Meteorite-catalyzed syntheses of nucleosides and of other prebiotic compounds from formamide under proton irradiation.

Liquid formamide has been irradiated by high-energy proton beams in the presence of powdered meteorites, and the products of the catalyzed resulting syntheses were analyzed by mass spectrometry. Relative to the controls (no radiation, or no formamide, or no catalyst), an extremely rich, variegate, and prebiotically relevant panel of compounds was observed. The meteorites tested were representative of the four major classes: iron, stony iron, chondrites, and achondrites. The products obtained were amino acids, carboxylic acids, nucleobases, sugars, and, most notably, four nucleosides: cytidine, uridine, adenosine, and thymidine. In accordance with theoretical studies, the detection of HCN oligomers suggests the occurrence of mechanisms based on the generation of radical cyanide species (CN·) for the synthesis of nucleobases. Given that many of the compounds obtained are key components of extant organisms, these observations contribute to outline plausible exogenous high-energy-based prebiotic scenarios and their possible boundary conditions, as discussed.

Optical Detection of Photorelease Kinetics on Gold and Glass Surfaces using Streptavidin-Coupled Biotinylated Photolabile Protecting Groups for Nucleosides.

Five biotinylated photolabile compounds of the general structure Bt-L1 -NPPOC-X-L2 were synthesized, in which Bt represents a biotin unit, L1 is a 3,6-dioxa-n-octane or an n-hexane spacer, NPPOC is the photolabile protecting group 2-(2-nitrophenyl)propoxycarbonyl, and X is a thymidine unit as a representative nucleoside or a direct linkage to L2 , an ω-mercapto- or ω-aminohexoyl linker, for coupling to a substrate surface. These compounds served for testing the photocleavage kinetics in self-assembled monolayers on gold or glass by using surface plasmon resonance (SPR) on gold or reflectometric interference spectroscopy (RIfS) on glass, whereby the biotin moiety offered the possibility to increase the bulkiness of the leaving group by binding to streptavidin, which thereby largely enhanced the SPR or RIfS signals. The photokinetics, found to consist in a dominating fast stage and a less contributing slow stage, were quantitatively analyzed, and the quantum yield of the fast part reached values up to almost 1 in favorable cases. A direct comparison of the results from SPR and RIfS yielded almost identical results. The present investigations pave the way to in situ monitoring of the photolithographic synthesis of DNA chips.

Ultrafast non-radiative decay of gas-phase nucleosides.

The ultrafast photo-physical properties of DNA are crucial in providing a stable basis for life. Although the DNA bases efficiently absorb ultraviolet (UV) radiation, this energy can be dissipated to the surrounding environment by the rapid conversion of electronic energy to vibrational energy within about a picosecond. The intrinsic nature of this internal conversion process has previously been demonstrated through gas phase experiments on the bases, supported by theoretical calculations. De-excitation rates appear to be accelerated when individual bases are hydrogen bonded to solvent molecules or their complementary Watson-Crick pair. In this paper, the first gas-phase measurements of electronic relaxation in DNA nucleosides following UV excitation are reported. Using a pump-probe ionization scheme, the lifetimes for internal conversion to the ground state following excitation at 267 nm are found to be reduced by around a factor of two for adenosine, cytidine and thymidine compared with the isolated bases. These results are discussed in terms of a recent proposition that a charge transfer state provides an additional internal conversion pathway mediated by proton transfer through a sugar to base hydrogen bond.

5-Ethyldeoxyuridine, a thymidine analog: photochemical transformation.

Irradiation at 254 millimicrons transforms 5-ethyldeoxyuridine to deoxyuridine by way of photohydration of the 5,6 bond and elimination of ethanol. At wavelengths to the red side of 265 millimicrons, photodimerization is the principal reaction, with a pronounced oxygen effect. The results are related to the photochemistry of thymidine and of bacteriophages containing incorporated 5-ethyluracil in place of thymine.

