Guanine Radical Cation Research Articles

Nitrosation mechanisms, kinetics, and dynamics of the guanine and 9-methylguanine radical cations by nitric oxide-Radical-radical combination at different electron configurations.

As a precursor to various reactive nitrogen species formed in biological systems, nitric oxide (•NO) participates in numerous processes, including enhancing DNA radiosensitivity in ionizing radiation-based radiotherapy. Forming guanine radical cations is another common DNA lesion resulting from ionization and oxidation damage. As such, the interaction of •NO with guanine radical cations (G•+) may contribute to the radiosensitization of •NO. An intriguing aspect of this process is the participation of multiple spin configurations in the reaction, including open-shell singlet 1,OS[G•+(↑)⋯(↓)•NO], closed-shell singlet 1,CS[G(↑↓)⋯NO+], and triplet 3[G•+(↑)⋯(↑)•NO]. In this study, the reactions of •NO with both unsubstituted guanine radical cations (in the 9HG•+ conformation) and 9-methylguanine radical cations (9MG•+, a guanosine-mimicking model compound) were investigated in the absence and presence of monohydration of radical cations. Kinetic-energy dependent reaction product ions and cross sections were measured using an electrospray ionization guided-ion beam tandem mass spectrometer. The reaction mechanisms, kinetics, and dynamics were comprehended by interpreting the reaction potential energy surface using spin-projected density functional theory, coupled cluster theory, and multiconfiguration complete active space second-order perturbation theory, followed by RRKM kinetics modeling. The combined experimental and computational findings revealed closed-shell singlet 1,CS[7-NO-9MG]+ as the major, exothermic product and triplet 3[8-NO-9MG]+ as the minor, endothermic product. Singlet biradical products were not detected due to high reaction endothermicities, activation barriers, and inherent instability.

Contrasting Photosensitized Processes of Ru(II) Polypyridyl Structural Isomers Containing Linear and Hooked Intercalating Ligands Bound to Guanine-Rich DNA.

The DNA binding and cellular uptake of the lambda enantiomer of two bis-tetraazaphenanthrene (TAP) Ru(II) polypyridyl complexes containing either a linear dppn (1) or a hooked bdppz (2) benzodipyridophenazine ligand are reported, and the role of different charge-transfer states of the structural isomers in the photo-oxidation of guanine is explored. Both complexes possess characteristic metal-to-ligand charge-transfer (MLCT) bands between 400 and 500 nm and emission at ca. 630 nm in an aerated aqueous solution. Transient visible absorption (TrA) spectroscopy reveals that 400 nm excitation of 1 yields a dppn-based metal-to-ligand charge-transfer (MLCT) state, which in turn populates a dppn intraligand (3IL) state. In contrast, photoexcitation of 2 results in an MLCT state on the TAP ligand and not the intercalating bdppz ligand. Both 1 and 2 bind strongly to double-stranded guanine-rich DNA with a loss of emission. Combined TrA and time-resolved infrared (TRIR) spectroscopy confirms formation of the guanine radical cation when 2 is bound to the d(G5C5)2 duplex, which is not the case when 1 is bound to the same duplex and indicates a different mechanism of action in DNA. Utilizing the long-lived triplet excited lifetime, we show good uptake and localization of 2 in live cells as well as isolated chromosomes. The observed shortening of the excited-state lifetime of 2 when internalized in cell chromosomes is consistent with DNA binding and luminescent quenching due to guanine photo-oxidation.

What tunes guanine ionization potential in a nucleosome? An all-in-one systematic QM/MM assessment

Guanine radical cations are precursors to oxidatively induced DNA lesions, and the determination of oxidative DNA hot spots beyond oligonucleotides remains a current challenge. In order to rationalize the finetuned ionization properties of the ∼60 guanines in a nucleosome core particle, we report a robust molecular dynamics-then-FO-DFTB/MM (fragment-orbital tight-binding density functional theory/molecular mechanics) simulation protocol spanning 20 μs. Our work allows us to identify several factors governing guanine ionization potential and map oxidative hotspots. Our results highlight the predominant role of the proximity of positively charged histone residues in the modulation of the guanine ionization potential up to 0.6 eV. Consequently, fast, long-range hole transfer in nucleosomal DNA could be tuned by the proximity of histone tails, which differs, from a biological point of view, on the chromatin state.

