nitrogen-dioxide and 8-nitroguanine

8-Nitroguanine (8-NitroG) is known to be a potent mutagenic product obtained from the guanine base of DNA that induces GC to AT transversion mutation. Its formation by the reaction of guanyl radical with nitrogen dioxide radical (NO(2)(*)) has been observed experimentally. The reaction was studied here theoretically considering different reactant complexes, transition states, intermediate complexes, and product complexes. The solvent effect due to one, two, three, and six specific water molecules and the bulk solvent effect of aqueous media on the reaction were considered. With one, two, and three complexed water molecules, geometry optimization calculations were performed using the B3LYP and BHandHLYP functionals of density functional theory along with the 6-31G (d,p) and the AUG-cc-pVDZ basis sets in gas phase which was followed by single point energy calculations at the MP2/AUG-cc-pVDZ level. With six complexed water molecules, geometry optimization was performed at the BHandHLYP/6-31G(d,p) level of theory followed by single point energy calculations at all the other levels of theory mentioned above. Solvation in bulk aqueous media was treated at all the above-mentioned levels of theory using the polarizable continuum model (PCM). Specific water molecules were found to play catalytic roles in proton transfer processes that usually lowered the barrier appreciably.

Topics: Catalysis; Free Radicals; Guanine; Nitrogen Dioxide; Thermodynamics

The reaction between nitrogen dioxide (NO2*) and guanine radical cation (G*+) yielding the mutagenic product 8-nitroguanine radical cation (8-nitroG*+) was studied in the presence of one or two water molecules. All the relevant extrema on the potential energy surface were located by fully optimizing the geometries of the reactant, intermediate, and product complexes as well as transition states at the B3LYP/6-31G**, B3PW91/6-31G**, B3LYP/AUG-cc-pVDZ, and B3PW91/AUG-cc-pVDZ levels of density functional theory in gas phase. Zero point energy-corrected total energies and the corresponding Gibbs free energies at 298.15 K were obtained at the B3LYP/AUG-cc-pVDZ and B3PW91/AUG-cc-pVDZ levels of theory. Single point energy calculations were performed for all the optimized geometries at the MP2/AUG-cc-pVDZ level of theory in gas phase. Solvent effect of aqueous media was treated by performing single point energy calculations at the B3LYP/ AUG-cc-pVDZ, B3PW91/AUG-cc-pVDZ, and MP2/AUG-cc-pVDZ levels of theory employing the polarizable continuum model. The solvent effect of bulk water as well as that due to specific water molecules were found to play very important roles in lowering down many barrier energies appreciably. It is found that 8-nitroG*+ complexed with water molecules would be formed due to the reaction of G*+ with NO2* in aqueous media. The possible biological significance of the results obtained has been examined by studying binding energies of several normal and abnormal base pairs.

Topics: Algorithms; Cations; Computational Biology; Free Radicals; Guanine; Models, Theoretical; Molecular Conformation; Mutagenesis; Mutagens; Mutation; Nitrogen Dioxide; Software; Solvents; Temperature; Thermodynamics

We have developed an analytical method to quantitate urinary 8-nitroguanine, a product of nitrative nucleic acid damage caused by reactive nitrogen species such as peroxynitrite and nitrogen dioxide. 8-Nitroguanine was purified from human urine using immunoaffinity columns with an anti-8-nitroguanine antibody, followed by quantitation by high-performance liquid chromatography-electrochemical detection. Four sequential electrodes were used to (a) oxidize interfering compounds (+250 mV), (b) reduce nitrated bases (two online electrodes at -1000 mV), and (c) quantitate reduced derivatives (+150 mV). Using this system 8-nitroxanthine can also be detected, with the detection limits being 25 and 50 fmol/injection for 8-nitroguanine and 8-nitroxanthine, respectively. The method was used to analyze both adducts in the urine of smokers (n=12) and nonsmokers (n=17). We found that smokers excrete more 8-nitroguanine [median, 6.1 fmol/mg creatinine; interquartile range (IQR), 23.8] than nonsmokers (0; IQR, 0.90) (p=0.018), and although 8-nitroxanthine was detected in human urine, its level was not related to smoking status. This is the first report to show that 8-nitroguanine is present in human urine and the methodology developed can be used to study the pathogenic roles of this adduct in the etiology of cancers associated with cigarette smoking and inflammation.

Topics: Adult; Antibodies, Monoclonal; Ascites; Biological Assay; Biomarkers; Chromatography, Affinity; Chromatography, High Pressure Liquid; DNA Damage; Electrochemistry; Female; Guanine; Humans; Male; Nitrogen Dioxide; Nucleic Acids; Peroxynitrous Acid; Reactive Nitrogen Species; Smoking; Spectrometry, Mass, Electrospray Ionization; Xanthines

A novel photochemical approach is described for synthesizing site-specific 8-nitro-2'-deoxyguanosine (8-nitro-dG) adducts DNA. The method is based on the bimolecular reaction of a neutral, deprotonated guanine radical [G(-H)*] in DNA and nitrogen dioxide (*NO(2)) radicals. This approach is illustrated using the single-stranded oligodeoxyribonucleotide 5'-d(CCATCGCTACC) dissolved in an aqueous solution of nitrite and bicarbonate anions at pH 7.5. The photochemical synthesis was triggered by the selective photodissociation of persulfate anions to yield SO(4)(*-) radical anions by either 308 nm XeCl excimer laser pulses or by a continuous irradiation with 290-340 nm light from a 1000 W Xe lamp. The sulfate radicals formed generate the CO(3)(*-) and *NO(2) radicals by one-electron oxidation of the bicarbonate and nitrite anions. In turn, the CO(3)(*-) radicals site-selectively generate G(-H)* radicals in DNA that combine with *NO(2) to form 8-nitro-dG lesions in the oligonucleotide. The nitrated oligonucleotides were purified by reversed-phase HPLC techniques and are stable at 4 degrees C for at least 4 days, but depurinate at ambient temperatures of 23 degrees C at pH 7 with a half-life of approximately 20 h. The nature of the reaction and decomposition products were studied by a combination of ESI and MALDI-TOF mass spectrometric techniques.

Topics: DNA Adducts; DNA, Single-Stranded; Free Radicals; Guanine; Mutagenesis, Site-Directed; Nitrogen Dioxide; Photochemistry; Spectrometry, Mass, Electrospray Ionization; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization