Metal oxide-based enrichment combined with gas-phase ion-electron reactions for improved mass spectrometric characterization of protein phosphorylation.
Gas-phase ion-electron reactions, including electron capture dissociation (ECD) and electron detachment dissociation (EDD), are advantageous for characterization of protein posttranslational modifications (PTMs), because labile modifications are not lost during the fragmentation process. However, at least two positive charges and relatively abundant precursor ions are required for ECD due to charge reduction and lower fragmentation efficiency compared to conventional gas-phase fragmentation techniques. Both these criteria are difficult to fulfill for phosphopeptides due to their acidic character. The negative ion mode operation of EDD is more compatible with phosphopeptide ionization, but EDD suffers from a fragmentation efficiency even lower than that of ECD. Recently, metal oxides such as ZrO 2 and TiO 2 have been shown to provide selective enrichment of phosphopeptides from proteolytic digests. Here, we utilize this enrichment strategy to improve ECD and EDD of phosphopeptides. This approach allowed determination of the locations of phosphorylation sites in highly acidic, multiply phosphorylated peptides from complex peptide mixtures by ECD. For singly phosphorylated peptides, EDD provided complementary sequence information compared to ECD.
Kweon
Hye Kyong
HK
Department of Chemistry, University of Michigan, 930 North University Avenue, Ann Arbor, Michigan 48109-1055, USA.
Håkansson
Kristina
K
D000595:Amino Acid Sequence; D004583:Electrons; D005740:Gases; D007477:Ions; D008969:Molecular Sequence Data; D044367:Phase Transition; D010748:Phosphopeptides / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D017550:Spectroscopy, Fourier Transform Infrared; D014025:Titanium; D015040:Zirconium
D005740:Gases; D007477:Ions; D010748:Phosphopeptides; D010750:Phosphoproteins; C009495:titanium dioxide; D015040:Zirconium; D014025:Titanium; C028541:zirconium oxide