Spin-orbit charge transfer from guanine and 9-methylguanine radical cations to nitric oxide radicals and the induced triplet-to-singlet intersystem crossing.

Nitric oxide (●NO) participates in many biological activities, including enhancing DNA radiosensitivity in ionizing radiation-based radiotherapy. To help understand the radiosensitization of ●NO, we report reaction dynamics between ●NO and the radical cations of guanine (a 9HG●+ conformer) and 9-methylguanine (9MG●+). On the basis of the formation of 9HG●+ and 9MG●+ in the gas phase and the collisions of the radical cations with ●NO in a guided-ion beam mass spectrometer, the charge transfer reactions of 9HG●+ and 9MG●+ with ●NO were examined. For both reactions, the kinetic energy-dependent product ion cross sections revealed a threshold energy that is 0.24 (or 0.37) eV above the 0K product 9HG (or 9MG) + NO+ asymptote. To interrogate this abnormal threshold behavior, the reaction potential energy surface for [9MG + NO]+ was mapped out at closed-shell singlet, open-shell singlet, and triplet states using density functional and coupled cluster theories. The results showed that the charge transfer reaction requires the interaction of a triplet-state surface originating from a reactant-like precursor complex 3[9MG●+(↑)⋅(↑)●NO] with a closed-shell singlet-state surface evolving from a charge-transferred complex 1[9MG⋅NO+]. During the reaction, an electron is transferred from π∗(NO) to perpendicular π∗(9MG), which introduces a change in orbital angular momentum. The latter offsets the change in electron spin angular momentum and facilitates intersystem crossing. The reaction threshold in excess of the 0K thermochemistry and the low charge-transfer efficiency are rationalized by the vibrational excitation in the product ion NO+ and the kinetic shift arising from a long-lived triplet intermediate.

Radical Cascade Dissociation Pathways to Unusual Nucleobase Cation Radicals.

We report unusual dissociations of protonated RNA nucleosides tagged with radical initiator groups at ribose 5'-O and furnished with a 2',3'-O-isopropylidene protecting group. The ions undergo collision-induced radical cascade dissociations starting at the radical initiator that break down the dioxolane ring and trigger the formation of nucleobase cations and cation radicals. The adenine cation radical that was formed by radical cascade dissociations was identified by MS5 UV-vis photodissociation action spectroscopy to be a higher-energy N-3-H tautomer of the canonical ionized nucleobase. The guanine cation radical was formed by radical cascade dissociations as the N-7-H tautomer. In contrast to adenosine and guanosine, radical cascade dissociations of the tagged ribocytidine ion produced protonated cytosine, whereas tagged ribothymidine showed yet different dissociations resulting in predominant thymine loss. Reaction mechanisms were suggested for the cascade dissociations that were based on Born-Oppenheimer molecular dynamics and density functional theory calculations that were used to map the relevant parts of the potential energy surfaces for adenosine, guanosine, and cytidine radical ions. The reported radical cascade dissociations represent a new, nonredox approach to nucleobase and nucleoside cation radicals that has the potential of being expanded to the generation of various oligonucleotide cation radicals.

Interstrand Crosslinking Involving Guanine: A New Major UVC Laser-Induced Biphotonic Oxidatively Generated DNA Damage.

Several classes of oxidatively generated DNA damage including oxidized purine and pyrimidine bases, interstrand base crosslinks and DNA-protein crosslinks have been previously shown to be generated in both isolated DNA and cellular DNA upon exposure to either 266-nm laser irradiation or one-electron oxidants. In this study, we provide evidence that biphotonic ionization of guanine bases by UVC laser irradiation of double-stranded deoxyoligonucleotides in aerated aqueous solutions induces the formation of interstrand crosslinks (ICLs). This is supported by various experiments including sequencing gel analyses of formed photoproducts and effects of UVC laser intensity on their formation. This constitutes a novel example of the diversity of reactions of guanine radical cation that can be generated by various one-electron oxidants including UVC laser biphotonic ionization, direct effect of ionization radiation and type I photosensitizers. However, the exact structure of the interstrand base adducts that is a challenging analytical issue remains to be further established. Examples of relevant biochemical/structural applications of biphotonic induction of ICLs in DNA samples by high-intensity UVC laser pulses are provided.

