Differentiating alpha- and beta-aspartic acids by electrospray ionization and low-energy tandem mass spectrometry

Spectra obtained by low-energy electrospray ionization tandem mass spectrom etry (ESI-MS/MS) of 34 peptides containing aspartic acids at position n wer e studied and unambiguously differentiated. beta -Aspartic acid yields an i nternal rearrangement similar to that of the C-terminal rearrangements of p rotonated and cationized peptides, As a result of this rearrangement, two d ifferent ions containing the N- and the C-terminal ends of the original pep tide are formed, namely, the b(n-1)+ H2O and y"(l-n+1) - 46 ions, respectiv ely, where l is the number of amino acid residues in the peptide. The struc ture suggested for the y"(l-n+1) - 46 ion is identical to that proposed for the v(n) ions observed upon high-energy collision-induced dissociation (CI D) experiments. The intensity of these ions in the low-energy MS/MS spectra is greatly influenced by the presence and position of basic amino acids wi thin the sequences, Peptides with a basic amino acid residue at position it - 1 with respect to the beta -aspartic acid yield very intense b(n-1) + H2 O ions, while the y"(l-n+1) - 46 ion was observed mostly in tryptic peptide s. Comparison between the high- and low-energy MS/MS spectra of several iso peptides suggests that a metastable fragmentation process is the main contr ibutor to this rearrangement, whereas for long peptides (40 AA) CID plays a more important role. We also found that alpha -aspartic acid containing pe ptides yield the normal immonium ion at 88 La, while peptides containing be ta -aspartic acid yield an ion at m/z 70, and a mechanism to explain this p henomenon is proposed. Derivatizing isopeptides to form quaternary amines, and performing MS/MS on the sodium adducts of isopeptides, both improve the relative intensity of the b(n+1) + H2O ions. Based on the above findings, it was possible to determine the isomerization sites of two aged recombinan t growth proteins. Copyright (C) 2000 John Wiley & Sons, Ltd.