DFT and MP2 study of low barrier proton transfer in hydrazide schiff base tautomers via water bridges and in the gas

MP2 and DFT studies were performed on the tautomers of N′-ethylideneacetohydrazide in different environments including gas phase, continuum solvent and microhydrated environment. The ground electronic state structures of the tautomers were optimized at the MP2 and B3LYP levels of theory using 6-311++G(d,p), separately. The optimized geometries of the transition states of different tautomerism processes, which occur through the proton transfer (PT) reaction, were determined using the QST3 approach at the same levels of theory. The same stability order as T1Z〉 T1E〉 T2ZE〉 T2ZZ〉 T2EE〉 T2EZ〉 T6〉 T4E was found for the tautomers in the gas phase, continuum solvent and microhydrated environment for both B3LYP and MP2 levels of theory. In addition, the variations of the Gibbs free energies of tautomeric processes, the activation Gibbs free energies of the forward and reverse tautomeric processes with the polarity of the solvent (in continuum solvent model) and the number of water molecules (in microhydrated environment) were investigated. It was found that the reverse tautomeric process is more favorable in all considered environments thermodynamically and kinetically. In addition, it was shown that the rate constants of the reverse and forward considered tautomeric processes decrease with the solvent polarity in the continuum solvent model and the process becomes more difficult than the gas phase. The opposite trend is seen in the microhydrated environment.     

Mutagenic mechanism of the A-T to G-C transition induced by 5-bromouracil: an ab Initio Study

The tautomerisms of uracil, 5-bromouracil (BrU), G-U, G-BrU, A-U, and A-BrU have been studied theoretically in an effort to investigate the mutagenicity of BrU. The ab initio calculations have been performed using HF and B3LYP methods with various basis sets. The relative stability of all tautomers was established. The intermolecular interactions between U, BrU, U*, BrU* (asterisks denote enol forms), and water have been studied. It shows that the possibility of tautomerism from BrU to BrU* is much more likely than that from U to U*. Further research indicates that BrU* tends to pair with guanine more easily than U*. The proton transfer process has been investigated by potential energy surface (PES) scan and transition state analysis. The results show that the proton transfer between G-U* and G*-U is monodirectional barrier-free proton transfer (BFPT), which terminates the base mismatch induced by U*. On the other hand, the proton transfer between G-BrU* and G*-BrU is bidirectional BFPT, which makes the base mismatch induced by BrU* sustained. On the basis of all of these calculated results, a new mutagenic mechanism for the A-T to G-C transition induced by 5-bromouracil is described in detail for the first time. It might give a new insight into the origin of the mutagenicity of the 5-Br derivative.     

Variations of the tautomeric preferences and π-electron delocalization for the neutral and redox forms of purine when proceeding from the gas phase (DFT) to water (PCM)

Quantum-chemical calculations were performed for all possible nine neutral tautomers of purine and their oxidized and reduced forms in water {PCM//DFT(B3LYP)/6?311+G(d,p)} and compared to those in the gas phase {DFT(B3LYP)/6?311+G(d,p)}. PCM hydration influences geometries, π-electron delocalization, and relative energies of purine tautomers in different ways. Generally, the harmonic oscillator model of electron delocalization (HOMED) indices increase when proceeding from the gas phase to aequeous solution for the neutral and redox forms of purine. Their changes for the neutral and oxidized tautomers are almost parallel to the relative energies showing that aromaticity plays an important role in the tautomeric preferences. Tautomeric stabilities and tautomeric preferences vary when proceeding from the gas phase to water indicating additionally that intra- and intermolecular interactions affect tautomeric equilibria. The tautomeric mixture of neutral purine in the gas phase consists mainly of the N9H tautomer, whereas two tautomers (N9H and N7H) dominate in water. For oxidized purine, N9H is favored in the gas phase, whereas N1H in water. A gain of one electron dramatically changes the relative stabilities of the CH and NH tautomers that C6H and C8H dominate in the tautomeric mixture in the gas phase, whereas N3H in water. These variations show exceptional sensitivity of the tautomeric purine system on environment in the electron-transfer reactions.     