Photo-induced telomeric DNA damage in human cancer cells.

Herein we report on the study of novel dinuclear ruthenium(ii) complexes designed to target and to photo-react with G-quadruplex telomeric DNA. Upon irradiation, complexes efficiently generate guanine radical cation sites as photo-oxidation products. The compounds also display efficient cell penetration with localization to the nucleus and show strong photocytotoxicity toward osteosarcoma cells. Thanks to a microscopic-based telomere dysfunction assay, which allows the direct visualization of DNA damage in cells, we brought the first evidence of forming photo-oxidative damage at telomeres in cellulo. This emphasizes interesting prospects for the development of future cancer phototherapies.

Early steps of oxidative damage in DNA quadruplexes are position-dependent: Quantum mechanical and molecular dynamics analysis of human telomeric sequence containing ionized guanine

Guanine radical cation (G•+) is a key intermediate in many oxidative processes occurring in nucleic acids. Here, by combining mixed Quantum Mechanical/Molecular Mechanics calculations and Molecular Dynamics (MD) simulations, we study how the structural behaviour of a tract GGG(TTAGGG)3 (hereafter Tel21) of the human telomeric sequence, folded in an antiparallel quadruple helix, changes when one of the G bases is ionized to G•+ (Tel21+). Once assessed that the electron-hole is localized on a single G, we perform MD simulations of twelve Tel21+ systems, differing in the position of G•+ in the sequence. When G•+ is located in the tetrad adjacent to the diagonal loop, we observe substantial structural rearrangements, which can decrease the electrostatic repulsion with the inner Na+ ions and increase the solvent exposed surface of G•+. Analysis of solvation patterns of G•+ provides new insights on the main reactions of G•+, i.e. the deprotonation at two different sites and hydration at the C8 atom, the first steps of the processes producing 8oxo-Guanine. We suggest the main structural determinants of the relative reactivity of each position and our conclusions, consistent with the available experimental trends, can help rationalizing the reactivity of other G-quadruplex topologies.

Deprotonation Dynamics of Guanine Radical Cations.

This review is dedicated to guanine radical cations (G+ )· that are precursors to oxidatively generated damage to DNA. (G+ )· are unstable in neutral aqueous solution and tend to lose a proton. The deprotonation process has been studied by time-resolved absorption experiments in which (G+ )· radicals are produced either by an electron abstraction reaction, using an external oxidant, or by low-energy/low-intensity photoionization of DNA. Both the position of the released proton and the dynamics of the process depend on the secondary DNA structure. While deprotonation in duplex DNA leads to (G-H1)· radicals, in guanine quadruplexes the (G-H2)· analogs are observed. Deprotonation in monomeric guanosine proceeds with a time constant of ~60 ns; in genomic DNA, it is completed within 2 µs; and in guanine quadruplexes, it spans from at least 30 ns to over 50 µs. Such a deprotonation dynamics in four-stranded structures, extended over more than three decades of times, is correlated with the anisotropic structure of DNA and the mobility of its hydration shell. In this case, commonly used second-order reaction models are inappropriate for its description.

Dynamics and Multiconfiguration Potential Energy Surface for the Singlet O2 Reactions with Radical Cations of Guanine, 9-Methylguanine, 2'-Deoxyguanosine, and Guanosine.

