Development of phosphopeptide enrichment techniques for phosphoproteome analysis.

Protein phosphorylation is one of the most biologically relevant and ubiquitous post-translational modifications. The analysis of protein phosphorylation is very challenging due to its highly dynamic nature and low stoichiometry. In this article, recent techniques developed for phosphoproteome analysis are reviewed with an emphasis on the new developments in this field in China. To improve the performance of phosphoproteome analysis, many novel methods, either by application of new separation mechanisms or by adoption of new separation materials, were developed to specifically enrich phosphopeptides from complex protein digests. A series of new materials, including nanostructure materials, magnetic materials, and monolithic materials, were applied to prepare immobilized affinity chromatography or metal oxide affinity chromatography to improve the performance of phosphopeptide enrichment. Besides, new software tools were also developed to validate phosphopeptide identification and predict kinase specific phosphorylation sites.