Electron Transfer Reduction of the Diazirine Ring in Gas-Phase Peptide Ions. On the Peculiar Loss of [NH₄O] from Photoleucine

Abstract

Abstract. Electron transfer to gas-phase peptide ions with diazirine-containing amino acid residue photoleucine (L*) triggers diazirine ring reduction followed by cascades of residue-specific radical reactions. Upon electron transfer, substantial fractions of (GL*GGR +2H) +• cation-radicals undergo elimination of [NH O-labeled peptide ions and found to specifically involve the amide oxygen of the N-terminal residue. The structures, energies, and electronic states of the peptide radical species were elucidated by a combination of near-UV photodissociation experiments and electron structure calculations combining ab initio and density functional theory methods. Electron transfer reaching the ground electronic states of charge reduced (GL*GGR +2H) +• cation-radicals was found to reduce the diazirine ring. In contrast, backbone N−C α bond dissociations that represent a 60%-75% majority of all dissociations because of electron transfer are predicted to occur from excited electronic states.

Combining Near-UV Photodissociation with Electron Transfer. Reduction of the Diazirine Ring in a Photomethionine-Labeled Peptide Ion

“…The m/z 415 ion relative intensity amounted to 25% of combined relative intensities of z 1 -z 4 ions and the m/z 469 (loss of ammonia) fragment ion. The ETD spectrum of (GM*GGR + 2H) 2+ displayed only very minor peaks of fragment ions at m/z 454 (loss of N 2 H 4 ) and m/z 452 (loss of NH 4 O) that were both prominent in the ETD mass spectra of L*-containing peptide ions [7,8]. Hence, a minor structure change of replacing L* with M* had a major effect on ETD.…”

“…To accomplish photodissociation of trapped ions in the LIT, the LTQ-XL ETD mass spectrometer was modified as described previously [5,8]. The typical light intensity used in the photodissociation experiments was 15-20 mJ/pulse.…”

“…The Figure 1a The low abundance of the long-lived m/z 486 (GM*GGR + 2H) +• cation radical in the Figure 1a ETD spectrum was an impediment to further experimental studies of this ion's structure and electronic properties. To further elucidate the nature of the m/z 486 ion, we employed our previously introduced method of selective cation radical preparation using noncovalent complexes with 18-crown-6-ether (CE) [7,8]. ETD of the (GM*GGR + CE + 2H) 2+ complex (m/z 375) resulted in charge reduction accompanied by an abundant loss of the CE molecule, forming the (GM*GGR + 2H) +• cation-radical at m/z 486 (Figure 2a), which was characterized by accurate mass measurements (Table S1, Supplementary Data).…”

“…ETD of the (GM*GGR + CE + 2H) 2+ complex (m/z 375) resulted in charge reduction accompanied by an abundant loss of the CE molecule, forming the (GM*GGR + 2H) +• cation-radical at m/z 486 (Figure 2a), which was characterized by accurate mass measurements (Table S1, Supplementary Data). It should be noted that although ETD has sometimes been presented as a method that preserves noncovalent interactions [28], the ETD spectra of this and several other peptide-CE complexes [5,8,29,30] attest to the opposite. The (GM*GGR + 2H) +• ion was selected by mass and subjected to CID ( Figure 2b) and UVPD ( Figure 2c).…”

“…Photodissociation of L* and M* is analogous to the general behavior of aliphatic diazirines in that it results in N 2 elimination, creating highly reactive carbenes positioned in the amino acid side chains that undergo competing insertions into X-H bonds and isomerization to nonreactive olefins [4]. In our previous studies of photodissociation [5], collision-induced dissociation [6], and electron transfer dissociation [7,8] of model peptide ions containing photoleucine, we discovered several unusual reactions involving the diazirine ring. In particular, ETD of L*-containing peptide ions (Scheme 1, n = 1) resulted in diazirine ring fission and losses of small molecules such as N 2 H 4 , N 2 H 3 , and [NH 4 O] that competed with backbone dissociations.…”