Journal
JOURNAL OF PROTEOME RESEARCH
Volume 4, Issue 1, Pages 101-108Publisher
AMER CHEMICAL SOC
DOI: 10.1021/pr049837+
Keywords
dimethyl stable isotope labeling; quantitative proteomics; de novo sequencing; mass spectrometry; reductive methylation
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Stable isotope-based dimethyl labeling that produces a dimethyl labeled terminal amine or a monomethylated proline N-terminus by reductive methylation (Anal. Chem. 2003, 75, 6843-6852) was reported as a promising strategy for global quantitative proteomics because of the simplicity of the process and its fast and complete reaction. This labeling strategy provides a signal enhancement for the produced a, ions, which are usually hard to detect among most of the nonderivatized fragments. To assist peptide sequencing, in this study, the enhanced a, ion produced under either collision induced dissociation (CID) or post source decay (PSD) modes was further characterized and applied as a mass tag for fingerprinting the identity of N-terminal amino acid. On the basis of the analysis of standard peptides, tryptic digests of hemoglobin and cell lysates, it was proved that such signal enhancement occurred to a, ions derived from all 20 of the amino acids residues and this phenomenon was explained based the formation of stable quaternary immoniun ions. Accurate determination of a, ions was shown to increase the chance for peptide de novo sequencing and also provided higher confidence in the scores obtained when identifying a protein through database searching. In addition, the a, ion was further demonstrated to be used as a universal tag for precursor ion scan in a Q-TOF instrument, leading to a greater number of peptide ions sequenced. Combined with the capability for differential quantitation, the stable isotope-based dimethyl labeling increases the usefulness of the labeling method for MS-based proteomics.
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