Differentiating alpha- and beta-aspartic acids by electrospray ionization and low-energy tandem mass spectrometry.
Spectra obtained by low-energy electrospray ionization tandem mass spectrometry (ESI-MS/MS) of 34 peptides containing aspartic acids at position n were studied and unambiguously differentiated. beta-Aspartic acid yields an internal rearrangement similar to that of the C-terminal rearrangements of protonated and cationized peptides. As a result of this rearrangement, two different ions containing the N- and the C-terminal ends of the original peptide are formed, namely, the bn-1 + H2O and y"l - n + 1 - 46 ions, respectively, where e is the number of amino acid residues in the peptide. The structure suggested for the y"l - n + 1 - 46 ion is identical to that proposed for the vn ions observed upon high-energy collision-induced dissociation (CID) 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 within the sequences. Peptides with a basic amino acid residue at position n - 1 with respect to the beta-aspartic acid yield very intense bn-1 + H2O ions, while the y"l - n + 1 - 46 ion was observed mostly in tryptic peptides. Comparison between the high- and low-energy MS/MS spectra of several isopeptides suggests that a metastable fragmentation process is the main contributor to this rearrangement, whereas for long peptides (40 AA) CID plays a more important role. We also found that alpha-aspartic acid containing peptides yield the normal immonium ion at 88 Da, while peptides containing beta-aspartic acid yield an ion at m/z 70, and a mechanism to explain this phenomenon 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 bn + 1 + H2O ions. Based on the above findings, it was possible to determine the isomerization sites of two aged recombinant growth proteins.
González
L J
LJ
Center for Genetic Engineering and Biotechnology, Havana, Cuba. luis.javier@cigb.edu.cu
Shimizu
T
T
Satomi
Y
Y
Betancourt
L
L
Besada
V
V
Padrón
G
G
Orlando
R
R
Shirasawa
T
T
Shimonishi
Y
Y
Takao
T
T
D000595:Amino Acid Sequence; D001224:Aspartic Acid / Q000032:analysis* / Q000737:chemistry; D006868:Hydrolysis; D008969:Molecular Sequence Data; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry; D021241:Spectrometry, Mass, Electrospray Ionization; D014357:Trypsin
D010455:Peptides; D001224:Aspartic Acid; D014357:Trypsin