Ultrafast reversible photo-cross-linking reaction: toward in situ DNA manipulation.

We describe a novel ultrafast reversible DNA interstrand photo-cross-linking reaction via 3-cyanovinylcarbazole nucleoside ( (CNV)K). Oligodeoxynucleotide (ODN) containing (CNV)K can be photo-cross-linked by irradiation at 366 nm for 1 s, and the photo-cross-linked ODN can be split by irradiation at 312 nm for 60 s.

Base release in nucleosides induced by low-energy electrons: a DFT study.

Low-energy electrons are known to induce strand breaks and base damage in DNA and RNA through fragmentation of molecular bonding. Recently the glycosidic bond cleavage of nucleosides by low-energy electrons has been reported. These experimental results call for a theoretical investigation of the strength of the C(1)'-N link in nucleosides (dA, dC and dT) between the base and deoxyribose before and after electron attachment. Through density functional theory (DFT) calculations, we compare the C(1)'-N bond strength, i.e., the bond dissociation energy of the neutral and its anionic radical, and find that an excess electron effectively weakens the C(1)'- N bond strength in nucleosides by 61-75 kcal/mol in the gas phase and 76-83 kcal/mol in the solvated environment. As a result, electron-induced fragmentation of the C(1)'-N bond in the gas phase is exergonic for dA (DeltaG=-14 kcal/mol) and for dT (DeltaG=-6 kcal/mol) and is endergonic (DeltaG=+1 kcal/ mol) only for dC. In the gas phase all the anionic nucleosides are found to be in valence states. Solvation is found to increase the exergonic nature by an additional 20 kcal, making the fragmentation both exothermic and exergonic for all nucleoside anion radicals. Thus C(1)'-N bond breaking in nucleoside anion radicals is found to be thermodynamically favorable both in the gas phase and under solvation. The activation barrier for the C(1)'-N bond breaking process was found to be about 20 kcal/mol in every case examined, suggesting that a 1 eV electron would induce spontaneous cleavage of the bond and that stabilized anion radicals on the DNA strand would undergo base release at only a modest rate at room temperature. These results suggest that base release from nucleosides and DNA is an expected consequence of low-energy electron-induced damage but that the high barrier would inhibit this process in the stable anion radicals.

Increasing urinary levels of modified nucleosides and bases during tumor development in mice.

Based on the fact that human cancer patients excrete increased amounts of various modified nucleosides and bases in their urine, we investigated whether the same phenomenon takes place in mice bearing experimentally induced tumors. We did indeed find that mice with MCA-induced skin tumors and mice exhibiting leukemia after X-ray irradiation excrete severalfold higher levels of modified nucleosides and bases than do untreated control mice. Comparison of the time course of altered urinary excretion of these RNA catabolites with the appearance of a recognizable tumor after MCA application revealed that the onset of the altered excretion rate of these compounds precedes tumor diagnosis. At present, the time-course studies in our mice exposed to a single X-ray dose to induce lymphoblastic leukemia seem to indicate a similar situation. Mice exhibiting preleukemic histological features already excrete increased amounts of various modified nucleosides and bases. Confirmation of our results by analysis of further irradiation-exposed mice in our present detailed time-course studies and of tumors experimentally induced in other organs of mice and other species will be taken as a basis for developing an in vivo test for carcinogenicity. Furthermore, the results could provide a foundation for the setting up of a noninvasive, early screening method for cancer in human beings.

Photoionization of DNA and RNA bases, nucleosides and nucleotides through a combination of one- and two-photon pathways upon 266 nm nanosecond laser excitation.

