While the base (i.e., the chromophore) itself is not charged in its native state, a
nucleic acid can be easily charged by deprotonation of phosphate groups, producing
a nucleotide. In addition, protonated species can be produced at low pH values
(<4). The focus of this chapter is on ionic systems isolated in vacuo that can be
studied in experiments based on mass-spectrometry techniques. One motivation for
gas-phase investigations is to shed light on the electronic properties and the energytransfer processes in DNA at the molecular level in the absence of perturbations
from chemical environments (e.g., other biomolecules, solvent molecules or ions).
The effect of water molecules can then be disentangled through comparisons with
the behaviour of solutions. Also, since most excited-state calculations are
performed on isolated molecules, gas-phase reference data are needed as
benchmarks for electronic structure theory [12–16]. In particular, the description
of multibase systems is nontrivial due to the electronic couplings between two or
more bases in the excited state. Last but not least, gas-phase experiments on DNA
strands are important for the understanding of bioanalytical techniques such as
mass spectrometric sequencing of oligonucleotides. These techniques rely predominantly on collisional activation of electrosprayed DNA strands, but more recent
approaches combine UV excitation with collisions to obtain better sequence
coverage.
10.2 Photodissociation Mass Spectra and Fragmentation
Mechanisms
Nucleotide ions can be conveniently prepared in vacuo by both electrospray
ionisation (ESI) and matrix-assisted laser desorption/ionisation (MALDI). However, photodissociation experiments have been carried out mainly with ions starting
from ESI sources [17–32], both as native anions and as cations in the form of
protonated nucleic acids.
UV-excitation of DNA [26, 28] and RNA mononucleotides, abbreviated
[M–H]
À (M ¼ neutral mononucleotide, see also Table 10.1), as well as cyclic
derivatives [27, 28] leads to the same fragments as those found in collision-induced
dissociation (CID) experiments. The observed fragment ion masses are collected in
Table 10.1.
Fragment ion species can be grouped into two general classes according to the
primary fragmentation sites. The first group consists of products that are based on
breaking the phosphate-sugar link, which results in the formation of PO 3
À and
H 2 PO 4
À . The second class involves fragments formed after cleavage of the relatively weak CN glycosidic bond between the nucleobase and sugar subunits and a
number of subsequent reactions. Cleavage of this bond can result in formation of
the deprotonated base, B
À (B ¼ A, C, G, T), or an ion of the form [M–H–BH]
À that
corresponds to the loss of neutral, intact base (BH) from the parent mononucleotide.
Another fragment ion formed at lower abundance is the species [M–H–BH–H 2 O]
À ,
which is thought to arise from unimolecular dissociation of the primary fragment
ion [M–H–BH]
À [33].
10 UV Photophysics of DNA and RNA Nucleotides In Vacuo: Dissociation. . .
183
nucleic acid can be easily charged by deprotonation of phosphate groups, producing
a nucleotide. In addition, protonated species can be produced at low pH values
(<4). The focus of this chapter is on ionic systems isolated in vacuo that can be
studied in experiments based on mass-spectrometry techniques. One motivation for
gas-phase investigations is to shed light on the electronic properties and the energytransfer processes in DNA at the molecular level in the absence of perturbations
from chemical environments (e.g., other biomolecules, solvent molecules or ions).
The effect of water molecules can then be disentangled through comparisons with
the behaviour of solutions. Also, since most excited-state calculations are
performed on isolated molecules, gas-phase reference data are needed as
benchmarks for electronic structure theory [12–16]. In particular, the description
of multibase systems is nontrivial due to the electronic couplings between two or
more bases in the excited state. Last but not least, gas-phase experiments on DNA
strands are important for the understanding of bioanalytical techniques such as
mass spectrometric sequencing of oligonucleotides. These techniques rely predominantly on collisional activation of electrosprayed DNA strands, but more recent
approaches combine UV excitation with collisions to obtain better sequence
coverage.
10.2 Photodissociation Mass Spectra and Fragmentation
Mechanisms
Nucleotide ions can be conveniently prepared in vacuo by both electrospray
ionisation (ESI) and matrix-assisted laser desorption/ionisation (MALDI). However, photodissociation experiments have been carried out mainly with ions starting
from ESI sources [17–32], both as native anions and as cations in the form of
protonated nucleic acids.
UV-excitation of DNA [26, 28] and RNA mononucleotides, abbreviated
[M–H]
À (M ¼ neutral mononucleotide, see also Table 10.1), as well as cyclic
derivatives [27, 28] leads to the same fragments as those found in collision-induced
dissociation (CID) experiments. The observed fragment ion masses are collected in
Table 10.1.
Fragment ion species can be grouped into two general classes according to the
primary fragmentation sites. The first group consists of products that are based on
breaking the phosphate-sugar link, which results in the formation of PO 3
À and
H 2 PO 4
À . The second class involves fragments formed after cleavage of the relatively weak CN glycosidic bond between the nucleobase and sugar subunits and a
number of subsequent reactions. Cleavage of this bond can result in formation of
the deprotonated base, B
À (B ¼ A, C, G, T), or an ion of the form [M–H–BH]
À that
corresponds to the loss of neutral, intact base (BH) from the parent mononucleotide.
Another fragment ion formed at lower abundance is the species [M–H–BH–H 2 O]
À ,
which is thought to arise from unimolecular dissociation of the primary fragment
ion [M–H–BH]
À [33].
10 UV Photophysics of DNA and RNA Nucleotides In Vacuo: Dissociation. . .
183