The relative influence of phosphorylation and methylation on responsiveness of peptides to MALDI and ESI mass spectrometry
The relative influence of phosphorylation and methylation on responsiveness of peptides to MALDI and ESI mass spectrometry
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DOI：
10.1002/jms.1581
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发表时间：
2009-05-01
影响因子：
2.3
通讯作者：
Krause, Eberhard
中科院分区：
文献类型：
--
作者：
Gropengiesser, Jan;Varadarajan, Balamurugan T.;Krause, Eberhard
通过质谱法对翻译后蛋白质修饰进行定性和定量分析常常受到氨基酸修饰引起的电离/检测效率变化的阻碍。本文报道了一个全面的研究的影响，磷酸化和甲基化的肽响应基质辅助激光解吸/电离（MALDI）和电喷雾电离（ESI）质谱。使用充分表征的合成肽混合物组成的修饰肽和它们的未修饰的类似物，磷酸化，单甲基化，和二甲基化的肽的相对电离/检测效率进行了测定。我们的研究结果清楚地证实，离子产率一般较低，信号强度降低与磷酸化肽比与他们的非磷酸化类似物，这必须考虑到在MALDI和ESI质谱。然而，由磷酸化引起的离子产率的平均降低在MALDI中比在ESI中更明显。磷酸化的不可预测的影响并不取决于肽的疏水性和净电荷，这表明通过质谱法可靠地定量磷酸化需要使用内标。与磷酸化相反，单-和二甲基化肽在MALDI质谱（MALDI-MS）中经常表现出增加的信号强度。尽管有微小的基质依赖性变异，MALDI方法非常适合于二甲基化精氨酸和赖氨酸肽的灵敏检测。精氨酸胍基的单-和二甲基化不显著影响ESI-MS中肽的电离效率。
Qualitative and quantitative analysis of post-translational protein modifications by mass spectrometry is often hampered by changes in the ionization/detection efficiencies caused by amino acid modifications. This paper reports a comprehensive study of the influence of phosphorylation and methylation on the responsiveness of peptides to matrix-assisted laser desorption/ionization (MALDI) and electrospray ionization (ESI) mass spectrometry. Using well-characterized synthetic peptide mixtures consisting of modified peptides and their unmodified analogs, relative ionization/detection efficiencies of phosphorylated, monomethylated, and dimethylated peptides were determined. Our results clearly confirm that the ion yields are generally lower and the signal intensities are reduced with phosphopeptides than with their nonphosphorylated analogs and that this has to be taken into account in MALDI and ESI mass spectrometry. However, the average reduction of ion yield caused by phosphorylation is more pronounced with MALDI than with ESI. The unpredictable impact of phosphorylation does not depend on the hydrophobicity and net charge of the peptide, indicating that reliable quantification of phosphorylation by mass spectrometry requires the use of internal standards. In contrast to phosphorylation, mono- and dimethylated peptides frequently exhibit increased signal intensities in MALDI mass spectrometry (MALDI-MS). Despite minor matrix-dependent variability, MALDI methods are well suited for the sensitive detection of dimethylated arginine and lysine peptides. Mono- and dimethylation of the arginine guanidino group did not significantly influence the ionization efficiency of peptides in ESI-MS. Copyright (C) 2009 John Wiley & Sons, Ltd.