Influence of the metals and ligands in dinuclear complexes on phosphopeptide sequencing by electron-transfer dissociation tandem mass spectrometry
Influence of the metals and ligands in dinuclear complexes on phosphopeptide sequencing by electron-transfer dissociation tandem mass spectrometry
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DOI：
10.1039/c8cp04516j
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发表时间：
2018-11-07
影响因子：
3.3
通讯作者：
Gabelica, Valerie
中科院分区：
文献类型：
--
作者：
Asakawa, Daiki;Miyazato, Akio;Gabelica, Valerie
磷酸化修饰是蛋白质修饰中最重要的一种，电子转移解离串联质谱（ETD MS/MS）是一种潜在的磷酸化肽序列分析方法，包括磷酸化位点的测定。值得注意的是，当前体含有多于三个正电荷时，ETD-MS/MS通常提供有用的信息。由于在具有三个以上正电荷的酸性磷酸肽的生产中出现困难，它尚未用作大规模磷酸肽生产的分析方法。为了增加磷酸肽的电荷状态，我们使用双核金属配合物，其通过添加正电荷选择性地结合磷酸肽中的磷酸基团。双核铜，锌，镓配合物进行了测试，发现存在于复杂的金属的类型强烈影响的亲和力的磷酸化的化合物和它们的ETD片段。双核铜配合物弱相互作用的磷酸基团和ETD诱导的肽片段在很大程度上抑制了Cu 2+的存在下，作为一个电子陷阱。双核镓配合物与磷酸基团牢固结合。然而，与镓结合的配体充当电子陷阱，并且在ETD-MS/MS的前体中存在双核镓络合物阻碍了磷酸肽的测序，如在双核铜络合物的情况下。与此相反，双核锌配合物有效地结合到磷酸肽的电荷状态的增加，促进磷酸肽测序ETD-MS/MS。在双核锌-磷酸肽复合物的配体和肽骨架的碎片竞争诱导ETD。这些过程是受配体结构的影响，因此详细的ETD断裂途径进行了研究，使用密度泛函理论计算。
Phosphorylation is one of the most important protein modifications, and electron-transfer dissociation tandem mass spectrometry (ETD-MS/MS) is a potentially useful method for the sequencing of phosphopeptides, including determination of the phosphorylation site. Notably, ETD-MS/MS typically provides useful information when the precursor contains more than three positive charges. It is not yet used as an analysis method for large-scale phosphopeptide production due to difficulties occurring in the production of acidic phosphopeptides having more than three positive charges. To increase the charge state of phosphopeptides, we used dinuclear metal complexes, which selectively bind to the phosphate group in phosphopeptides with the addition of positive charge(s). Dinuclear copper, zinc, and gallium complexes were tested and it was found that the type of metal present in the complex strongly affected the affinity of the phosphorylated compounds and their ETD fragmentation. The dinuclear copper complex interacted weakly with the phosphate groups and ETD-induced peptide fragmentation was largely suppressed by the presence of Cu2+, which worked as an electron trap. The dinuclear gallium complex was strongly bound to a phosphate group. However, the ligand binding to gallium acted as an electron trap and the presence of dinuclear gallium complex in the precursor for ETD-MS/MS hampered the sequencing of the phosphopeptides, as in the case of dinuclear copper complexes. In contrast, dinuclear zinc complexes efficiently bind to phosphopeptides with an increase in the charge state, facilitating phosphopeptide sequencing by ETD-MS/MS. The fragmentation of the ligand and peptide backbone in the dinuclear zinc-phosphopeptide complex were competitively induced by ETD. These processes are influenced by the ligand structure and so the detailed ETD fragmentation pathways were investigated using density functional theory calculations.