Thymine Research Articles

A Theoretical Exploration of the Photoinduced Breaking Mechanism of the Glycosidic Bond in Thymine Nucleotide.

DNA glycosidic bond cleavage may induce cancer under the ultraviolet (UV) effect. Yet, the mechanism of glycosidic bond cleavage remains unclear and requires more detailed clarification. Herein, quantum chemical studies on its photoinduced mechanism are performed using a 5'-thymidine monophosphate (5'-dTMPH) model. In this study, four possible paths were examined to study the glycosidic bond cleavage. The results showed that, upon excitation, the electronic transition from the π bonding to π antibonding orbitals of the thymine ring leads to the damage of the thymine ring. Afterwards, the glycosidic bond is cleaved. At first, the doublet ground state (GS) path of glycosidic bond cleavage widely studied by other groups is caused by free electron generated by photoirradiation, with a kinetically feasible energy barrier of ~23 kcal/mol. Additionally, then, the other three paths were proposed that also might cause the glycosidic bond cleavage. The first one is the doublet excited state (ES) path, triggered by free electron along with UV excitation, which can result in a very-high-energy barrier ~49 kcal/mol that is kinetically unfavorable. The second one is the singlet ES path, induced by direct UV excitation, which assumes DNA is directly excited by UV light, which features a very low-energy barrier ~16 kcal/mol that is favored in kinetics. The third one is the triplet ES path, from the singlet state via intersystem crossing (ISC), which refers to a feasible ~27 kcal/mol energy barrier. This study emphasizes the pivotal role of the DNA glycosidic bond cleavage by our proposed direct UV excitation (especially singlet ES path) in addition to the authorized indirect free-electron-induced path, which should provide essential insights to future mechanistic comprehension and novel anti-cancer drug design.

Identifying mutation region of herbicide-resistant Mutant Quinoa (Chenopodium quinoa Willd.) lines by molecular characterization

Quinoa can be considered as an alternative plant with its features being compatible with unfavorable climate and soil conditions. Moreover, the nutrient content of its seed is great. But there is not a selective herbicide for weeds in quinoa fields. This is a significant problem in the quinoa cultivation. Therefore, it is necessary to develop herbicide-resistant quinoa lines In the study, Tititcaca, a variety of quinoa was used. Sodium azide was used 1.5 mM for mutagenesis. Herbicide-resistant plants were determined by applying herbicide belonging to the IMI group in the M3 generation. These were named ET-6, ET-7, OT-11 and T-103. The 4 mutant lines obtained as a result of seed mutagenesis, and mutant line OT-11 plants occurred in the form of replacement of the cytosine nucleotide with the adenine nucleotide (CA) in the ALS gene. Line ET-6 plants, the thymine nucleotide was replaced with a guanine nucleotide (TG). The mutation occurring in the OT-11 and ET-6 genotypes was evaluated as a transversion-type mutation. A transition type mutation occurred in mutant line T-103 plants as well, and this mutation occurred in the form of substitution of thymine nucleotide with cytosine nucleotide (TC) at nucleotide 1114. According to the results, weed control in agriculture can be done effectively by developing varieties resistant to IMI group herbicides in quinoa plants. Herbicide resistance studies should be continued in the next generations with the ET-6, OT-11, and T-103 lines.

Sequence-Dependent Surface Coverage of ssDNA Coatings on Single-Wall Carbon Nanotubes.

A combination of experimental measurements and molecular dynamics (MD) simulations was used to investigate how the surfaces of single-wall carbon nanotubes (SWCNTs) are covered by adsorbed ssDNA oligos with different base compositions and lengths. By analyzing the UV absorption spectra of ssDNA-coated SWCNTs before and after coating displacement by a transparent surfactant, the mass ratios of adsorbed ssDNA to SWCNTs were determined for poly-T, poly-C, GT-containing, and AT-containing ssDNA oligos. Based on the measured mass ratios, it is estimated that an average of 20, 22, 26, or 32 carbon atoms are covered by one adsorbed thymine, cytosine, adenine, or guanine nucleotide, respectively. In addition, the UV spectra revealed electronic interactions of varying strengths between the nucleobase aromatic rings and the nanotube π-systems. Short poly-T DNA oligos show stronger π-π stacking interactions with SWCNT surfaces than do short poly-C DNA oligos, whereas both long poly-C and poly-T DNA oligos show strong interactions. These experiments were complemented by MD computations on simulated systems that were constrained to match the measured ssDNA/SWCNT mass ratios. The surface coverages computed from the MD results varied with oligo composition in a pattern that correlates higher measured yields of nanotube fluorescence with greater surface coverage.