The 266 nm nanosecond laser photolysis of various purine and pyrimidine derivatives results in their photoionization (PI) as one of the primary photochemical pathways. Electron photoejection occurs through a combination of one- and two-photon mechanisms. The PI values depend on the substituents attached to the chromophore of the base. The net PI of the purine bases at 266 nm are of the same order of magnitude (10(-2)) as those of the pyrimidine bases under similar experimental conditions. The monophotonic component is approximately one-third of the net PI yield of the bases. A nonrelaxed singlet excited state intermediate is tentatively proposed for this pathway. It is proposed that this state is significantly stabilized by water solvation, transforming it into a charge transfer to solvent state from which the hydrated electron evolves.

Alkali-labile photolesions mapping to purine sites in ultraviolet-irradiated DNA.

Gel sequencing experiments with the 5'- and 3'-end-labeled oligonucleotides d(A3GA4GA5GA6GA3G) and d(AT)10 have demonstrated that dimeric adenine photoproducts and thymine-adenine photoadducts constitute alkali-labile lesions in UV-irradiated DNA. On treatment with hot piperidine, DNA strand breakage occurs predominantly at the sites of 5'-adenines in the dimeric photoproducts and of 3'-adenines in the thymine-adenine photoadducts. With 5'-end-labeled oligonucleotides of mixed sequence, major UV-induced loci for alkaline cleavage map to purine bases flanked on their 5'-side by two pyrimidines. This behavior does not arise from enhanced photoreactivity of purines in this sequence context as has been inferred from photofootprinting studies. Instead, as shown by 3'-labeling and selective substitution with 5-methylcytosine, it results from the anomalous electrophoretic mobility of 5'-end-labeled fragments produced by alkaline cleavage of DNA at adjacent pyrimidine (6-4) pyrimidone photoproducts.

Enhancement of radiation-induced base release from nucleosides in alkaline solution: essential role of the O.- radical.

The effect of pH on base release in the gamma-radiolysis of N2O-saturated solutions of a number of nucleosides (including uridine, 3-methyluridine, 2',3'-O-isopropylidene-uridine, and adenosine) has been investigated. For all these nucleotides, independent of the base or sugar moiety, base release is very low at pH below 10 (G approximately (0.3-0.7) x 10(-7) mol J-1), but increases drastically to G approximately (3-4) x 10(-7) mol J-1 at pH greater than or equal to 13. This phenomenon had already been previously reported and attributed to an OH(-)-induced transfer of a base radical into a sugar radical. However, it is now shown that at pH 12, where base release starts to increase, a lowering of the dose-rate does not affect the yield of free base. The increase in base release is accompanied by an overall reduction of chromophore loss of similar magnitude (with 2',3'-O-isopropylidene-uridine and 3-methyluridine), as well as by an increase in the yield of oxidizing radicals by a factor of 2 (with uridine). The measured rate constant of the reaction of .OH/O.- with the nucleosides is also pH-dependent, as .OH reacts faster than O.- with the nucleosides by a factor of 6-7. It is concluded that the increase in base release at high pH is caused by the increasing participation of O.-, which, unlike .OH, attacks the nucleosides preferentially at their sugar moieties, and is not due to an OH(-)-induced radical transfer from the base to the sugar moiety.

Mutagenicity of irradiated solutions of nucleic acid bases and nucleosides in Salmonella typhimurium.

Solutions of nucleic acid bases, nucleosides and a nucleotide, saturated with either N2, N2O or O2, were irradiated and tested for mutagenicity towards Salmonella typhimurium, with and without pre-incubation. Irradiated solutions of the nucleic acid bases were all non-mutagenic. Irradiated solutions of the nucleosides showed mutagenicity in S. typhimurium TA100 (pre-incubation assay). Generally, the mutagenicity followed the order: N2O greater than N2 greater than O2. The results show that the formation of mutagenic radiolytic products is initiated by attack of mainly OH radicals on the 2-deoxy-D-ribose moiety of the nucleosides. With irradiated solutions of the nucleotide, thymidine-5'-monophosphate, no mutagenicity could be detected.

Solid-state radiation chemistry of DNA and its constituents.