Reactions of electronically excited singlet oxygen (1O2) with the radical cations of guanine (9HG•+), 9-methylguanine (9MG•+), 2'-deoxyguanosine (dGuo•+), and guanosine (Guo•+) were studied in the gas phase by a combination of guided-ion-beam mass spectrometric measurement of product ions and cross sections as a function of collision energy (Ecol) and electronic structure calculations of the reaction potential energy surface (PES) at various levels of theory. No product could be captured in the 1O2 reaction with bare 9HG•+ or 9MG•+, because energized products decayed rapidly to reactants before being detected. To overcome this unfavorable kinetics, monohydrated 9HG•+·H2O and 9MG•+·H2O were used as reactant ions, of which the peroxide product ions were stabilized by energy relaxation via elimination of the water ligand. Reaction cross sections for 9HG•+·H2O and 9MG•+·H2O decrease with increasing Ecol, becoming negligible above 0.6 eV. This indicates that the reactions are exothermic with no barriers above reactants and the heat of formation of the products is sufficiently large to overcome their water ligand elimination energy (0.7 eV). Peroxide product ions were also detected in the 1O2 reactions with unhydrated dGuo•+ and Guo•+, in which intramolecular vibrational redistribution was able to stabilize oxidation products. 9MG•+ was utilized as a model system to explore the reaction PES for the initial 1O2 addition to the guanine radical cation. Calculations were carried out using single-reference ωB97XD, RI-MP2, and DLPNO-CCSD(T) and multireference CASSCF and CASPT2. Although the same PES profile was obtained at different levels of theory, the energies of the mixed open- and closed-shell 1O2 reactant and the open-shell reaction intermediates, transition states, and products are sensitive to the theories. By taking into account both static and dynamic electron correlations, the CASPT2 PES has provided the best agreement with the experimentally measured reaction thermodynamics and predicted 8-peroxide as the most probable initial oxidation product of the guanine radical cation.

Reaction mechanism and dynamics for C8-hydroxylation of 9-methylguanine radical cation by water molecules.

In contrast to their spontaneous deprotonation in aqueous solution, reactions of guanine and guanosine radical cations with water in the gas phase are exclusively initiated by hydration of the radical cations as reported in recent work (Y. Sun et al., Phys. Chem. Chem. Phys., 2018, 20, 27510). As gas-phase hydration reactions closely mimic the actual scenario for guanine radical cations in double-stranded DNA, exploration of subsequent reactions within their water complexes can provide an insight into the resulting oxidative damage to nucleosides. Herein guided-ion beam mass spectrometry experiment and direct dynamics trajectory simulations were carried out to examine prototype complexes of the 9-methylguanine radical cation with one and two water ligands (i.e., 9MG˙+·(H2O)1-2) in the gas phase, wherein the complexes were activated by collisional activation in the experiment and by thermal excitation at high temperatures in the simulations. Guided by mass spectroscopic measurements, trajectory results and reaction potential energy surface, three reaction pathways were identified. The first two reaction pathways start with H-atom abstraction from water by the O6 and N7 atoms in 9MG˙+ and are referred to as HAO6 and HAN7, respectively. The primary products of HAO6 and HAN7 reactions, including [9MG + HO6]+/[9MG + HN7]+ and ˙OH, react further to either form [8OH-9MG + HO6]˙+ and [8OH-9MG + HN7]˙+via C8-hydroxylation or form radical cations of 6-enol-guanine (6-enol-G˙+) and 7H-guanine (7HG˙+) via SN2-type methanol elimination. The third reaction pathway corresponds to the formation of 8OH-9MG+ by H elimination from the complex, referred to as HE. Among these product channels, [8OH-9MG + HN7]˙+ has the most favorable formation probability, especially in the presence of additional water molecules. This product may serve as a preceding structure to the 8-oxo-7,8-dihydroguanine lesion in DNA and has implications for health effects of radiation exposure and radiation therapy.

Computational Study of the Precursor Spectroelectrochemistry of Guanine

Precursor spectroelectrochemical behavior of guanine is investigated based on UV-Vis absorption and fluorescence, and IR and Raman spectra of guanine and its radical cation in the presence of a model electrode, computed using (TD)M06/6-31++G** method. Effects of electrode potential ( ), molecule-electrode distance (d) and molecular orientations (θ) on this behavior are investigated. Results indicate that application of electric potential causes changes in the molecular structure and distribution of charge and spin densities, which consequently changes the electronic and vibrational characteristics of the system. Also, perturbation due to the applied electric potential, changes both the intensities and frequencies of the vibrational bands of the studied species. The absorption wavelength, and the peak intensity and width of the electronic spectra of guanine and its radical cation also show sensitivity to the applied electrode potential. Presence of solvent both as electrostatic medium and as explicit solvent (molecules) have significant effects on the spectroelectrochemical properties of guanine, and change the chemical activity of guanine radical cation formed by the electrode reaction. Furthermore, population and orbital analyses show that for all orientations, application of the electric potential by the electrode increases contribution of the inter-molecular (guanine→water) charge density displacement to the UV-Vis transitions.

