Method Development for Qualitative, Quantitative and Computational Proteomics

Protein phosphorylation is an important posttranslational modification that plays a regulatory role within numerous biological processes. The simultaneous identification, localization, and quantification of phosphorylated proteins is vital for understanding this dynamic control mechanism. The application of isobaric labeling strategies, e.g., iTRAQ, for quantitative phosphopeptide analysis requires (i) optimal peptide fragmentation conditions, (ii) sophisticated computational proteomics algorithms to identify (phosphorylated) iTRAQ labeled peptides, and (iii) the ability to use more than one Peptide-Spectra-Match and phosphopeptide sequence to guarantee accurate phosphorylation specific quantification. \nThese three demands were combined into a platform to relatively quantify iTRAQ-4Plex labeled (phospho)proteins on a LTQ Orbitrap Velos.