The molecular structure as well as the mechanisms of formation and decay of free radicals produced in DNA and its constituents by ionizing radiation is reviewed. Starting with the description of the spectral parameters for cations and anions in natural nucleic acid bases, emphasis is given to the comparable species formed in the group of the 5-halogen substituted uracil derivatives. The consequences of the attachment of a ribose or ribosephosphate group to the bases is discussed in terms of the distribution of primary radicals which, again, is shown to be different from those of the subunits in DNA itself. The quantitative aspects of radical formation are discussed in terms of G values and their dependence on the temperature of irradiation. Finally, a schematic presentation of the major modes of radical reactions is given occurring upon warming of the primary species in the DNA subunits and in DNA itself.

Free radicals induced in solid DNA by heavy ion bombardment.

Free radical formation after heavy-ion bombardment was studied in solid, polycristalline pellets of DNA-constituents by analysing the dose-yield curves and the spectra obtained by ESR-spectroscopy at low (< 100 K) and ambient temperatures. The dose-yield curves were found to correlate with those found after X-irradiation but shifted to higher doses and lower saturation concentrations. The corresponding radical yields (per 100 eV) exhibit values which are one to two orders of magnitudes lower. The structural aspects as revealed from powder ESR-spectra gave a complex inter-relation between substance, LET, dose and irradiation temperature, which is discussed in terms of mechanistic models.

Abiogenic synthesis of pyrimidine nucleotides in solid state by vacuum ultraviolet radiation.

The abiogenic synthesis of pyrimidine nucleotides in solid state has been investigated. Our experiment indicates that natural nucleotides are produced in thin films prepared from nucleoside and inorganic phosphate by irradiating with vacuum ultraviolet light (VUV, lambda=100-200 nm). We have investigated the influence of the type of nucleic acids base (thymidine, cytosine, uracil) and the structure of sugar moiety (ribose or deoxyribose) on the course and yield of reaction. We compared the action of vacuum ultraviolet light with action of gamma-radiation, heat and biology significant UV (254 nm) which have been investigated earlier. The occurrence of these reaction in open space is discussed.

Far UV photolysis of uracil and cytosine in phosphate solution.

The photolysis of nucleobases, nucleosides and nucleotides (NA) was enhanced by phosphate under irradiation of medium pressure mercury lamp (MPML). Uracil and Cytosine in phosphate solution were selected to study the mechanism of phosphate effect. Photoproducts were produced in the irradiated uracil and cytosine of phosphate solution which have been isolated by anion exchange resin. Ultraviolet irradiation (190-220nm) of uracil in 0.05 mol/dm3 phosphate buffered solution at pH 8-9 leads to the production of a novel compound C4H5N2O6P which has been identified by UV, 1H-NMR spectroscopy and LC/MS/MS. The formation mechanism of the photoproduct and the kinetics were studied.

[The role of ultraviolet and gamma radiation in the abiogenic synthesis of nucleotides in the solid state]. / Rol' UF- i gamma-radiatsii v abiogennom sinteze nukleotidov v tverdom sostoianii.

A possibility of abiogenic synthesis of natural nucleotides in solid state under the influence of UV and gamma radiation has been shown for a mixture of nucleosides and dihydrophosphate. Adenosine and deoxyadenosine were used as initial substances. During adenosine and deoxyadenosine phosphorylation under the action of UV radiation (lambda = 254 nm), only 5'-monophosphates were formed, a yield of 0.094 and 0.36% respectively. Under the action of gamma radiation, 2',3'-cAMP (0.12%), 2'-AMP (0.08%) and 3'-AMP (0.14%) were produced together with 5'AMP (0.41%).

Microwave-assisted preparation of nucleoside-phosphoramidites.

Microwave-assisted phosphitylation of sterically hindered nucleosides is demonstrated to be an efficient method for the preparation of corresponding phosphoramidites (otherwise onerous under standard conditions) and is shown to be general in its applicability.