A facile synthesis of iron oxide nanoparticles as a nano-sensor to detect levofloxacin and ciprofloxacin in human blood and evaluation of their biological activities.

A rapid synthesis of a pH-stable magnetic nano-sensor (iron oxide nanoparticles, Fe-NPs, ∼2.6 nm) encapsulated with 3-aminobenzoic acid (3-ABA) was achieved. 3-ABA was prepared for the first time through the reduction of 3-nitrobenzoic acid (3-NBA) in the presence of HCl and tin. Electron-impact mass spectrometry (EIMS), Fourier-transform infrared (FTIR), nuclear magnetic resonance (NMR), ultraviolet visible (UV) spectroscopy and atomic force microscopy (AFM) were used for characterization. Ten drugs, namely, ciprofloxacin (CPF), levocetirizine (LCT), levofloxacin (LVF), sulbactam sodium (SBS), ephedrine (EPH), thymine (THM), sertraline (SRT), pyridoxine (PRX), cefotaxime (CFX) and ceftriaxone (CFT) were screened with Fe-NPs. A pronounced hypsochromic shift was observed for levofloxacin and ciprofloxacin, proving that 3-ABA-coated Fe-NPs were an efficient nano-sensor for levofloxacin and ciprofloxacin up to the limit of 0.5 and 0.7 μM, respectively. The stoichiometry of the complexes was conclusively determined as 1 : 1 using Job's plot analysis. Furthermore, the drugs were successfully detected in real samples, including tap water, well water, and human blood. Moreover, the antioxidant activity, urease and lipoxygenase inhibitory potential of these nanoparticles were evaluated, exhibiting promising antioxidant potential.

Determination of isohydrafural and methyluracil in ear drops by high performance liquid chromatography

Abstract Objective The aim of current study was the development and validation of a reversed-phase high-pressure liquid chromatography (RPHPLC) method for the quantitative determination of two active ingredients, isohydrafural (IHF) and methyluracil (MU) in fixed dose combination ear drops. Methods: An efficient separation of the two compounds was achieved on a Teknokroma C1 5μm (150*4,6mm) column, with a methanol: water 60:40 (V: V) mobile phase, at 300C temperature and 0.6 mL/min flow-rate. The total analysis time was 5.5 min. Result: The verified validation parameters were: linearity, selectivity, specificity, precision (repeatability and reproducibility), robustness, limit of detection and quantification. Conclusions: Good separation and lack of interference from other chromatographic peaks, rapidanalysis times were obtained. The newly developed chromatographic method can be applied for the analysis of fixed dose combination ear drops with isohydrafural and methyluracil.

Myricetin causes site-specific DNA damage via reactive oxygen species generation by redox interactions with copper ions.

Myricetin (MYR), found in tea and berries, may have preventive effects on diseases, including Alzheimer's disease and cancer. However, MYR is also a mutagen, inducing DNA damage in the presence of metal ions. We have studied the molecular mechanisms of DNA damage by MYR in the presence of Cu(II) (MYR+Cu). Using 32P-5'-end-labeled DNA fragments, we analyzed site-specific DNA damage caused by MYR+Cu. MYR+Cu caused concentration-dependent DNA strand breaks and base alterations, leading to cleavage of DNA at thymine, cytosine, and guanine nucleotides. Formation of the oxidative DNA damage indicator, 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG), in calf thymus DNA was increased by MYR+Cu. The production of 8-oxodG in MYR-treated HL-60 cells was significantly higher than in HP100 cells, which are more resistant to H2O2 than are HL-60 cells. Reactive oxygen species (ROS) scavengers were used to elucidate the mechanism of DNA damage. DNA damage was not inhibited by typical free hydroxyl radical (•OH) scavengers such as ethanol, mannitol, or sodium formate. However, methional, catalase, and bathocuproine inhibited DNA damage induced by MYR+Cu. These results suggest that H2O2, Cu(I), and ROS other than •OH are involved in MYR+Cu-induced DNA damage. We conclude that the Cu(I)/Cu(II) redox cycle and concomitant H2O2 production via autoxidation of MYR generate a complex of H2O2 and Cu(I), probably Cu(I)-hydroperoxide, which induces oxidative DNA damage.

