Photodissociation Action Spectroscopy Research Articles

Charge-Tagged DNA Radicals in the Gas Phase Characterized by UV/Vis Photodissociation Action Spectroscopy.

Adenosine radicals tagged with a fixed-charge group were generated in the gas phase and structurally characterized by tandem mass spectrometry, deuterium labeling, and UV/Vis action spectroscopy. Experimental results in combination with Born-Oppenheimer molecular dynamics, ab initio, and excited-state calculations led to unambiguous assignment of adenosine radicals as N-7 hydrogen atom adducts. The charge-tagged radicals were found to be electronically equivalent to natural DNA nucleoside radicals.

Guanine-adenine interactions in DNA tetranucleotide cation radicals revealed by UV/vis photodissociation action spectroscopy and theory.

Hydrogen-rich cation radicals (GATT + 2H)+˙ and (AGTT + 2H)+˙ represent oligonucleotide models of charged hydrogen atom adducts to DNA. These tetranucleotide cation radicals were generated in the gas phase by one-electron reduction of the respective (GATT + 2H)2+ and (AGTT + 2H)2+ dications in which the charging protons were placed on the guanine and adenine nucleobases. We used wavelength-dependent UV/Vis photodissociation in the valence-electron excitation region of 210-700 nm to produce action spectra of (GATT + 2H)+˙ and (AGTT + 2H)+˙ that showed radical-associated absorption bands in the near-UV (330 nm) and visible (400-440 nm) regions. Born-Oppenheimer molecular dynamics and density-functional theory calculations were used to obtain and rank by energy multiple (GATT + 2H) dication and cation-radical structures. Time-dependent density functional theory (TD-DFT) calculations of excited-state energies and electronic transitions in (GATT + 2H)+˙ were augmented by vibronic spectra calculations at 310 K for selected low-energy cation radicals to provide a match with the action spectrum. The stable product of one-electron reduction was identified as having a 7,8-dihydroguanine cation radical moiety, formed by intramolecular hydrogen atom migration from adenine N-1-H. The hydrogen migration was calculated to have a transition state with a low activation energy, Ea = 96.5 kJ mol-1, and positive activation entropy, ΔS‡ = 75 J mol-1 K-1. This allowed for a fast isomerization of the primary reduction products on the ion-trap time scale of 150 ms that was substantially accelerated by highly exothermic electron transfer.

Color tuning of chlorophyll a and b pigments revealed from gas-phase spectroscopy

Chlorophyll (Chl) pigments are responsible for vital mechanisms in photosynthetic proteins: light harvesting, energy transfer and charge separation. A complex interplay between the Chl molecule and its microenvironment determines its transition energy. Interactions such as excitonic coupling with one or more pigments (Chls or carotenoids), axial ligation to the magnesium center, or electrostatic interactions between Chl and nearby amino-acid residues all influence the photophysical properties. Here we use time-resolved photodissociation action spectroscopy to determine transition energies of Chla/b complexes in vacuo to directly compare the impact of a negatively charged axial ligand (formate) to that of exciton coupling between two Chls. Experiments carried out at the electrostatic ion storage ring ELISA allow dissociation to be sampled on hundreds of milliseconds time scale. Absorption-band maxima of Chla-formate complexes are found at 433 ± 4 nm/2.86 ± 0.03 eV (Soret band) and in the region 654-675 nm/1.84-1.90 eV (Q band) and those of Chla dimers tagged by a quaternary ammonium ion at 419 ± 5 nm/2.96 ± 0.04 eV (Soret band) and 647 nm/1.92 eV (Q band). The axial ligand strongly affects the Chla transition energies causing redshifts of 0.21 eV of the Soret band and 0.04-0.1 eV of the Q band compared to Chla tagged by a quaternary ammonium. Slightly smaller shifts were found in case of Chlb. The redshifts are approximately twice that induced by excitonic coupling between two Chlas, also tagged by a quaternary ammonium ion. Axial ligation brings the absorption by isolated Chls very close to that of photosynthetic proteins.

