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The identification of peptide modifications derived from gel‐separated proteins using electrospray triple quadrupole and ion trap analyses
Author(s) -
Swiderek Kristine M.,
Davis Michael T.,
Lee Terry D.
Publication year - 1998
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.1150190614
Subject(s) - chemistry , fragmentation (computing) , mass spectrum , mass spectrometry , tandem mass spectrometry , electrospray , collision induced dissociation , ion trap , chromatography , ion , peptide mass fingerprinting , protein mass spectrometry , electrospray ionization , top down proteomics , peptide , bottom up proteomics , triple quadrupole mass spectrometer , quadrupole ion trap , database search engine , selected reaction monitoring , proteomics , organic chemistry , biochemistry , search engine , information retrieval , computer science , gene , operating system
Microspray tandem mass spectrometry (MS/MS) in combination with database search routines has become a powerful tool for the identification of proteins from femtomole amounts of material following gel electrophoresis and in‐gel digestion procedures. However, artifactual modification of susceptible residues can arise during gel electrophoresis, leading to unexpected peptide mass shifts during mass analysis. Consequently, collision‐induced dissociation (CID) spectra generated from these derivatized peptides can defy direct interpretation by automated database search routines and remain unidentified. Here, we evaluate the MS/MS spectra of peptides carrying oxidized derivatives of tryptophane and methionine residues, and various modifications of cysteine. We demonstrate that certain of these modifications generate characteristic fragmentation patterns or “fingerprints”, during CID analysis, the knowledge of which can facilitate the interpretation of the spectra. We will show that these signature fragment ions are predominantly produced during the CID analysis of singly charged ions although they can be observed in the MS/MS spectra of the doubly charged species as well. In other cases, the CID spectrum lacks a characteristic fingerprint and the modification remains silent. However, CID spectra of related peptides, differing only by their modifications, are similar and all or part of the fragment ion spectra will have shifted by a discreet mass, which facilitates the identification of the modified residue. At the same time, the comparison of related spectra can prevent misinterpretations such as the assignment of a residue mass to the wrong amino acid or a neutral loss fragment ion to a y ‐ or b ‐ion.