Investigation of the presence of b ions in electron capture dissociation mass spectra

Abstract

Previously, we have indicated (Cooper, H.J., et al. Int. J. Mass Spectrom., 2003, 228, 723-728) that electron capture dissociation (ECD) of the doubly protonated peptides, Leu 4 -Sar-Leu 3 -Lys-OH, Leu 4 -Ala-Leu 3 -Lys-OH, Gly 4 -Sar-Gly 3 -Lys-NH 2 , and Gly 3 -Pro-Sar-Gly 3 -Lys-NH 2 , results in abundant b ions, which derive from fragmentation of backbone amide bonds, a nonstandard fragmentation channel in ECD. The instrumental conditions were such that the possibility that collision-induced dissociation processes were contributing to the observed spectra was eliminated. In a separate study (Fung, Y.M.E., et al. Eur. J. Mass Spectrom., 2004, 10, 449 -457. ECD of peptides Arg-(Gly) n -Xxx-(Gly) n -Arg, where Xxx is the amino acid of interest, did not result in b ions. The variation in ECD observed for strikingly similar peptides suggests that the nature of the charge carrier (Arg or Lys) is instrumental in governing the fragmentation channels. Here, we describe the ECD behavior of a suite of model peptides designed such that the nature and position of the charge carrier could be probed. The results suggest that the presence of b ions in ECD spectra is a consequence of both charge carrier and peptide structure. The mechanism by which ECD of NOC␣ bonds in peptides and proteins occurs has been a matter of considerable debate [1, 5, 14 -23]. One proposed mechanism [1] involves capture of an electron by a charged site in the ion, for example, a protonated lysine or arginine residue, followed by H· transfer to (or recapture by) an amide carbonyl resulting in an aminoketyl radical. The radical subsequently dissociates via cleavage of the NOC␣ bond. Originally, it was suggested that c/z· fragment ion formation proceeded faster than vibrational energy redistribution, and that the process was nonergodic [1]. Recent work by Tureček suggests that NOC␣ bond dissociation is particularly facile in thermalized radicals and radical cations, that is, nonergodicity does not apply [21]. McLafferty and coworkers [24] showed that infrared multiphoton dissociation of thermalized [M ϩ 12H] 11ϩ· ions results in the dominant loss of H· atoms in marked contrast to the c/z ions produced following electron capture by [M ϩ 12H] 12ϩ ions to form [M ϩ 12H] 11ϩ· radical ions. These researchers concluded that ECD was nonergodic and the mechanism involves electron capture in a high-n Rydberg state followed by cooling to a dissociative electronic state. A direct mechanism has been suggested in which electron capture at C OH ϩ is accompanied by cleavage of the NOC␣ bond. Uggerud and coworkers [25,26] have shown theoretically and experimentally that both hydrogen transfer and direct processes play a role in ECD. Recently, Syrstad and Tureček [27] have proposed a mechanism for ECD that explains the low selectivity of NOC␣ cleavage and applies equally to peptide ions whose charge carriers are not protons. Either electron capture in a ground electronic state results in hydrogen atom transfer from the ammonium group to the amide carbonyl ...

Electron transfer dissociation of amide nitrogen methylated polypeptide cations

“…These ions match the predicted masses of b-ions. The presence of b ions in ETD/ECD spectra has been previously discussed [30,31]. However, in this case, a more likely origin of the b-ions is from collisional activation of the precursor ion upon injection into the collision cell.…”

Topoisomer Differentiation of Molecular Knots by FTICR MS: Lessons from Class II Lasso Peptides

“…Indeed, electron capture is expected to yield a distonic charge-reduced species [M + 3H]• 2+ that consecutively undergo c/z • cleavage. The formation of b-type product ions in ECD experiments has been reported for peptides containing or not basic residues [34][35][36][37]. They have been proposed to result from H• loss from the charge-reduced species and subsequent dissociation of the nonradical species by cleavage of the peptide backbone.…”

“…However, the formation of radical b' • through ECD of peptides has never been reported previously. The b'• series and bipeptic product ions permitted to propose a competitive fragmentation pattern involving two isomeric charge-reduced species [ [34][35][36][37]. In MccJ25 and MccJ25-lcm, the b-type product ions could also be generated from H• migration from the b'• series.…”

Electron Capture Dissociation of Hydrogen-Deficient Peptide Radical Cations

“…Although double resonance experiments with c-ion ejection eliminated the latter possibility (i.e., formation of b-ions was still observed after c-ion ejection in ECD of even electron precursor ions [48]) we cannot exclude this possibility in ECD of hydrogen-deficient precursor ions. Considering that the c-type product ions detected in ECD of hydrogen-deficient precursor ions were mainly radical species, we can postulate that they are more readily amenable to small neutral losses compared with even electron c-type product ions.…”

“…The majority of product ions detected for the even electron species were also present in the ECD spectrum of the hydrogen-deficient species, but their intensities were noticeably decreased. * = noise peak, @ = amino acid side chain losses In previous reports, the unexpected formation of b-ions in ECD of even electron precursor ions was rationalized by the presence of a vibrationally excited charge-reduced species [48][49][50][51]. These reports proposed that electron capture by the precursor ions followed by hydrogen atom loss leads to the formation of a vibrationally excited even electron species, [M + (n-1)H] (n-1)+ , which dissociates via the mobile proton pathway [48][49][50][51].…”