Phosphoproteomic Analysis of Peptides

Posttranslational modifications represent a major mechanism by which cells control molecular events. Sample preparation is one of the biggest keys in terms of proteomic analysis, and especially when looking at phosphoproteomics. Here we describe two techniques that are used to identify phosphopeptides. These are Titanium Dioxide and Immobilized Metal Affinity Chromatography (IMAC) which enrich for phosphopeptides. These enrichment techniques typically often identify ~35 % of the same and ~65 % unique phosphopeptides. In this chapter, we describe the use of an ion-trap mass spectrometer for the identification of phosphopeptides.