N-Terminal amino acid side-chain cleavage of chemically modified peptides in the gas phase: A mass spectrometry technique for N-terminus identification
peer-reviewed · Bioorganic & Medicinal Chemistry · 2006
| Date | 2006-09-01 |
| Type | peer-reviewed |
| Venue | Bioorganic & Medicinal Chemistry |
| Publisher | Elsevier BV |
| Contribution | adjacent |
| DOI | 10.1016/j.bmc.2006.05.060 |
| Citations (OpenAlex) | 28 |
Abstract
Although genome databases have become the key for proteomic analyses, de novo sequencing remains essential for the study of organisms whose genomes have not been completed. In addition, post-translational modifications present a challenge in database searching. Recognition of the b or y-ion series in a peptide MS/MS spectrum as well as identification of the b1 - and yn-1 -ions can facilitate de novo analyses. Therefore, it is valuable to identify either amino-acid terminus. In previous work, we have demonstrated that peptides modified at the epsilon-amino group of lysine as a t-butyl peroxycarbamate derivative undergo free radical promoted peptide backbone fragmentation under low-energy collision-induced dissociation (CID) conditions. Here we explore the chemistry of the N-terminal amino group modified as a t-butyl peroxycarbamate. The conversion of N-terminal amines to peroxycarbamates of simple amino acids and peptides was studied with aryl t-butyl peroxycarbonates. ESI-MS/MS analysis of the peroxycarbamate adducts gave evidence of a product ion corresponding to the neutral loss of the N-terminal side chain (R), thus identifying this residue. Further fragmentation (MS3) of product ions formed by N-terminal residue side-chain loss (-R) exhibited an m/z shift of the b-ions equal to the neutral loss of R, therefore labeling the b-ion series. The study was extended to the analysis of a protein tryptic digest where the SALSA algorithm was used to identify spectra containing these neutral losses. The method for N-terminus identification presented here has the potential for improvement of de novo analyses as well as in constraining peptide mass mapping database searches.
Methods and tools
- Peroxycarbamate N-terminal labelling: N-terminal t-butyl peroxycarbamate derivatives lose the N-terminal side chain in CID, identifying that residue and labelling the b-ion series to aid de novo sequencing.