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Chromatographic and mass spectrometric methods for the identification of phosphorylation sites in phosphoproteins
Author(s) -
Hunter Ann P.,
Games David E.
Publication year - 1994
Publication title -
rapid communications in mass spectrometry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.528
H-Index - 136
eISSN - 1097-0231
pISSN - 0951-4198
DOI - 10.1002/rcm.1290080713
Subject(s) - chemistry , chromatography , identification (biology) , phosphorylation , mass spectrometry , biochemistry , botany , biology
The phosphorylation sites in a model phosphoprotein, α sl ‐casein from bovine milk, have been identified by tryptic peptide mapping (Gibson and Cohen, Methods Enzymol . vol. 193, p. 480 (1990)) employing reversed‐phase high performance liquid chromatography (RPHPLC)/electrospray ionization mass spectrometry (ES‐MS); by infusion tandem mass spectrometry (MS/MS) and LC/MS/MS in neutral loss mode of tryptic digests of α sl ‐casein, in which the characteristic neutral loss of phosphoric acid by phosphopeptides under collision‐induced dissociation (CID) conditions is exploited to highlight phosphopeptides in a tryptic digest (Covey et al. , in Methods in Protein Sequence Analysis , Jörnvall et al. (Eds), Birkhäuser Verlag, Basel 1991), and by a novel method, termed LC/CID‐MS, in which phosphopeptides are located in mixtures of peptides by the generation and detection of phosphate‐specific fragment ions during LC/ES‐MS (Huddleston et al., J. Am. Soc. Mass Spectrom . vol. 4, p. 710 (1993)). An appraisal of the efficiency, sensitivity and practicality of each of these methods in the identification of phosphorylation sites in post‐translationally modified proteins is given.