Collision‐induced fragmentations of the (M‐H)⁻ parent anions of underivatized peptides: An aid to structure determination and some unusual negative ion cleavages

Abstract

This article describes the fundamental cleavage reactions of (M-H)(-) anions of underivatized peptides that contain up to 25 amino acid residues. The experimental observations of these cleavages have been backed up by molecular modeling, generally at the AM1 level of theory. The basic cleavages are the ubiquitous alpha- and beta-backbone cleavage reactions, which provide information similar to that of the B and Y + 2 cleavages of MH(+) ions of peptides. The residues Asp and Asn also effect cleavages of the backbone (called delta- and gamma-cleavages), by reactions initiated from side chain enolate anions, causing elimination reactions that cleave the backbone between the Asp (Asn) N bond;C backbone bond. Glu and Gln also direct analogous delta- and gamma-cleavages of the backbone, but in this case the processes are initiated by attack of the side chain CO(2) (-) (CONH(-)) to form a lactone (lactam). Ser and Thr residues undergo characteristic fragmentations of the side chain. These processes, losses of CH(2)O (Ser) and MeCHO (Thr), convert these residues into Gly. In larger peptides, Ser and Thr can effect two backbone cleavage reactions, called gamma- and epsilon -processes. The C-terminal CO(2) (-) (or CONH(-)) forms a hydrogen bond with the side chain OH (of Ser or Thr), placing the C-terminal residue in a position where it may affect S(N) (2) attack at the electrophilic backbone CH of Ser, with concomitant cleavage of the backbone. All of the above negative ion cleavages require the peptide backbone to be conformationally flexible. However, there is a backbone cleavage that requires the peptide to have an alpha-helical conformation in order for the two reacting centers to approach. This cleavage is illustrated for the Glu 23-initiated backbone cleavage at Ile 21 for the (M-H)(-) anion of the antimicrobial peptide caerin 1.1.

CαC Backbone Fragmentation Dominates in Electron Detachment Dissociation of Gas‐Phase Polypeptide Polyanions

“…[20] The evenelectron species [MÀH] À are likely to be responsible for the observed prominent water loss. [18] A radical-initiated mechanism of CO 2 loss from oxidized species, another prominent EDD feature, has been suggested elsewhere. [20] In comparison, CAD of the [MÀH] À ions led to abundant neutral losses (H 2 O and NH 3 ) from both fragments and precursor ions, but no loss of CO 2 .…”

“…Although a number of recent publications in the scientific literature have shown that useful sequence information can be derived from CAD mass spectra of negative polypeptide ions, [17,18] negative-ion fragmentation is still rarely used in practice. This is because the spectra obtained from negativeion MS/MS experiments are rife with ion peaks resulting from internal fragmentations, extensive side-chain losses, and neutral losses from fragments, and therefore these spectra are not always easy to interpret.…”

Charge inversion of polypeptide anions using protein and dendrimer cations as charge inversion reagents

“…It has been observed that fragmentation of ions of opposite polarity can provide complementary structural information [34]. Interrogating protein or peptide ions of both polarities is seldom performed, however, even when potentially valuable information might be obtained, presumably due to the additional time and experimental effort associated with analysis of the sample in both polarities.…”

Collision‐induced dissociation of the [M2H]2− ion of C‐peptide

“…4 In conclusion, guided by the study of Bowie's group on typical fragmentation patterns in negative ion spectra of peptides and proteins, we were able to elucidate the fragmentation pathway of C-peptide with negative ESI and CID monitoring of the doubly charged deprotonated molecule [ Letter to the Editor 3141…”