Endothelial Dysfunction in Adult Patients of Sickle Cell Disease

Sickle cell disease is one of the most common hemoglobinopathy worldwide. It is an autosomal recessive genetic disorder caused by replacement of adenine to thymine nucleotide in the beta chain of hemoglobin results in replacement of valine for glutamic acid at 6th position. This substitution results in formation of sickle hemoglobin (HbS) which in turn leads to a reduced lifespan of red blood cells (RBC). In hypoxic conditions, HbS has a tendency to aggregate and form fibrillar structure called tactoid within the red cells

Synthesis, antiviral evaluation, molecular docking study and cytotoxicity of 5′-phosphorylated 1,2,3-triazolyl nucleoside analogues with thymine and 6-methyl uracil moieties

Synthesis, antiviral evaluation, molecular docking study and cytotoxicity of 5′-phosphorylated 1,2,3-triazolyl nucleoside analogues with thymine and 6-methyl uracil moieties

Potential use of LIAD time-of-flight mass spectrometry for the detection of biomolecules: An example of detecting nucleobases in DNA

Deoxyribonucleic acid (DNA) carries the genetic information necessary for the synthesis of RNA and proteins; it is a biological macromolecule essential for the development and proper functioning of living organisms and is composed of nucleobases, deoxyribose, and phosphate. The four nucleobases in DNA are adenine (AD), guanine (GU), thymine (TY), and cytosine (CY). Abnormal concentrations of these four nucleobases in an organism have a significant impact on disease diagnosis. Therefore, the qualitative and quantitative detection of these DNA nucleobases in organisms is helpful to diagnose certain diseases. In this work, we report the simultaneous determination of purine (AD, GU) and pyrimidine (TY, CY) nucleobases in DNA using laser-induced acoustic desorption (LIAD) with electron ionization (EI)/time-of-flight mass spectrometry (TOFMS). The purine (MW 120 Da) samples were used as model compounds to assess the sensitivity and quantitative performance of the instrument. Its limits of detection assessed using the LIAD/EI/MS method were ∼0.5–1.2 pg under optimal conditions, and their calibration curves exhibited good linearity (R2 = 0.98). The LIAD/TOFMS was successfully applied in the simultaneous detection of AD, GU, TY, and CY in real DNA samples. The advantage of this technique is simple, fast, and without complex pre-treatment processes. In addition, a quartz-enhanced LIAD (QE-LIAD) source was used to improve the signal strength. The desorption for complex biomolecules shows that the QE-LIAD is still a “gentle” desorption source.

Transmission of poly(dT60 ) single-stranded DNA through polycarbonate track-etched ultrafiltration membranes.

The objective of this study was to examine membrane filtration of a single stranded DNA (ssDNA) with 60 thymine nucleotides, and to elucidate the variables controlling its transmission across track-etched porous membranes. Dead end filtration measurements were performed using different pore size membranes (10, 15, and 30 nm) at different transmembrane pressures in solutions with ionic strength ranging from 0 to 1000 mM NaCl. The diffusivity of the ssDNA was determined using fluorescence recovery after photobleaching, yielding hydrodynamic radii ranging from 1.6 to 2.8nm, with values decreasing with increasing solution ionic strength. Despite the small ssDNA/membrane pore size, nearly 100% rejection was observed for measurements performed with the 10 and 15 nm pore size membranes under low-ionic strength conditions. These high rejections can be attributed to strong repulsive electrostatic ssDNA-membrane interactions. With increasing ionic strength, electrostatic interactions as well as the effective size of the ssDNA decreases and the flexibility of the ssDNA increases, leading to a reduction in ssDNA rejection. A design of experiments approach was used to plan filtration experiments that adequately covered the variable space with a manageable number of experiments. The results yielded an empirical expression relating ssDNA rejection to pore size, solution ionic strength and transmembrane pressure. There was evidence of flow induced elongation at high-transmembrane pressures in the 30 nm pore size membranes, but not in the smaller pore size membranes. These results are consistent with critical flux estimates developed using a free draining model for the ssDNA.

Can Hindered Transport Models for Rigid Spheres Predict the Rejection of Single Stranded DNA from Porous Membranes?

