Niobium(V) Oxide (Nb<sub>2</sub>O<sub>5</sub>): Application to Phosphoproteomics

Proteomics-based analysis of signaling cascades relies on a growing suite of affinity resins and methods aimed at efficient enrichment of phosphorylated peptides from complex biological mixtures. Given the heterogeneity of phosphopeptides and the overlap in chemical properties between phospho- and unmodified peptides, it is likely that the use of multiple resins will provide the best combination of specificity, yield, and coverage for large-scale proteomics studies. Recently titanium and zirconium dioxides have been used successfully for enrichment of phosphopeptides. Here we report the first demonstration that niobium pentoxide (Nb<sub>2</sub>O<sub>5</sub>) provides for efficient enrichment and recovery (∼50−100%) of phosphopeptides from simple mixtures and facilitates identification of several hundred putative sites of phosphorylation from cell lysate. Comparison of phosphorylated peptides identified from Nb<sub>2</sub>O<sub>5</sub> and TiO<sub>2</sub> with sequences in the PhosphoELM database suggests a useful degree of divergence in the selectivity of these metal oxide resins. Collectively our data indicate that Nb<sub>2</sub>O<sub>5</sub> provides efficient enrichment for phosphopeptides and offers a complementary approach for large-scale phosphoproteomics studies.