UV-Vis Photodissociation Action Spectroscopy on Thermo LTQ-XL ETD and Bruker amaZon Ion Trap Mass Spectrometers: a Practical Guide.

We report automated procedures for multiple tandem mass spectra acquisition allowing UV-Vis photodissociation action spectroscopy measurements of ions and radicals. The procedures were developed for two commercial ion trap mass spectrometers and applied to collision-induced and electron-transfer dissociation tandem mass spectrometry modes of ion generation.

Selecting and identifying gas-phase protonation isomers of nicotineH+ using combined laser, ion mobility and mass spectrometry techniques.

The detection and assignment of protonation isomers, termed protomers, of gas-phase ions remains a challenge in mass spectrometry. With the emergence of ion-mobility techniques combined with tuneable-laser photodissociation spectroscopy, new experimental combinations are possible to now meet this challenge. In this paper, the differences in fragmentation and electronic spectroscopy of singly protonated (S)-nicotine (nicH+) ions are analysed using action spectroscopy in the ultraviolet region and field asymmetric ion mobility spectrometry (FAIMS). Experiments are supported by quantum chemical calculations (DFT, TD-DFT and CC2) of both spectroscopic and thermochemical properties. Electrospray ionisation (ESI) of (S)-nicotine from different solvents leads to different populations of two nicH+ protomers corresponding to protonation on the pyridine nitrogen and pyrrolidine nitrogen, respectively. FAIMS gives partial resolution of these protomers and enables characteristic product ions to be identified for each isomer as verified directly by analysis of product-ion specific action spectroscopy. It is shown that while characteristic, these product ions are not exclusive to each protomer. Calculations of vertical electronic transitions assist in rationalising the photodissociation action spectra. The integration of photodissociation action spectroscopy with FAIMS-mass spectrometry is anticipated to be a useful approach for separating and assigning protonation isomers of many other small molecular ions.

Near-UV Water Splitting by Cu, Ni, and Co Complexes in the Gas Phase.

(2,2'-Bipyridine)M═O+ ions (M = Cu, Ni, Co) were generated by collision-induced dissociation and near-UV photodissociation of readily available [(2,2'-bipyridine)MII(NO3)]+ ions in the gas phase, and their structure was confirmed by ion-molecule reactions combined with isotope labeling. Upon storage in a quadrupole ion trap, the (2,2'-bipyridine)M═O+ ions spontaneously added water, and the formed [(2,2'-bipyridine)M═O + H2O]+ complexes eliminated OH upon further near-UV photodissociation. This reaction sequence can be accomplished at a single laser wavelength in the range of 260-340 nm to achieve stoichiometric homolytic cleavage of gaseous water. Structures, spin states, and electronic excitations of the metal complexes were characterized by ion-molecule reactions using 2H and 18O labeling, photodissociation action spectroscopy, and density functional theory calculations.

Spontaneous Isomerization of Peptide Cation Radicals Following Electron Transfer Dissociation Revealed by UV-Vis Photodissociation Action Spectroscopy.

Peptide cation radicals of the z-type were produced by electron transfer dissociation (ETD) of peptide dications and studied by UV-Vis photodissociation (UVPD) action spectroscopy. Cation radicals containing the Asp (D), Asn (N), Glu (E), and Gln (Q) residues were found to spontaneously isomerize by hydrogen atom migrations upon ETD. Canonical N-terminal [z4 + H]+● fragment ion-radicals of the R-C●H-CONH- type, initially formed by N-Cα bond cleavage, were found to be minor components of the stable ion fraction. Vibronically broadened UV-Vis absorption spectra were calculated by time-dependent density functional theory for several [●DAAR + H]+ isomers and used to assign structures to the action spectra. The potential energy surface of [●DAAR + H]+ isomers was mapped by ab initio and density functional theory calculations that revealed multiple isomerization pathways by hydrogen atom migrations. The transition-state energies for the isomerizations were found to be lower than the dissociation thresholds, accounting for the isomerization in non-dissociating ions. The facile isomerization in [●XAAR + H]+ ions (X = D, N, E, and Q) was attributed to low-energy intermediates having the radical defect in the side chain that can promote hydrogen migration along backbone Cα positions. A similar side-chain mediated mechanism is suggested for the facile intermolecular hydrogen migration between the c- and [z + H]●-ETD fragments containing Asp, Asn, Glu, and Gln residues. Graphical Abstract ᅟ.