Deprotonation of Guanine Radical Cation G•+ Mediated by the Protonated Water Cluster.

Proton transfer is regarded as a fundamental process in chemical reactions of DNA molecules and continues to be an active research theme due to the connection with charge transport and oxidation damage of DNA. For the guanine radical cation (G•+) derived from one-electron oxidation, experiments suggest a facile proton transfer within the G•+:C base pair, and a rapid deprotonation from N1 in free base or single-strand DNA. To address the deprotonation mechanism, we perform a thorough investigation on deprotonation of G•+ in free G base by combining density functional theory (DFT) and laser flash photolysis spectroscopy. Experimentally, kinetics of deprotonation is monitored at temperatures varying from 280 to 298 K, from which the activation energy of 15.1 ± 1.5 kJ/mol is determined for the first time. Theoretically, four solvation models incorporating explicit waters and the polarized continuum model (PCM), i.e., 3H2O-PCM, 4H2O-PCM, 5H2O-PCM, and 7H2O-PCM models are used to calculate deprotonation potential energy profile, and the barriers of 5.5, 13.4, 14.4, and 13.7 kJ/mol are obtained, respectively. It is shown that at least four explicit waters are required for properly simulating the deprotonation reaction, where the participation of protonated water cluster plays key roles in facilitating the proton release from G•+.

Electron detachment dynamics of the iodide-guanine cluster: does ionization occur from the iodide or from guanine?

Laser photodissociation spectroscopy of the I‐·guanine complex has been conducted for the first time across the regions above the electron detachment threshold to explore the excited states and whether vertical ionisation occurs from the iodide or the nucleobase. The photofragment spectra reveal a prominent dipole-bound excited state (I) close to the calculated vertical electron detachment energy (∼4.0 eV) and a second excited (II) centred around 4.8 eV, which we assign to π-π* nucleobase-localised transitions. The ionic photofragments are identified as I‐ and I‐·[G-H], with the later fragment being produced significantly more strongly than the former. Both photofragments are observed across the two excited states, with production of the iodide being attributed to internal conversion to the ground state followed by evaporation. We trace the formation of the I‐·[G-H] photofragment to initial vertical ionisation of guanine, followed by ejection of a proton. This two-step process is important as it follows known steps in radiation-induced damage to DNA, namely initial formation of a guanine radical cation which then forms a free radical [G-H] moiety through deprotonation. Production of the I‐·[G-H] photofragment is pronounced through II indicating that its formation is enhanced by coupling of the π-π* transitions to the electron detachment continuum.

Neighboring base sequence effect on DNA damage

Guanine is the most strongly oxidized base in DNA; generation of a guanine radical cation as an intermediate in an oxidation reaction leads to migration through a resulting cationic hole in the DNA π-stack until it is trapped by irreversible reaction with water or other free radicals. In the case of normal sequences, the primary position of Guanine oxidations by one-electron oxidants such as carbonate radical anions, BPT(7,8,9,10-tetrahydroxytetrahydrobenzo[a]pyrene), and riboflavin are 5′-G in GG doublets and the central G in a GGG triplet. According to results, the properties of guanine oxidation on abasic site containing sequences are independent from the position of AP(apurinic/apyrimidinic) site in the presence of carbonate radical anions under a short irradiation time, although this radical is exposed to solvent by the existence of an abasic site. The lack of abasic site effect on guanine oxidative damage by the carbonate radical may be due to a sequence-independent property of the initial electron transfer rate in the hole injection step, or may relate to an electron transfer mechanism with large reorganization energy dependency. Consequently, the carbonate radical anions may easily migrate to another single G in the charge re-distribution step. Meanwhile, there is a strong dependency on the presence of an AP(apurinic/apyrimidinic) site in the cleavage patterns of guanine oxidations by physically large oxidizing agents, such as BPT(7,8,9,10-tetrahydroxytetrahydrobenzo[a]pyrene) and riboflavin. These radicals show strong AP(apurinic/apyrimidinic) site dependency and clear G-site selectivity.Communicated by Ramaswamy H. Sarma

Computational Study of the Formation of C8, C5, and C4 Guanine:Lysine Adducts via Oxidation of Guanine by Sulfate Radical Anion.

