Dissociation of multiply charged negative ions for hirudin (54–65), fibrinopeptide B, and insulin A (oxidized)

Abstract

Collision-induced dissociation (CID) was performed on multiply deprotonated ions from three commercial peptides: hirudin (54-65), fibrinopeptide B, and oxidized insulin chain A. Ions were produced by electrospray ionization in a Fourier transform ion cyclotron resonance mass spectrometer. Each of these peptides contains multiple acidic residues, which makes them very difficult to ionize in the positive mode. However, the peptides deprotonate readily making negative ion studies a viable alternative. The CID spectra indicated that the likely deprotonation sites are acidic residues (aspartic, glutamic, and cysteic acids) and the C-terminus. The spectra are rife with c, y, and internal ions, although some a, b, x, and z ions form. Many of the fragment ions were formed from cleavage adjacent to acidic residues, both N- and C-terminal to the acidic site. In addition, neutral loss (e.g., NH3, CH3, H2O, and CO2) was prevalent from both the parent ions and from fragment ions. These neutral eliminations were often indicative of specific amino acid residues. The fragmentation patterns from several charge states of the parent ions, when combined, provide significant primary sequence information. These results suggest that negative mode CID of multiply deprotonated ions provides useful structural information and can be worthwhile for highly acidic peptides that do not form positive ions in abundance.

Characterization of alkali induced formation of lanthionine, trisulfides, and tetrasulfides from peptide disulfides using negative ion mass spectrometry

“…Abstraction of the C ␣ -hydrogen results in formation of a peptide enolate at Cys residues, which can subsequently cleave to yield dehydroalanine residue with loss of H 2 S or H 2 S 2 . The use of negative ion mass spectrometry [13][14][15][16][17][18][19][20][21][22][23][24][25][26][27][28][29] for the study of disulfide containing peptide natural products has been established in an extensive series of investigations by Bowie and coworkers [30 -35] and other groups [36 -38]. During the course of recent attempts to fragment the negative ions of disulfide bridged peptides under mass spectrometric conditions, we noted that the cleavage reactions closely resemble those observed during breakage of disulfide bonds under alkaline conditions [39,40].…”

A Comparison of the Effects of Amide and Acid Groups at the C-Terminus on the Collision-Induced Dissociation of Deprotonated Peptides

“…Penta-alanine also produces a n − and ″b n − , which is typical of oligoalanine peptides in the negative ion mode [41]. Neutral loss peaks common to specific amino acid residues are abundant in the spectra; for example, elimination of H 2 O and CH 2 O from serine [42][43][44][45][46][47][48][49], the guanidino group (HN=C=NH) from arginine [40,49,50], H 2 O from aspartic acid [51][52][53][54], O=C 6 H 4 =CH 2 from tyrosine [54], and CH 3 SH, CH 3 SCH 3 , and CH 2 CH 2 SCH 3 from methionine [55].…”

Electron Induced Dissociation of Singly Deprotonated Peptides

“…In EID, only the c 9 product ion was observed. Formation of c-type product ions is generally observed in negative ion mode CAD [21,29,60]. Some y-and b-type ions, y 4 , b 4 , and b 7 , observed in CAD were absent in EID.…”

“…However, utilization of negative ion mode is often desired, given that~50% of naturally occurring proteins are acidic. Peptides containing numerous acidic residues (e.g., glutamic and aspartic acid) may be challenging to detect in positive ion mode [21][22][23]. On the other hand, it has been shown that highly basic peptides can easily form singly deprotonated ions with sufficient intensities for fragmentation experiments [24].…”