Side-chain involvement in the fragmentation reactions of the protonated methyl esters of histidine and its peptides

The fragmentation reactions of the protonated dipeptides Gly-Arg and Arg-Gly have been studied using collision-induced dissociation (CID) in a quadrupole ion trap, by in-source CID in a single-quadrupole mass spectrometer and by CID in the quadrupole cell of a QqTOF mass spectrometer. In agreement with earlier quadrupole ion trap studies (Farrugia, J. M.; O'Hair, R. A. J., Int. J. Mass Spectrom., 2003, 222, 229), the CID mass spectra obtained with the ion trap for the MH ϩ ions and major fragment ions are very similar for the two isomers indicating rearrangement to a common structure before fragmentation. In contrast, in-source CID of the MH ϩ ions and QqTOF CID of the MH ϩ , [MH Ϫ NH 3 ] ϩ and [MH Ͻ23 HN ϭ C(NH 2 ) 2 ] ϩ ions provide distinctly different spectra for the isomeric dipeptides, indicating that rearrangement to a common structure has not occurred to a significant extent under these conditions even near the threshold for fragmentation in the QqTOF instrument. Clearly, under normal operating conditions significantly different fragmentation behavior is observed in the ion trap and beam-type experiments. This different behavior probably can be attributed to the shorter observation times and concomitant higher excitation energies in the in-source and QqTOF experiments compared to the long observation times and lower excitation energies relevant to the ion trap experiments. Based largely on elemental compositions derived from accurate mass measurements in QqTOF studies fragmentation schemes are proposed for the MH ϩ , [MH Ϫ

Section: Resultsmentioning

confidence: 89%

Fragmentation of protonated dipeptides containing arginine. Effect of activation method

Forbes

Jockusch

Young

et al. 2007

“…A plausible mechanism that explains the observed experimental behavior includes proton transfer to the C-terminal adjacent amide N and subsequent nucleophilic attack by the His side-chain on the carbon center of the protonated amide bond [13]. O'Hair in another study [25] demonstrated that CID of the b 2 ion of Gly-His and His-Gly produces the same fragmentation pattern that parallels that of the diketopiperazine derivative cyclo-GH.…”

mentioning

confidence: 86%

“…The amide bond cleavage pathway initiated by the His side-chain (referred to as His-b n -y m ) was studied in detail by Wysocki, O'Hair, and coworkers [11,25,26]. Based on these investigations, we proposed a mechanism that involves transfer of the extra ionizing proton from the His side-chain to the C-terminal adjacent amide nitrogen and subsequent nucleophilic attack of the imidazole nitrogen on the carbon of the protonated amide bond [13].…”

Section: His Side-chain Initiated Amide Bond Cleavage: Formation Of Bmentioning

confidence: 99%

“…Fragmentation of histidine containing peptides attracted some attention [11,25,26] in the past, still many aspects of the related dissociation chemistry are not clarified yet. The histidine side chain has a relatively high proton affinity [27,28] and, therefore, it is likely to be protonated if the number of added protons is higher than the number of arginines and lysines present in the peptide ion under study.…”

mentioning

confidence: 99%

See 1 more Smart Citation

Fragmentation of doubly-protonated Pro-His-Xaa tripeptides: Formation of b₂²⁺ ions