Concerning the solvent effect in the tautomerism of uracil, 5-fluorouracil, and thymine by density-functional theory and ab initio calculations

The tautomerism of uracil, 5-fluorouracil, and thymine has been investigated in the gas phase and in solution. Electron correlation effects were included in ab initio computations at the MP2 level, and DFT calculations were performed using the B3LYP level. Full geometry optimizations were conducted at the HF/6-31G**, HF/6-31+G**, and B3LYP/6-31+G** levels. Single-point MP2/6-31+G** calculations were performed on the HF/6-31+G** optimized geometries. The influence of the solvent was examined from self-consistent reaction field calculations performed with )=2.21 (1,4-dioxane) and )=78.54 (water). The calculated relative free energies ((G) indicate that substitution of uracil at the position group does not change the relative free energy order of the uracil tautomers in the gas phase and in 1,4-dioxane (except at the MP2 level) whereas this ordering changes in water. Attachment of a fluorine atom changes the relative free energy order of uracil tautomers in the gas phase and in solution.     

Deamination features of 5-hydroxymethylcytosine,a radical and enzymatic DNA oxidation product

Geometric consequences of electron delocalization were studied for all possible adenine tautomers in aqueous solution by means of ab initio methods {PCM(water)//DFT(B3LYP)/6-311+G(d,p)} and compared to those in the gas phase {DFT(B3LYP)/6-311+G(d,p)}. To measure the consequences of any type of resonance conjugation (π-π, n-π, and σ-π), the geometry-based harmonic oscillator model of electron delocalization (HOMED) index, recently extended to the isolated (DFT) and hydrated (PCM//DFT) molecules, was applied to the molecular fragments (imidazole, pyrimidine, 4-aminopyrimidine, and purine) and also to the whole tautomeric system. For individual tautomers, the resonance conjugations and consequently the bond lengths strongly depend on the position of the labile protons. The HOMED indices are larger for tautomers (or their fragments) possessing the labile proton(s) at the N rather than C atom. Solvent interactions with adenine tautomers slightly increase the resonance conjugations. Consequently, they slightly shorten the single bonds and lengthen the double bonds. When going from the gas phase to water solution, the HOMED indices increase (by less than 0.15 units). There is a good relation between the HOMED indices estimated in water solution and those in the gas phase for the neutral and ionized forms of adenine. Subtle effects, being a consequence of intramolecular interactions between the neighboring groups, are so strongly reduced by solvent that the relation between the HOMED indices and the relative energies for the neutral adenine tautomers seems to be better in water solution than in the gas phase.

The eosinophil peroxidase-hydrogen peroxide-bromide system of human eosinophils generates 5-bromouracil, a mutagenic thymine analogue

Eosinophils use eosinophil peroxidase, hydrogen peroxide (H(2)O(2)), and bromide ion (Br(-)) to generate hypobromous acid (HOBr), a brominating intermediate. This potent oxidant may play a role in host defenses against invading parasites and eosinophil-mediated tissue damage. In this study, we explore the possibility that HOBr generated by eosinophil peroxidase might oxidize nucleic acids. When we exposed uracil, uridine, or deoxyuridine to reagent HOBr, each reaction mixture yielded a single major oxidation product that comigrated on reversed-phase HPLC with the corresponding authentic brominated pyrimidine. The eosinophil peroxidase-H(2)O(2)-Br(-) system also converted uracil into a single major oxidation product, and the yield was near-quantitative. Mass spectrometry, HPLC, UV--visible spectroscopy, and NMR spectroscopy identified the product as 5-bromouracil. Eosinophil peroxidase required H(2)O(2) and Br(-) to produce 5-bromouracil, implicating HOBr as an intermediate in the reaction. Primary and secondary bromamines also brominated uracil, suggesting that long-lived bromamines also might be physiologically relevant brominating intermediates. Human eosinophils used the eosinophil peroxidase-H(2)O(2)-Br(-) system to oxidize uracil. The product was identified as 5-bromouracil by mass spectrometry, HPLC, and UV--visible spectroscopy. Collectively, these results indicate that HOBr generated by eosinophil peroxidase oxidizes uracil to 5-bromouracil. Thymidine phosphorylase, a pyrimidine salvage enzyme, transforms 5-bromouracil to 5-bromodeoxyridine, a mutagenic analogue of thymidine. These findings raise the possibility that halogenated nucleobases generated by eosinophil peroxidase exert cytotoxic and mutagenic effects at eosinophil-rich sites of inflammation.     

Effects of p-fluorophenylalanine on the induction of mutations in bacteriophage T4: I. 5-Bromouracil mutagenesis

The amino acid analogue p-fluorophenylalanine (PFPA) was found to have no mutagenic activity in the r system of bacteriophage T4. However, under standard conditions for 5-bromouracil (5-BU) mutagenesis, PFPA depressed the induced frequencies for both forward and reverse mutations. When the folate antagonist sulphanilamide (SU) was omitted from the mutagenic treatment medium or when it was replaced by Trimethoprim (TM), another folate antagonist, this depressive effect was abolished. It was proposed that PFPA alleviated the inhibitory action of SU.     

