Conversion of arginine to ornithine for improving the fragmentation pattern of peptides labeled with the N-terminal tris(2,4,6-trimethoxyphenyl)phosphoniumgroup in tandem mass spectrometry

Abstract

We propose a method of converting arginine to ornithine residues by controlled hydrazinolysis, in order to facilitate the sequencing of peptides by tandem mass spectrometry (MS/MS). Whereas the presence of C-terminal arginine residue occurring in a tryptic peptide is generally desirable in MS and MS/MS analyses, that of arginine in the middle of a peptide often adversely affects the appearance of fragment peaks. This problem still arises in peptides, which are acylated with a reagent containing the tris(2,4,6-trimethoxyphenyl)phosphonium (TMPP) group at the N-termini. Using four model peptides, angiotensin III (RVYIHPF), a mucin-related peptide (APDTRPAPG), α-bag cell peptide (1–9) (APRLRFYSL), and [des-Pro²] bradykinin (RPGFSPFR), we optimized the protocol of hydrazinolysis to remove the guanidino group of arginine. Owing to this derivatization, we obtained much simpler MS/MS spectra composing mainly a-type ions characteristic of peptides tagged with the TMPP group. Formylation of the δ-amino group of ornithine further enhanced the efficacy of the derivatization, which would be applicable to the sequencing of non-tryptic peptides.