Oxidative damage to DNA can lead to DNA-protein cross-links which can interfere with DNA transcription, replication, and repair. In experimental studies modeling oxidative damage to DNA, oxidation of guanosine by sulfate radical anion in the presence of lysine produced a mixture of lysine (Lys)-substituted spiroiminodihydantoins (Sp): ∼65% 5-Lys-Sp, ∼30% 8-Lys-Sp, and ∼5% 5,8-diLys-Sp. Pathways for formation of the lysine adducts during the oxidation of guanine by sulfate radical anions have been mapped out using B3LYP density functional theory and the SMD solvation model. Methylamine was used as a model for lysine, and imidazole served as a proton acceptor. The lowest barrier for methylamine reaction with guanine radical is addition at C8, yielding mainly 8-NHR-Sp and some 5,8-diNR-Sp. This is in good agreement with the cross-link ratios for mild oxidations mediated by type I photosensitizers such as benzophenone, but this is not in agreement with the product ratios for strong oxidants such as sulfate radical anion. The calculations explored pathways for oxidation of guanine by sulfate radical anion that produced guanine radical and radical cation and doubly oxidized guanine (Gox) and its cation. Sulfate radical anion can also oxidize methylamine to produce neutral methylamine radical (CH3NH•) after deprotonation. The calculations qualitatively reproduced the observed product ratio at pH 7 via a pathway involving the barrierless addition of methylamine radical at C5 and C8 of guanine radical. After C5 addition of methylamine radical, the lowest barrier is for H2O addition at C8 leading exclusively to 5-NHR-Sp. After C8 addition of methylamine radical, H2O and methylamine addition to C5 lead to 8-NHR-Sp and some 5,8-diNR-Sp.

Monitoring the Structure-Dependent Reaction Pathways of Guanine Radical Cations in Triplex DNA: Deprotonation Versus Hydration.

Exposure of DNA to one-electron oxidants leads initially to the formation of guanine radical cations (G•+), which may degrade by deprotonation or hydration and ultimately cause strand breaks or 8-oxoG lesions. As the structure is dramatically changed by binding of the third strand in the major groove of the target duplex, it makes the triplex an interesting DNA structure to be examined and compared with the duplex on the G•+ degradation pathways. Here, we report for the first time the time-resolved spectroscopy study on the G•+ reaction dynamics in triplex DNA together with the Fourier transform infrared characterization of steady-state products, from which structural effects on the reactivity of G•+ are unraveled. For an antiparallel triplex-containing GGC motif, G•+ mainly suffers from fast deprotonation (9.8 ± 0.2) × 106 s-1, featuring release of both N1-H and N2-H of G in the third strand directly into bulk water. The much faster and distinct deprotonation behavior compared to the duplex should be related to long-resident water spines in the third strand. The G•+ hydration product 8-oxoG is negligible for an antiparallel triplex; instead, the 5-HOO-(G-H) hydroperoxide formed after G•+ deprotonation is identified by its vibrational marker band. In contrast, in a parallel triplex (C+GC), the deprotonation of G•+ occurs slowly (6.0 ± 0.3) × 105 s-1 with the release of N1-H, while G•+ hydration becomes the major pathway with yields of 8-oxoG larger than in the duplex. The increased positive charge brought by the third strand makes the G radical in the parallel triplex sustain more cation character and prone for hydration. These results indicate that non-B DNA (triplex) plays an important role in DNA damage formation and provide mechanistic insights to rationalize why triplex structures might become hot spots for mutagenesis.