Uracil-DNA glycosylase causes 5-bromodeoxyuridine photosensitization in Escherichia coli K-12.

An Escherichia coli uracil-DNA glycosylase-defective mutant (ung-1 thyA) was more resistant than its wild-type counterpart (ung+ thyA) to the killing effect of UV light when cultured in medium containing 5-bromouracil or 5-bromo-2'-deoxyuridine (BrdUrd). The phenotype of resistance to BrdUrd photosensitization and the uracil-DNA glycosylase deficiency appeared to be 100% cotransduced by P1 phage. During growth with BrdUrd, both strains exhibited similar growth rates and 5-bromouracil incorporation into DNA. The resistant phenotype of the ung-1 mutant was observed primarily during the stationary phase. In cells carrying 5-bromouracil-substituted DNA, mutations causing resistance to rifampin and valine were induced by UV irradiation at a higher frequency in the wild type than in the ung-1 mutant. This Ung-dependent UV mutagenesis required UmuC function. These results suggest that the action of the uracil-DNA glycosylase on UV-irradiated 5-bromouracil-substituted DNA produces lethal and mutagenic lesions. The BrdUrd photosensitization-resistant phenotype allowed us to develop a new, efficient method for enriching and screening ung mutants.     

Studies on tautomeric forms of Guanine-Cytosine base pairs of nucleic acids and their interactions with water molecules

The relative stabilities of Guanine-Cytosine (G-C) DNA bare base pairs, its tautomeric forms and microhydrated base pairs are theoretically investigated with a focus on the keto-enol tautomerism as well as on the cis-trans isomerism using ab initio and density functional theory methods. The stabilities of the G-C bare base pairs, its tautomeric forms and microhydrated base pairs were affected by various factors including keto-enol tautomerization, cis-trans enol isomerization, and steric hindrance between the base pair and water molecules. The Atoms in Molecules theory (AIM) is employed to investigate H-bonding patterns both in bare and microhydrated base pairs. From the above topological results, an excellent linear correlation is shown between electron density [rho(r)], and its Laplacian [V2rho(r)] at the bond critical points. NBO analysis has been carried out to study the charge transfer between proton acceptor to the antibonding orbital of the X-H bond both in bare and microhydrated base pairs.     

Replication of DNA containing 5-bromouracil can be mutagenic in Syrian hamster cells.

A new protocol for inducing mutations in mammalian cells in culture by exposure to the thymidine analog 5-bromodeoxyuridine (BrdUrd) was established. This protocol, called "DNA-dependent" mutagenesis, involved the incorporation of BrdUrd into DNA under nonmutagenic conditions and the subsequent replication of the 5-bromouracil (BrUra)-containing DNA under mutagenic conditions but with no BrdUrd present in the culture medium. The mutagenic conditions were induced by allowing BrUra-containing DNA to replicate in the presence of high concentrations of thymidine. This generated high intracellular levels of dTTP and dGTP, causing nucleotide pool imbalance. The mutagenesis induced by this protocol was found to correlate with the level of BrUra substituted for thymine in DNA.     

DFT study of the gas phase proton transfer in guanine assisted by water, methanol, and hydrogen peroxide

A computational study of hydrogen-bonded complexes between the oxo-/hydroxo-amino N7/9H tautomers of guanine and water, methanol, and hydrogen peroxide has been performed at the B3LYP/6-31+G(d) level of theory. The mechanisms of the water-, methanol-, and hydrogen peroxide-assisted proton transfers in guanine were studied and compared with the intramolecular proton transfer in guanine in the gas phase. It was found that the assisted proton transfers pass through about three times lower energy barriers than those found for isolated guanine tautomers. Figure DFT study of the gas phase proton transfer in guanine assisted by water, methanol and hydrogen peroxide     

Quinolone compounds with carboxylic equivalents and semiempirical calculations on their tautomers

Quinolone antibiotic compounds having no carboxylic group but its equivalents at the 3-position have been synthesized. Energies of tautomers with the carboxylic mimics were calculated in gas phase and aqueous solution with AM1-SCRF method. Antibacterial activity of the compounds was rationalized with tautomeric protons and atomic charges of calculated tautomers.     