Elucidation of the intrinsic optical properties of hydrogen-bonded and protonated flavin chromophores by photodissociation action spectroscopy.

A model system of the flavin chromophore was synthesized and investigated for its intrinsic optical properties by gas phase action spectroscopy using an ion storage ring. An ammonium group was anchored to this flavin chromophore to allow its transfer to the gas phase by electrospray ionization and for studying the influence of hydrogen bonding and a nearby positive charge. According to calculations one of the hydrogen atoms of the ammonium group favorably forms an intramolecular ionic hydrogen bond to one of the oxygen atoms of the flavin chromophore, and this interaction was found to cause a blueshift of the S0 → S1 transition and a redshift of the S0 → S2 transition. For comparison, the S0 → S1 transition shows little solvent dependence (only in regard to the degree of fine structure). In addition, the influence of protonation of the flavin chromophore was elucidated by experimental and theoretical studies of a simple flavin system. While the position of the S0 → S1 absorption was at identical positions in the gas phase for the intramolecularly hydrogen-bonded and protonated flavin systems, the S0 → S2 absorption was further redshifted for the protonated species. This redshift resulting from protonation was also observed in solution.

Experimental Evidence for Noncanonical Thymine Cation Radicals in the Gas Phase.

Thymine cation radicals were generated in the gas phase by collision-induced intramolecular electron transfer in [Cu(2,2':6,2″-terpyridine)(thymine)]2+• complexes and characterized by ion-molecule reactions, UV-vis photodissociation action spectroscopy, and ab initio and density functional theory calculations. The experimental results indicated the formation of a tautomer mixture consisting chiefly (77%) of noncanonical tautomers with a C-7-H2 group. The canonical 2,4-dioxo-N-1,N-3-H isomer was formed as a minor component at ca. 23%. Ab initio CCSD(T) calculations indicated that the canonical [thymine]+• ion was not the lowest-energy isomer. This contrasts with neutral thymine, for which the canonical isomer is the lowest-energy structure. Exothermic unimolecular isomerization by a methyl hydrogen migration in the canonical [thymine]+• ion required a low energy barrier, forming a C-7-H2,O-4-H isomer. Noncanonical thymine tautomers with a C-7-H2 group were also identified by calculations as low-energy isomers of 2'-deoxythymidine phosphate cation radicals. The relative energies of thymidine ion isomers were sensitive to the computational method used and were affected by solvation. The noncanonical [thymine]+• ions have extremely low adiabatic recombination energies (REadiab < 5.9 eV), making them potential ionization hole traps in ionized nucleic acids.

UV-Vis Action Spectroscopy Reveals a Conformational Collapse in Hydrogen-Rich Dinucleotide Cation Radicals.

We report the generation of deoxyriboadenosine dinucleotide cation radicals by gas-phase electron transfer to dinucleotide dications and their noncovalent complexes with crown ether ligands. Stable dinucleotide cation radicals of a novel hydrogen-rich type were generated and characterized by tandem mass spectrometry and UV-vis photodissociation (UVPD) action spectroscopy. Electron structure theory analysis indicated that upon electron attachment the dinucleotide dications underwent a conformational collapse followed by intramolecular proton migrations between the nucleobases to give species whose calculated UV-vis absorption spectra matched the UVPD action spectra. Hydrogen-rich cation radicals generated from chimeric riboadenosine 5'-diesters gave UVPD action spectra that pointed to novel zwitterionic structures consisting of aromatic π-electron anion radicals intercalated between stacked positively charged adenine rings. Analogies with DNA ionization are discussed.

