Characterization of the Phosphoproteome in Androgen-Repressed Human Prostate Cancer Cells by Fourier Transform Ion Cyclotron Resonance Mass Spectrometry

Androgen-repressed human prostate cancer, ARCaP, grows and is highly metastatic to bone and soft tissues in castrated mice. The molecular mechanisms underlying the aberrant responses to androgen are not fully understood. Here, we apply state-of-the-art mass spectrometry methods to investigate the phosphoproteome profiles in ARCaP cells. Because protein biological phosphorylation is always substoichiometric and the ionization efficiency of phosphopeptides is low, selective enrichment of phosphorylated proteins/peptides is required for mass spectrometric analysis of phosphorylation from complex biological samples. Therefore, we compare the sensitivity, efficiency, and specificity for three established enrichment strategies: calcium phosphate precipitation (CPP), immobilized metal ion affinity chromatography (IMAC), and TiO2-modified metal oxide chromatography. Calcium phosphate precipitation coupled with the TiO2 approach offers the best strategy to characterize phosphorylation in ARCaP cells. We analyzed p...