Soluble nanopolymer-based phosphoproteomics for studying protein phosphatase

Protein phosphorylation is a vital reversible post-translational modification that regulates protein-protein interactions, enzymatic activity, subcellular localization, complex formation and protein stability. The emerging field of mass spectrometry-based proteomics allows us to investigate phosphorylation and dephosphorylation on a global scale. In this review, we describe a new strategy based on soluble nanopolymers that have been used to selectively isolate phosphopeptides for mass spectrometric analysis. Functionalized soluble nanopolymers provide a homogeneous environment and linear reaction kinetics for chemical derivatization to isolate phosphopeptides with high specificity. Combined with phosphatase inhibitors and stable isotopic labeling, the approach has the capability of quantitatively measuring phosphorylation and dephosphorylation on individual sites. We provide experimental details for the approach and describe some other complementary techniques that can be used.