Selective zirconium dioxide-based enrichment of phosphorylated peptides for mass spectrometric analysis.
Due to the dynamic nature and low stoichiometry of protein phosphorylation, enrichment of phosphorylated peptides from proteolytic mixtures is often necessary prior to their characterization by mass spectrometry. Several phosphopeptide isolation strategies have been presented in the literature, including immobilized metal ion affinity chromatography. However, that technique suffers from poor selectivity and reproducibility. Recently, titanium dioxide-based columns have been successfully employed for phosphopeptide enrichment by several research groups. Here, we present, to our knowledge, the first demonstration of the utility of zirconium dioxide microtips for phosphopeptide isolation prior to mass spectrometric analysis. These microtips display similar overall performance as TiO2 microtips. However, more selective isolation of singly phosphorylated peptides was observed with ZrO2 compared to TiO2 whereas TiO2 preferentially enriched multiply phosphorylated peptides. Thus, these two chromatographic materials possess complementary properties. For alpha- and beta-casein, Glu-C digestion provided no evident advantage compared to trypsin digestion when combined with TiO2 or ZrO2 phosphopeptide enrichment.
Kweon
Hye Kyong
HK
Department of Chemistry, University of Michigan, 930 North University Avenue, Ann Arbor, Michigan 48109-1055, USA.
Håkansson
Kristina
K
D013058:Mass Spectrometry / Q000379:methods*; D010748:Phosphopeptides / Q000032:analysis* / Q000302:isolation & purification*; D010750:Phosphoproteins / Q000032:analysis; D012680:Sensitivity and Specificity; D014025:Titanium / Q000737:chemistry; D015040:Zirconium / Q000737:chemistry*
D010748:Phosphopeptides; D010750:Phosphoproteins; C009495:titanium dioxide; D015040:Zirconium; D014025:Titanium; C028541:zirconium oxide