5-Methyltryptophan resistance mutations in Escherichia coli K-12. Mutagenic activity of monofunctional alkylating agents including organophosphorus insecticides

The induction of 5-methyltryptophan (5-MT) resistance mutations was assayed as a test system for mutagenic chemicals in Escherichia coli. It is assumed that different premutational alterations in several genes of the Escherichia coli chromosome will lead to 5-MT-resistant mutants. The chemicals used were three monofunctional alkylating agents as reference compounds, namely β-propiolactone (β-PL), N-methyl-N′-nitro-N-nitrosoguanidine (MNNG), and methyl methanesulfonate (MMS), which are all mutagenic in the 5-MT system; of the eight organophosphorus insecticides tested, four have definite mutagenic activity (Dichlorvos, Oxydemetonmethyl, Dimethoate, and Bidrin), one is probably mutagenic (Methylparathion) and the remaining three (Parathion, Malathion and Diazinon) do not induce 5-MT resistance mutations in the conditions used here (< 30% survival). The relative mutagenic activity after a treatment time of 60 min is (in decreasing order) MNNG > MMS > Dichlorvos > Oxydemetonmethyl, Dimethoate and Bidrin. The concentration-dependent mutagenic activity of all mutagenic compounds is nearly linear when plotted on a log-log scale (with slopes varying from 1.0 to 1.5) and could be taken as an indication that one premutational reaction will be sufficient for the induction of one 5-MT-resistant mutant.     

Quantum chemical investigation of intramolecular thione-thiol tautomerism of 1,2,4-triazole-3-thione and its disubstituted derivatives

The one step intramolecular thione-thiol tautomerism of 1,2,4-triazole-3-thione and its disubstituted derivatives has been studied through the use of electronic structure methods. Due to the absence of experimental data for the parent molecule of 1,2,4-triazole-3-thione the structure and energetics of aforementioned tautomers were derived using various basis sets and levels including HF, B3LYP, and MP2 methods. The gas phase results show that in all different levels of theory the most stable tautomer is the thione form. It has also been revealed that B3LYP/6-31G(d,p) level is quite well suited and reliable to investigate these kinds of tautomerism. To account the influence of substituents on the mentioned tautomerization, the tautomerism and conformational properties as well as vibrational analysis of 20 halophenyl and isopyridyl derivatives were investigated using B3LYP/6-31G(d,p) calculations. In all cases the calculations indicate that substituents have no considerable effects on relative stabilities and energy barriers for the thione-thiol proton transfer and the thione forms are the predominant species in the gas phase. In order to figure out the relative stabilities of the species involved in the tautomerism, geometrical and natural bond orbital (NBO) analyses have been employed. It has also been shown that the computed vibrational frequencies of tautomers with different scaling factors could be used to interpret the vibrational frequencies in IR spectrum of similar species.     

Conformation and tautomerizm of the 2-methyl-4-pyridin-2′-yl-1,5-benzodiazepine molecule. An ab initio study

The protomeric tautomerizm and conformation of the 2-methyl-4-pyridin-2′-yl-1,5-benzodiazepine molecule were investigated, and its three neutral tautomers (B₁,B₂,B₃) and their rotamers (C₁,C₂,C₃) were considered. Full geometry optimizations were carried out at the HF/6-31G* and B3LYP/6-31G* levels in gas phase and in water. The tautomerization processes in water (ɛ = 78.54) were studied by using self-consistent reaction field theory. The calculation showed that the boat conformation is dominant for the seven-membered diazepine ring in all of the structures, even with different double bond positions. The calculated relative free energies (ΔG) showed that the tautomer C₁ was the most stable structure, and its conformer B₁ was the second most stable in the gas phase and in water. Figure 2-Methyl-4-pyridin-2′-yl-1,5-benzodiazepine     

Synthesis and antitumor activity of 5-bromo-1-mesyluracil

Large-scale preparation of 5-bromo-1-mesyluracil (BMsU) 4 has been optimized. BMsU was synthesized by condensation of silylated 5-bromouracil and MsCl in acetonitrile or by the reaction of 5-bromouracil with MsCl in pyridine. The same product was obtained by bromination of 1-mesyluracil. The purpose of this study was to elucidate the effects of BMsU on the biosynthetic activity of tumor cell enzymes involved in DNA, RNA and protein syntheses, and in de novo and salvage pyrimidine and purine syntheses. Investigations were performed in vitro on human cervix carcinoma cells (HeLa). BMsU displayed inhibitory effects on DNA and RNA syntheses in HeLa cells after 24 h of treatment. De nova biosynthesis of pyrimidine and purine was also affected. Antitumor activity of BMsU is closely associated with its inhibitory activity on the enzymes that play an important role in the metabolism of tumor cells. In vivo antitumor activity of BMsU was also investigated. The model used in investigations was a mouse anaplastic mammary carcinoma transplanted into the thigh of the right leg of CBA mice. Significant reduction in tumor growth time was achieved with BmsU administered at a dose of 50 mg/kg.     