In this paper, predictions from a theoretical model describing the rejection of a rigid spherical solute from porous membranes are compared to experimental results for a single stranded DNA (ssDNA) with 60 thymine nucleotides. Experiments were conducted with different pore size track-etched membranes at different transmembrane pressures and different NaCl concentrations. The model includes both hydrodynamic and electrostatic solute-pore wall interactions; predictions were made using different size parameters for the ssDNA (radius of gyration, hydrodynamic radius, and root mean square end-to-end distance). At low transmembrane pressures, experimental results are in good agreement with rejection predictions made using the hard sphere model for the ssDNA when the solute size is described using its root mean square end-to-end distance. When the ssDNA size is characterized using the radius of gyration or the hydrodynamic radius, the hard sphere model under-predicts rejection. Not surprisingly, the model overestimates ssDNA rejection at conditions where flow induced elongation of the DNA is expected. The results from this study are encouraging because they mean that a relatively simple hindered transport model can be used to estimate the rejection of a small DNA from porous membranes.

A photochemically covalent lock stabilizes aptamer conformation and strengthens its performance for biomedicine.

Aptamers’ vast conformation ensemble consisting of interconverting substates severely impairs their performance and applications in biomedicine. Therefore, developing new chemistries stabilizing aptamer conformation and exploring the conformation–performance relationship are highly desired. Herein, we developed an 8-methoxypsoralen-based photochemically covalent lock to stabilize aptamer conformation via crosslinking the inter-stranded thymine nucleotides at TpA sites. Systematical studies and molecular dynamics simulations were performed to explore the conformation–performance relationship of aptamers, revealing that conformation-stabilized aptamers displayed better ability to bind targets, adapt to physiological environment, resist macrophage uptake, prolong circulation half-life, accumulate in and penetrate into tumor than their counterparts. As expected, conformation-stabilized aptamers efficiently improved the therapeutic efficacy of aptamer-drug conjugation on tumor-bearing mice. Collectively, our study has developed a general, simple and economic strategy to stabilize aptamer conformation and shed light on the conformation–performance relationship of aptamers, laying a basis for promoting their basic researches and applications in biomedicine.

Nucleotide mismatches prevent intrinsic self-silencing of hpRNA transgenes to enhance RNAi stability in plants

Hairpin RNA (hpRNA) transgenes are the most successful RNA interference (RNAi) method in plants. Here, we show that hpRNA transgenes are invariably methylated in the inverted-repeat (IR) DNA and the adjacent promoter, causing transcriptional self-silencing. Nucleotide substitutions in the sense sequence, disrupting the IR structure, prevent the intrinsic DNA methylation resulting in more uniform and persistent RNAi. Substituting all cytosine with thymine nucleotides, in a G:U hpRNA design, prevents self-silencing but still allows for the formation of hpRNA due to G:U wobble base-pairing. The G:U design induces effective RNAi in 90–96% of transgenic lines, compared to 57–65% for the traditional hpRNA design. While a traditional hpRNA transgene shows increasing self-silencing from cotyledons to true leaves, its G:U counterpart avoids this and induce RNAi throughout plant growth. Furthermore, siRNAs from G:U and traditional hpRNA show different characteristics and appear to function via different pathways to induce target DNA methylation.

Tracking Infection and Genetic Divergence of Methicillin-Resistant Staphylococcus aureus at Pets, Pet Owners, and Environment Interface.

