电荷态。两种形式的CID中，酰胺键断裂程度变化的定性趋势以及与前体离子电荷态相关的断裂键的身份相似。然而，对于所研究的每个电荷态，束型CID产生的一级序列信息大约是离子阱CID的两倍。例如，使用束型CID对[M + 9H]⁹⁺前体离子观察到86%的蛋白质酰胺键发生断裂的证据，而使用离子阱CID对相同前体离子只有41%的键发生断裂。在束实验中以观察产物所需的速率驱动碎裂反应需要更高的能量，这能进入更多结构信息丰富的碎裂通道，这对全蛋白质串联质谱具有重要意义。