A dinuclear ruthenium(ii) phototherapeutic that targets duplex and quadruplex DNA.

With the aim of developing a sensitizer for photodynamic therapy, a previously reported luminescent dinuclear complex that functions as a DNA probe in live cells was modified to produce a new iso-structural derivative containing RuII(TAP)2 fragments (TAP = 1,4,5,8-tetraazaphenanthrene). The structure of the new complex has been confirmed by a variety of techniques including single crystal X-ray analysis. Unlike its parent, the new complex displays Ru → L-based 3MLCT emission in both MeCN and water. Results from electrochemical studies and emission quenching experiments involving guanosine monophosphate are consistent with an excited state located on a TAP moiety. This hypothesis is further supported by detailed DFT calculations, which take into account solvent effects on excited state dynamics. Cell-free steady-state and time-resolved optical studies on the interaction of the new complex with duplex and quadruplex DNA show that the complex binds with high affinity to both structures and indicate that its photoexcited state is also quenched by DNA, a process that is accompanied by the generation of the guanine radical cation sites as photo-oxidization products. Like the parent complex, this new compound is taken up by live cells where it primarily localizes within the nucleus and displays low cytotoxicity in the absence of light. However, in complete contrast to [{RuII(phen)2}2(tpphz)]4+, the new complex is therapeutically activated by light to become highly phototoxic toward malignant human melanoma cell lines showing that it is a promising lead for the treatment of this recalcitrant cancer.

Computational Study of the pH-Dependent Competition between Carbonate and Thymine Addition to the Guanine Radical.

When oligonucleotides are oxidized by carbonate radical, thymine and carbonate can add to guanine radical, yielding either a guanine-thymine cross-link product (G∧T) or 8-oxo-7,8-dehydroguanine (8oxoG) and its further oxidation products such as spiroiminodihydantoin (Sp) and guanidinohydantoin (Gh). The ratio of thymine addition to carbonate addition depends strongly on the pH. Details of the mechanism have been explored by density functional calculations using the ωB97XD/6-31+G(d,p) level of theory with the SMD implicit solvation method, augmented with a few explicit waters. Free energies of intermediates and transition states in aqueous solution have been calculated along the pathways for addition of thymine, CO32-/HCO3- and carbonate radical to guanine radical. The pH dependence was examined by using appropriate explicit proton donors/acceptors as computational models for buffers at pH 2.5, 7, and 10. Deprotonation of thymine is required for nucleophilic addition at C8 of guanine radical, and thus is favored at higher pH. The barrier for carbonate radical addition is lower than for bicarbonate or carbonate dianion addition; however, for low concentrations of carbonate radical, the reaction may proceed by addition of bicarbonate/carbonate dianion to guanine radical. Thymine and bicarbonate/carbonate dianion addition are followed by oxidation by O2, loss of a proton from C8 and decarboxylation of the carbonate adduct. At pH 2.5, guanine radical cation can be formed by oxidization with sulfate radical. Water addition to guanine radical cation is the preferred path for forming 8oxoG at pH 2.5.

Biphotonic Ionization of DNA: From Model Studies to Cell.

Oxidation reactions triggered by low-intensity UV photons represent a minor contribution with respect to the overwhelming pyrimidine base dimerization in both isolated and cellular DNA. The situation is totally different when DNA is exposed to high-intensity UVC radiation under conditions where biphotonic ionization of the four main purine and pyrimidine bases becomes predominant at the expense of singlet excitation processes. The present review article provides a critical survey of the main chemical reactions of the base radical cations thus generated by one-electron oxidation of nucleic acids in model systems and cells. These include oxidation of the bases with the predominant formation of 8-oxo-7,8-dihydroguanine as the result of preferential hole transfer to guanine bases that act as sinks in isolated and cellular DNA. In addition to hydration, other nucleophilic addition reactions involving the guanine radical cation give rise to intra- and interstrand cross-links together with DNA-protein cross-links. Information is provided on the utilization of high-intensity UV laser pulses as molecular biology tools for studying DNA conformational features, nucleic acid-protein interactions and nucleic acid reactivity through DNA-protein cross-links and DNA footprinting experiments.