Characterization of Oligodeoxynucleotides by Electron Detachment Dissociation Fourier Transform Ion Cyclotron Resonance Mass Spectrometry

Abstract

Electron detachment dissociation (EDD), recently introduced by Zubarev and co-workers for the dissociation of multiply charged biomolecular anions via a radical ion intermediate, has been shown to be analogous to electron capture dissociation (ECD) in several respects, including more random peptide fragmentation and retention of labile posttranslational modifications. We have previously demonstrated unique fragmentation behavior in ECD compared to vibrational excitation for oligodeoxynucleotide cations. However, that approach is limited by the poor sensitivity for oligonucleotide ionization in positive ion mode. Here, we show implementation of EDD on a commercial Fourier transform ion cyclotron resonance mass spectrometer utilizing two different configurations: a heated filament electron source and an indirectly heated hollow dispenser cathode electron source. The dispenser cathode configuration provides higher EDD efficiency and additional fragmentation channels for hexamer oligodeoxynucleotides. As in ECD, even-electron d/w ion series dominate the spectra, but we also detect numerous a/z (both even-electron and radical species), (a/z - B), c/x, (c/x - B), and (d/w - B) ions with minimal nucleobase loss from the precursor ions. In contrast to previous high-energy collision-activated dissociation (CAD) and ion trap CAD of radical oligonucleotide anions, we only observe minimum sugar cross-ring cleavage, possibly due to the short time scale of EDD, which limits secondary fragmentation. Thus, EDD provides fragmentation similar to ECD for oligodeoxynucleotides but at enhanced sensitivity. Finally, we show that noncovalent bonding in a DNA duplex can be preserved following EDD, illustrating another analogy with ECD. We believe the latter finding implies EDD has promise for characterization of nucleic acid structure and folding.

Electron photodetachment dissociation of DNA anions with covalently or noncovalently bound chromophores

“…Altogether, photodetachment of the DNA complexes upon chromophore excitation can be interpreted by the following mechanism: (1) ligands with sufficiently long excited-state lifetime undergo resonant two-photon excitation to reach the level of the DNA excited states, then (2) the excited-state must be coupled to the DNA excited states for photodetachment to occur. Our experiments also pave the way towards photodissociation probes of biomolecule conformation in the gas-phase by Since then, some major advances must be mentioned in electron activation methods [3][4][5] and in infrared photodissociation [6].…”

“…Altogether, photodetachment of the DNA complexes upon chromophore excitation can be interpreted by the following mechanism: (1) ligands with sufficiently long excited-state lifetime undergo resonant two-photon excitation to reach the level of the DNA excited states, then (2) the excited-state must be coupled to the DNA excited states for photodetachment to occur. Our experiments also pave the way towards photodissociation probes of biomolecule conformation in the gas-phase by Since then, some major advances must be mentioned in electron activation methods [3][4][5] and in infrared photodissociation [6].We recently started exploring the gas-phase reaction pathways of multiply charged DNA single strands and double strands upon UV irradiation around 260 nm [7,8]. To our surprise, we found out that, instead of fragmentation, electron detachment was the major reaction pathway with strands containing guanines.…”

“…Apart from the sequencing applications of EPD, numerous questions remain about the electron photodetachment mechanism, and how it compares with electron detachment dissociation [3][4][5] and thermal electron detachment [10,11]. We will briefly summarize our current understanding of the photodetachment mechanism [8].…”

“…The DNA fragmentation pathways resulting from the various activation methods have been reviewed in 2004 [2]. Since then, some major advances must be mentioned in electron activation methods [3][4][5] and in infrared photodissociation [6].…”

Electron detachment dissociation of neutral and sialylated oligosaccharides

“…In EDD, polyanions are irradiated with Ͼ10 eV electrons, resulting in electron detachment and subsequent product ions. EDD has been applied to peptides [45][46][47], oligonucleotides [48,49], gangliosides [50], and recently glycosaminoglycans (GAGs) [32,33]. GAGs are linear polysaccharides consisting of repeating disaccharide units and are frequently polysulfated.…”