Knapp-Mohammady

Young

Paizs

et al. 2009

When ionized by electrospray from acidic solutions, the tripeptides Pro-His-Xaa (Xaa ϭ Gly, Ala, Leu) form abundant doubly-protonated ions, [M ϩ 2H] 2ϩ . Collision-induced dissociation (CID) of these doubly-protonated species results, in part, in formation of b 2 2ϩ ions, which fragment further by loss of CO to form a 2 2ϩ ions; the latter fragment by loss of CO to form the Pro and His iminium [immonium is commonly used in peptide MS work] ions. Although larger doubly-charged b ions are known, this represents the first detailed study of b 2 2ϩ ions in CID of small doubly protonated peptides. The most abundant CID products of the studied doubly-protonated peptides arise mainly in charge separation involving two primary fragmentation channels, formation of the b 2 /y 1 pair and formation of the a 1 /y 2 pair. Combined molecular dynamics and density functional theory calculations are used to gain insight into the structures and fragmentation pathways of doubly-protonated Pro-His-Gly including the energetics of potential protonation sites, backbone cleavages, post-cleavage charge-separation reactions and the isomeric structures of b 2 2ϩ ions. Three possible structures are considered for the b 2 2ϩ ions: the oxazolone, diketopiperazine, and fused ring isomers. The last is formed by cleavage of the His-Gly amide bond on a pathway that is initiated by nucleophilic attack of one of the His side-chain imidazole nitrogens. Our calculations indicate the b 2 2ϩ ion population is dominated by the oxazolone and/or fused ring isomers. (J Am Soc Mass Spectrom 2009, 20, 2135-2143

“…Other alternative ion structures can be derived if His, Arg, Asp, or Lys occupies the C-terminal position in b ions [23][24][25][26]. In these cases the side-chain nucleophiles of the preceding amino acids are responsible for cleavage of the amide bond or reisomerization of the primarily formed oxazolone ring.…”

mentioning

confidence: 99%

Structure and reactivity of a_n and a*_n peptide fragments investigated using isotope labeling, tandem mass spectrometry, and density functional theory calculations

Bythell

Molesworth

Osburn

et al. 2008

Extensive 15 N labeling and multiple-stage tandem mass spectrometry were used to investigate the fragmentation pathways of the model peptide FGGFL during low-energy collisioninduced-dissociation (CID) in an ion-trap mass spectrometer. Of particular interest was formation of a 4 from b 4 and a 4 * (a 4 -NH 3 ) from a 4 ions correspondingly, and apparent rearrangement and scrambling of peptide sequence during CID. It is suggested that the original FGGF oxa b 4 structure undergoes b-type scrambling to form GGFF oxa . These two isomers fragment further by elimination of CO and 14 NH 3 or 15 NH 3 to form the corresponding a 4 and a 4 * isomers, respectively. For ( 15 N-F)GGFL and FGG( 15 N-F)L the a 4 * ion population appears as two distinct peaks separated by 1 mass unit. These two peaks could be separated and fragmented individually in subsequent CID stages to provide a useful tool for exploration of potential mechanisms along the a 4 ¡ a 4 * pathway reported previously in the literature . In most MS/MS experiments, protonated peptides are excited collisionally to induce dissociation (collision-induced-dissociation, CID) and the fragment ion spectrum is used to elucidate peptide sequences. The CID spectra of peptides in proteomics studies are commonly assigned by bioinformatics tools that implement sequencing algorithms and peptide fragmentation models. Regrettably, the existing sequencing programs are based on rather limited fragmentation models that poorly approximate the rich dissociation chemistry of protonated peptides [3]. These limitations often lead to erroneous assignment of peptides and proteins, and the resulting uncertainty in the evaluation of the raw MS/MS data is one of the major limiting factors in large-scale protein identification studies [4,5]. The incorporation of more detailed peptide fragmentation mechanisms and spectral characteristics into these sequencing algorithms would undoubtedly place MS/MS based sequencing on a much more robust basis.In general, fragmentation of protonated peptides under low-energy collision conditions involves protondriven reactions in which amide bonds are cleaved along the peptide backbone and b, y, and a ions [6,7] are formed. The energetics and kinetics of the necessary proton mobilization (mobile proton model [8,9]) and amide bond cleavage pathways [3, 10 -14] have received significant research interest. On the other hand, much less attention has been devoted to the structure and reactivity of the primary fragments formed by backbone cleavages. According to the recent pathways in competition (PIC) fragmentation model [3], the thermodynamic properties and the reactivity of these fragAddress reprint requests to Dr. M. Van Stipdonk,