Abstract
Reversible protein phosphorylation represents one of the most dynamic post-translational modifications. The importance of phosphoproteins is often not reflected in their abundance, as their non-phosphorylated counterparts are usually present in much greater quantities within the cell.
Moreover, phosphorylation is a transient modification, so the protein in question could be present in both native and phosphorylated forms. Phosphoprotein identification is further complicated by technical issues with mass spectrometry (MS).
To increase the number of phosphoproteins that can be identified, it is necessary to remove non-phosphorylated proteins or peptides from samples and enrich for the phosphorylated isoforms prior to MS. This task is carried out by the use of enriching techniques that can be performed at two levels-at the level of intact phosphoproteins, or at the level of peptides.
This article offers an overview of currently used enrichment protocols. It will pinpoint their advantages as well as their limitations.