Radical Reactions Affecting Polar Groups in Threonine Peptide Ions.

Peptide cation-radicals containing the threonine residue undergo radical-induced dissociations upon collisional activation and photon absorption in the 210-400 nm range. Peptide cation-radicals containing a radical defect at the N-terminal residue, [•Ala-Thr-Ala-Arg+H]+, were generated by electron transfer dissociation (ETD) of peptide dications and characterized by UV-vis photodissociation action spectroscopy combined with time-dependent density functional theory (TD-DFT) calculations of absorption spectra, including thermal vibronic band broadening. The action spectrum of [•Ala-Thr-Ala-Arg+H]+ ions was indicative of the canonical structure of an N-terminally deaminated radical whereas isomeric structures differing in the position of the radical defect and amide bond geometry were excluded. This indicated that exothermic electron transfer to threonine peptide ions did not induce radical isomerizations in the fragment cation-radicals. Several isomeric structures, ion-molecule complexes, and transition states for isomerizations and dissociations were generated and analyzed by DFT and Møller-Plesset perturbational ab initio calculations to aid interpretation of the major dissociations by loss of water, hydroxyl radical, C3H6NO•, C3H7NO, and backbone cleavages. Born-Oppenheimer molecular dynamics (BOMD) in combination with DFT gradient geometry optimizations and intrinsic reaction coordinate analysis were used to search for low-energy cation-radical conformers and transition states. BOMD was also employed to analyze the reaction trajectory for loss of water from ion-molecule complexes.

Accessing the Intrinsic Nature of Electronic Transitions from Gas‐Phase Spectroscopy of Molecular Ion/Zwitterion Complexes

AbstractA molecule's color is governed by the nature of its electronic transitions. Herein we show that the degree of charge transfer can be assessed by measuring the change in absorption induced by complexation with the betaine zwitterion. Our approach benefits from direct formation of complexes by electrospray of a mixture solution, followed by photodissociation action spectroscopy. We explored two ion groups: 1) No permanent dipole moment due to even charge delocalization (e.g. MnO4−) and 2) Non‐even charge distribution but where the charge according to resonance forms is either delocalized (e.g. oxyluciferin) or located at one site (e.g. m‐nitrophenolate, mNP). The maximal shift for ions from (1) was &lt;0.05 eV but as large as 0.3 eV and 0.2 eV for mNP and oxyluciferin. Hence our work supports the notion that oxyluciferin undergoes CT, and that the microenvironment can account for large variation in light emission from insects, ranging from green to red (shift of 0.3 eV).

Accessing the Intrinsic Nature of Electronic Transitions from Gas-Phase Spectroscopy of Molecular Ion/Zwitterion Complexes.

A molecule's color is governed by the nature of its electronic transitions. Herein we show that the degree of charge transfer can be assessed by measuring the change in absorption induced by complexation with the betaine zwitterion. Our approach benefits from direct formation of complexes by electrospray of a mixture solution, followed by photodissociation action spectroscopy. We explored two ion groups: 1) No permanent dipole moment due to even charge delocalization (e.g. MnO4- ) and 2) Non-even charge distribution but where the charge according to resonance forms is either delocalized (e.g. oxyluciferin) or located at one site (e.g. m-nitrophenolate, mNP). The maximal shift for ions from (1) was <0.05 eV but as large as 0.3 eV and 0.2 eV for mNP and oxyluciferin. Hence our work supports the notion that oxyluciferin undergoes CT, and that the microenvironment can account for large variation in light emission from insects, ranging from green to red (shift of 0.3 eV).

