Charge-state-dependent sequence analysis of protonated ubiquitin ions via ion trap tandem mass spectrometry
Charge-state-dependent sequence analysis of protonated ubiquitin ions via ion trap tandem mass spectrometry
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DOI：
10.1021/ac0101095
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发表时间：
2001-07-15
影响因子：
7.4
通讯作者：
McLuckey, SA
中科院分区：
文献类型：
--
作者：
Reid, GE;Wu, J;McLuckey, SA
中的一个;控制串联质谱法对完整多质子化蛋白质进行“自上而下”序列分析的主要因素是前体离子电荷状态对产物离子形成的影响。为了更全面地了解这种影响，将电喷雾电离与四极杆离子阱质谱仪联用，采用碰撞诱导解离和气相离子/离子反应来检查 牛泛素的 [M + 12H](12+)、[M + H](+) 离子断裂。在低电荷状态（+1 至 +6）下，观察到质子化前体中 NH3 或 H2O 的损失以及天冬氨酸残基的定向裂解。在中间电荷状态（+7、+8 和 +9），观察到蛋白质骨架的广泛非特异性断裂，仅从 [M + 8H](8+) 离子获得 50% 的序列覆盖率。在高电荷状态（+10、+11、+12），观察到的单一主导通道是单个脯氨酸残基的优先断裂。这些数据可以根据当前的分子内质子动员模型（即“移动质子模型”）、为质子化肽开发的酰胺键解离机制以及气相中泛素的多电荷离子的结构（通过离子淌度和氢/氘交换测量来检查）来容易地解释。
One of the; major factors governing the "top-down" sequence analysis of:intact multiply protonated proteins by tandem mass spectrometry is the effect of the precursor ion charge state on the formation of product ions., To more fully understand this effect electrospray ionization coupled to a quadrupole ion trap mass spectrometer, collision-induced dissociation, and gas-phase ion/ion reactions have been employed to examine the fragmentation of the [M + 12H](12+), [M + H](+) ions of bovine ubiquitin. At low charge states (+1 to +6), loss of NH3 or H2O from the protonated precursor and directed cleavage at:aspartic acid residues was observed. At intermediate charge states, (+7, +8, and +9), extensive nonspecific fragmentation of the protein backbone was observed, with 50% sequence coverage obtained from the [M + 8H](8+) ion alone, At high charge states, (+10, +11, +12), the single dominant channel that was observed was the preferential fragmentation of;a single proline residue. These data can be readily explained in terms of the current model for intramolecular proton mobilization, that is, the "mobile proton model", the mechanisms for amide bond dissociation developed for protonated peptides, as well as the structures of the multiply charged ions of ubiquitin in the gas phase, examined by ion mobility and hydrogen/deuterium exchange measurements.