Department of Analytical Chemistry, University of Vienna, Austria.
Anal Chim Acta. 2011 Oct 3;703(1):19-30. doi: 10.1016/j.aca.2011.07.012. Epub 2011 Jul 19.
Enrichment, separation and mass spectrometric analysis of biomolecules carrying a phosphate group plays an important role in current analytical chemistry. Application areas range from the preparative enrichment of phospholipids for biotechnological purposes and the separation and purification of plasmid DNA or mRNA to the specific preconcentration of phosphoproteins and -peptides to facilitate their later identification and characterization by mass spectrometry. Most of the recent improvements in this field were triggered by the need for phosphopeptide enrichment technology for the analysis of cellular protein phosphorylation events with the help of liquid chromatography-mass spectrometry. The high sensitivity of mass spectrometry and the possibility to combine this technique with different separation modes in liquid chromatography have made it the method of choice for proteome analysis. However, in the case of phosphoprotein analysis, the low abundance of the resulting phosphopeptides and their low quality fragment spectra interfere with the identification of phosphorylation events. Recent developments in phosphopeptide enrichment and fragmentation technologies successfully helped to overcome these limitations. In this review, we will focus on sample preparation techniques in the field of phosphoproteomics, but also highlight recent advancements for the analysis of other phosphorylated biomolecules.
在当前的分析化学中，对带有磷酸基团的生物分子进行富集、分离和质谱分析起着重要作用。应用领域从生物技术中磷脂的制备性富集、质粒 DNA 或 mRNA 的分离和纯化，到磷酸化蛋白和肽的特异性预浓缩，以促进其通过质谱进行后续鉴定和表征。该领域的大多数最新进展都是由于需要磷酸肽富集技术来帮助分析细胞蛋白质磷酸化事件，这是借助液相色谱-质谱完成的。质谱的高灵敏度以及将该技术与液相色谱中的不同分离模式相结合的可能性，使其成为蛋白质组分析的首选方法。然而，在磷酸蛋白分析中，所得磷酸肽的丰度低及其低质量的片段谱干扰了磷酸化事件的鉴定。磷酸肽富集和片段化技术的最新进展成功地帮助克服了这些限制。在本文中，我们将重点介绍磷酸蛋白质组学领域的样品制备技术，但也将突出介绍其他磷酸化生物分子分析的最新进展。