Strong Impact of an Axial Ligand on the Absorption by Chlorophyll a and b Pigments Determined by Gas-Phase Ion Spectroscopy Experiments.

The microenvironments in photosynthetic proteins affect the absorption by chlorophyll (Chl) pigments. It is, however, a challenge to disentangle the impact on the transition energies of different perturbations, for example, the global electrostatics of the protein (nonbonded environmental effects), exciton coupling between Chl's, conformational variations, and binding of an axial ligand to the magnesium center. This is needed to distinguish between the two most commonly proposed mechanisms for energy transport in photosynthetic proteins, relying on either weakly or strongly coupled pigments. Here, on the basis of photodissociation action spectroscopy, we establish that the redshift of the Soret absorption band due to binding of a negatively charged carboxylate (as present in aspartic acid and glutamic acid residues) is 0.1-0.2 eV for Chl a and b. This effect is almost enough to reproduce the well-known green color of plants and can account for the strong spectral variation between Chl's. The experimental data serve to benchmark future high-level calculations of excited-state energies. Finally, we demonstrate that complexes between Chl a and histidine, tagged by a quaternary ammonium ion, can be made in the gas phase by electrospray ionization, but more work is needed to produce enough ions for gas-phase spectroscopy.

Where Does the Electron Go? Stable and Metastable Peptide Cation Radicals Formed by Electron Transfer.

Electron transfer to doubly and triply charged heptapeptide ions containing polar residues Arg, Lys, and Asp in combination with nonpolar Gly, Ala, and Pro or Leu generates stable and metastable charge-reduced ions, (M + 2H)+●, in addition to standard electron-transfer dissociation (ETD) fragment ions. The metastable (M + 2H)+● ions spontaneously dissociate upon resonant ejection from the linear ion trap, giving irregularly shaped peaks with offset m/z values. The fractions of stable and metastable (M + 2H)+● ions and their mass shifts depend on the presence of Pro-4 and Leu-4 residues in the peptides, with the Pro-4 sequences giving larger fractions of the stable ions while showing smaller mass shifts for the metastables. Conversion of the Asp and C-terminal carboxyl groups to methyl esters further lowers the charge-reduced ion stability. Collisional activation and photodissociation at 355 nm of mass-selected (M + 2H)+● results in different dissociations that give sequence specific MS3 spectra. With a single exception of charge-reduced (LKGLADR + 2H)+●, the MS3 spectra do not produce ETD sequence fragments of the c and z type. Hence, these (M + 2H)+● ions are covalent radicals, not ion-molecule complexes, undergoing dramatically different dissociations in the ground and excited electronic states. The increased stability of the Pro-4 containing (M + 2H)+● ions is attributed to radicals formed by opening of the Pro ring and undergoing further stabilization by hydrogen atom migrations. UV-VIS photodissociation action spectroscopy and time-dependent density functional theory calculations are used in a case in point study of the stable (LKGPADR + 2H)+● ion produced by ETD. In contrast to singly-reduced peptide ions, doubly reduced (M + 3H)+ ions are stable only when formed from the Pro-4 precursors and show all characteristics of even electron ions regarding no photon absorption at 355 nm or ion-molecule reactions, and exhibiting proton driven collision induced dissociations. Graphical Abstract ᅟ.

UV/Vis Action Spectroscopy and Structures of Tyrosine Peptide Cation Radicals in the Gas Phase

AbstractWe report the first application of UV/Vis photodissociation action spectroscopy for the structure elucidation of tyrosine peptide cation radicals produced by oxidative intramolecular electron transfer in gas‐phase metal complexes. Oxidation of Tyr‐Ala‐Ala‐Ala‐Arg (YAAAR) produces Tyr‐O radicals by combined electron and proton transfer involving the phenol and carboxyl groups. Oxidation of Ala‐Ala‐Ala‐Tyr‐Arg (AAAYR) produces a mixture of cation radicals involving electron abstraction from the Tyr phenol ring and N‐terminal amino group in combination with hydrogen‐atom transfer from the Cα positions of the peptide backbone.

