Analysis of Protein Phosphorylation by Using High Mass Accuracy LC-MS Data Generated from Linear Ion Trap-Orbitrap Hybrid Mass Spectrometer

摘要

Protein phosphorylation is a vital post-translational modification and many mass spectrometry-based methods have been developed for phosphoprotein characterization. In recent years, mass spectrometers with high mass accuracy and fast scanning rate have improved the identification of phosphopeptides. However, the large scale phosphorylation study is still hindered by their low stoichiometry and the greater probability of false positive identification. We proposed here a strategy for determination of protein phosphorylation using liquid chromatography-mass spectrometry (LC-MS) data derived from a linear ion trap-Orbitrap hybrid mass spectrometer (LTQ/Orbitrap). Its accurate mass measurement was used to recognize exact mass shift after alkaline phosphatase treatment. Enzymatic dephosphorylation was used not only to provide clues to phosphopeptide signals but also to function as phosphopeptide validation.