Mutagenicity,Creatine and Nutrient Contents of Pan Fried Meat from Various Animal Species

The mutagenic activity in extracts of fried meat from 16 different animal species was studied in Salmonella typhimurium TA98. In each experiment, 1 meat sample together with a standard beef sample was fried, and the mutagenicity was expressed relative to the beef sample. All meat samples showed less mutagenic activity than beef. The contents of creatine, creatinine, water, protein, carbohydrate and fat in the meat samples were analyzed, but mutagenicity was not correlated with the concentration of any of these constituents. Beef meat treated with creatinase to remove creatine produced reduced mutagenic activity. Possibly a threshold concentration of creatine is necessary to give a high mutagenic response.     

SYNTHESIS AND ANTITUMOR ACTIVITY OF 5-BROMO-1-MESYLURACIL

Large-scale preparation of 5-bromo-1-mesyluracil (BMsU) 4 has been optimized. BMsU was synthesized by condensation of silylated 5-bromouracil and MsCl in acetonitrile or by the reaction of 5-bromouracil with MsCl in pyridine. The same product was obtained by bromination of 1-mesyluracil. The purpose of this study was to elucidate the effects of BMsU on the biosynthetic activity of tumor cell enzymes involved in DNA, RNA and protein syntheses, and in de novo and salvage pyrimidine and purine syntheses. Investigations were performed in vitro on human cervix carcinoma cells (HeLa). BMsU displayed inhibitory effects on DNA and RNA syntheses in HeLa cells after 24 h of treatment. De nova biosynthesis of pyrimidine and purine was also affected. Antitumor activity of BMsU is closely associated with its inhibitory activity on the enzymes that play an important role in the metabolism of tumor cells. In vivo antitumor activity of BMsU was also investigated. The model used in investigations was a mouse anaplastic mammary carcinoma transplanted into the thigh of the right leg of CBA mice. Significant reduction in tumor growth time was achieved with BmsU administered at a dose of 50 mg/kg.     

The electron affinities of deprotonated adenine, guanine, cytosine, uracil, and thymine

Electron attachment rates and gas phase acidities for the canonical tautomers of the nucleobases and electron affinities for thymine, deprotonated thymine, and cytosine are reported The latter are from a new analysis of published photoelectron spectra. The values for deprotonated thymine are (all in eV) keto-N1-H, 3.327(5); enol-N3-H, 3.250(5), enol-C2OH, 3.120(5) enol-N1-H, 3.013(5), and enol-C4OH,3.123(5). The values for deprotonated cytosine, keto-N1-H, 3.184(5); trans-NH-H, 3.008(5); cis-NH-H, 3.039(5); and enol-N1-H, 2.750(5) and enol-O-H, 2.950(5). The gas phase acidities from these values are obtained from these values using experimental or theoretical calculations of bond dissociation energies. Kinetic and thermodynamic properties for thermal electron attachment to thymine are obtained from mass spectrometric data. We report an activation energy of 0.60 eV and electron affinity of thymine, 1.0(1) eV.     

The Electron Affinities of Deprotonated Adenine,Guanine, Cytosine,Uracil, and Thymine

Electron attachment rates and gas phase acidities for the canonical tautomers of the nucleobases and electron affinities for thymine, deprotonated thymine, and cytosine are reported The latter are from a new analysis of published photoelectron spectra. The values for deprotonated thymine are (all in eV) keto-N1-H, 3.327(5); enol-N3-H, 3.250(5), enol-C2OH, 3.120(5) enol-N1-H, 3.013(5), and enol-C4OH,3.123(5). The values for deprotonated cytosine, keto-N1-H, 3.184(5); trans-NH-H, 3.008(5); cis-NH-H, 3.039(5); and enol-N1-H, 2.750(5) and enol-O-H, 2.950(5). The gas phase acidities from these values are obtained from these values using experimental or theoretical calculations of bond dissociation energies. Kinetic and thermodynamic properties for thermal electron attachment to thymine are obtained from mass spectrometric data. We report an activation energy of 0.60 eV and electron affinity of thymine, 1.0(1) eV.