UV/Vis Action Spectroscopy and Structures of Tyrosine Peptide Cation Radicals in the Gas Phase.

We report the first application of UV/Vis photodissociation action spectroscopy for the structure elucidation of tyrosine peptide cation radicals produced by oxidative intramolecular electron transfer in gas-phase metal complexes. Oxidation of Tyr-Ala-Ala-Ala-Arg (YAAAR) produces Tyr-O radicals by combined electron and proton transfer involving the phenol and carboxyl groups. Oxidation of Ala-Ala-Ala-Tyr-Arg (AAAYR) produces a mixture of cation radicals involving electron abstraction from the Tyr phenol ring and N-terminal amino group in combination with hydrogen-atom transfer from the Cα positions of the peptide backbone.

Combining UV photodissociation action spectroscopy with electron transfer dissociation for structure analysis of gas-phase peptide cation-radicals.

We report the first example of using ultraviolet (UV) photodissociation action spectroscopy for the investigation of gas-phase peptide cation-radicals produced by electron transfer dissociation. z-Type fragment ions (●) Gly-Gly-Lys(+), coordinated to 18-crown-6-ether (CE), are generated, selected by mass and photodissociated in the 200-400 nm region. The UVPD action spectra indicate the presence of valence-bond isomers differing in the position of the Cα radical defect, (α-Gly)-Gly-Lys(+) (CE), Gly-(α-Gly)-Lys(+) (CE) and Gly-Gly-(α-Lys(+))(CE). The isomers are readily distinguishable by UV absorption spectra obtained by time-dependent density functional theory (TD-DFT) calculations. In contrast, conformational isomers of these radical types are calculated to have similar UV spectra. UV photodissociation action spectroscopy represents a new tool for the investigation of transient intermediates of ion-electron reactions. Specifically, z-type cation radicals are shown to undergo spontaneous hydrogen atom migrations upon electron transfer dissociation.

Optical absorption spectrum of the chromium dimer cation: Measurements by photon-trap and photodissociation spectroscopy

Optical excitation in the chromium dimer cation, Cr2+, is investigated by photon-trap and photodissociation-action spectroscopy under a cryogenic condition in an ion trap. The former provides direct measurement of an absorption spectrum with high sensitivity, where cross sections as small as 0.4Å2 at the maximum are detectable. The dissociation cross sections are thus compared with those of absorption quantitatively. The spectra in the 2.2-eV region obtained by both methods are found to be equivalent. The present results are consistent with our assignment that the excited state is repulsive, in contrast to a previous report suggesting predissociation.

Moving in on the Action: An Experimental Comparison of Fluorescence Excitation and Photodissociation Action Spectroscopy.

Photodissociation action spectroscopy is often used as a proxy for measuring gas-phase absorption spectra of ions in a mass spectrometer. Although the potential discrepancy between linear optical and photodissociation spectra is generally acknowledged, direct experimental comparisons are lacking. In this work, we use a quadrupole ion trap that has been modified to enable both photodissociation and laser-induced fluorescence to assess how closely the visible photodissociation action spectrum of a fluorescent dye reflects its fluorescence excitation spectrum. Our results show the photodissociation action spectrum of gaseous rhodamine 110 is both substantially narrower and slightly red-shifted (∼120 cm(-1)) compared to its fluorescence excitation spectrum. Power dependence measurements reveal that the photodissociation of rhodamine 110 requires, on average, the absorption of three photons whereas fluorescence is a single-photon process. These differing power dependences are the key to interpreting the differences in the measured spectra. The experimental results provide much-needed quantification and insight into the differences between action spectra and linear optical spectra, and emphasize the utility of fluorescence excitation spectra to provide a more reliable benchmark for comparison with theory.