Wenjun Mo

Novel rearranged ions observed for protonated peptides via metastable decomposition in matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

C-terminal rearrangement ions [bn−1 + H2O] (where n refers to the total number of residues of peptides) are frequently observed for peptides which contain basic amino acid(s), especially arginine, at or near their N termini in low- and high-energy collision-induced dissociation or post-source decay (PSD) spectra. Here we report a novel rearrangement, associated with PSD for serine- or threonine-containing peptides that are susceptible to C-terminal rearrangement. Based on PSD analyses of serine- or threonine-containing bradykinin and its analogs, which have been ethyl-esterified or 18O labeled at their C termini, the [bk + H2O] (where k denotes the position adjacent to the left of the Ser/Thr residue) ion is generally thought to be formed by the transfer of the hydroxyl moiety of a serine or threonine residue to the carbonyl group of the residue to its left accompanied by the loss of the remaining C-terminal portion of the peptide. When the Ser/Thr is at or near the C terminus, the present [bk + H2O] ion could be formed via two pathways, i.e., the Ser/Thr-related rearrangement and the conventional C-terminal rearrangement, which has been clearly verified by 18O labeling at the C terminus. In addition, the ions which are formally designated as [ymbl + H2O], where ymbl denotes a b-type internal ion, are also briefly described.

Accurate peptide sequencing by post-source decay matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

Partial 18O-labeling of peptides has been applied to post-source decay experiments in a matrix-assisted laser desorption/ionization time-of-flight mass spectrometer. The ions which originate from the carboxyl terminus of a peptide partially retain 18O atoms which have readily been incorporated into the C-terminal carboxyl groups during enzymatic hydrolysis in a buffer containing 40 atom percent H218O. The isotopic resolution of singly charged precursor ions and their product ions obtained for peptides up to ca. 2800 Da has been achieved using the delayed extraction method, which permits the rapid identification and assignment of the 18O-labeled and non-labeled ion species in the PSD spectra. The results obtained from several 18O-labeled peptides, derived from an enzymatic digest, demonstrate the accuracy and reproducibility of the present method, which will be in widespread use for protein identification via database searching or for investigations of totally unknown proteins.

Accurate peptide sequencing by mass spectrometry

Mass spectrometry for the measurement of product ions produced upon collision-induced-dissociation (CID) or post-source decay (PSD) is a well accepted method for the characterization of biomolecules, especially for peptide sequencing and the analysis of post-translational modifications of proteins. However, the assignment of the product ions is not always definite, rendering the resulting data ambiguous. Moreover, the manual interpretation of the spectra requires considerable skill, without which, misassignments of ions can occur. To circumvent these issues, we [l–3] and others [4,5] earlier reported on the 18 O labeling of peptides for rapid and accurate sequence analysis of peptides and a software program designed to aid in peptide sequencing [6–9], which is based on lowand high-energy CID mass spectra. In this study, we describe an extension of this method to PSD experiments, using a high-resolution matrix-assisted laser desorption/ionization (MALDI) time-of-flight mass spectrometer (TOFMS), along with a new version of the software, ‘SeqMS ’, which has been modified for the automated interpretation of PSD spectra.