Staphylococcus aureus (S. aureus) has become a leading animal and public health pathogen that keeps on transferring from one host to other, giving rise to newer strains by genetic shifts. The current study was designed to investigate the epidemiology and genetic relatedness of mecA gene in S. aureus isolated from pets, immediate individuals in contact with pets, and veterinary clinic environments. A total of n = 300 samples were collected from different veterinary hospitals in Pakistan using convenience sampling. The collected samples were subjected to microbiological and biochemical examination for the isolation of S. aureus. Methicillin resistance was investigated by both phenotypically using oxacillin disk diffusion assay and by genotypically targeting mecA gene by PCR. PCR amplicons were subjected for sequencing by Sanger method of sequencing, which were subsequently submitted to NCBI GenBank under the accession numbers MT874770, MT874771, and MT874772. Sequence evolutionary analysis and mecA gene characterization was done using various bioinformatics tools. Overall, 33.66% mecA genes harboring S. aureus strains were isolated from all sources (33.33% from pets, 46.0% from surrounding, and 28.0% from immediate contact individuals). The bioinformatics analysis noted that one SNP was identified at position c.253C>A (Transvertion). The phylogenetic tree (two clades) of S. aureus mecA revealed a possibility of inter-transmission of disease between the environment and pets. Frequency of adenine and thymine nucleotide in motifs were found to be the same (0.334). Cytosine and guanine frequency were also the same (0.166). Threonine was replaced by asparagine (p.T84D) in each sample of cat, environment, and human. On the other hand, protein structures ofcat-1 and cat-2 proteins were found identical while cat-3, environmental, and human proteins shared identical structures. The study thus concludes rising circulation of methicillin-resistant S. aureus (MRSA) strains in animal-human-environment interfaces, forecasting the development of novel strains withmodified range of resistance.

Phytochemical and biological investigation of Litophyton arboreum

Three known compounds including, uracil (1), 5-methyl uracil (2) and uridine (3) have been isolated from the soft coral Litophyton arboreum collected from the red sea. The isolated compounds were identified using spectroscopic techniques predominately 1H-NMR and 13C-NMR analyses, in addition to HR-ESI-MS. Moreover, compounds 1-3 were in vitro estimated for their cytotoxic activity against three human cancer cell lines (A549, MCF-7 and HepG2), and antileishmanial potential against Leishmania major.

The effectiveness of the use of oxymethyl uracil on the model of acute alcohol intoxication

Introduction. Considering the prevalence of acute alcohol poisoning in Russia, it seems urgent to search for new effective means of correcting them. Along with taking measures to remove ethanol from the body, pathogenetic correction is effective. Oxymethyluracil and its derivatives have proven to be effective hepatoprotectors in various experimental models of liver damage. The aim of the research was the evaluation of the effectiveness of oxymethyl uracil on the model of acute alcohol intoxication. Material and methods. On the model of acute toxic liver injury of laboratory animals with ethanol, the efficiency of correction of pathological changes with oxymethyl uracil was studied compared to the drug “Mexidol”. A complex of biochemical, morphological and genetic studies was carried out. Results. The morphological studies showed that the correction with oxymethyluracil was more effective at both time points than the drug “Mexidol”, which was manifested in a lower intensity of damage to the liver parenchyma. In the group that received oxymethyluracil, a restoration of the frequency of expression of the Chek 1 gene was observed both after 24 and 72 hours. Upon acute exposure to ethanol, a slight decrease in the level of RIPK1 gene expression was observed. The level of expression of this gene decreased most significantly during the correction of oxy methyl uracil. A decrease in the frequency of expression of this gene can indicate a slowdown in necrosis processes and suppression of reactive oxygen species production in liver cells and, consequently, a curative effect of oxymethyluracil in this type of intoxication. Conclusion. Based on the complex biochemical, morphological and genetic studies carried out, it can be concluded that under acute exposure to ethanol, the corrective effect of oxymethyl uracil is more pronounced than Mexidol (ethylmethylhydroxypyridine succinate).

Electrochemical Detection of Oxacillin Resistance using Direct-Labeling Solid-Phase Isothermal Amplification.

Isothermal amplification reactions represent an important and exciting approach to achieve widespread, low cost, and easily implemented molecular diagnostics. This work presents a modified recombinase polymerase amplification (RPA) reaction, which can be directly coupled to a simple electrochemical measurement to ultimately allow development of a nucleic acid-based assay for antibiotic resistance genes. It is shown that use of reagents from a standard RPA reaction kit allows incorporation of horse radish peroxidase-labeled thymine nucleotides into amplified DNA strands, which can be detected via an amperometric signal readout for detection of important gene sequences. The assay is exemplified through detection of fragments of the oxacillin resistance gene in Escherichia coli cells bearing a drug resistance plasmid, achieving a potential limit of detection of 319 cfus/mL and an unoptimized time to result of 60 min. This work serves as a suitable demonstration of the potential for a system to deliver detection of key drug resistance genes at clinically relevant levels.

The Role of Folate-supplementation in Depression: A Narrative Review

Background: There is a relationship between the etiology and management of clinical depression and vitamin deficiencies. Aim: To review the effect of dietary folic acid supplementation on cause, severity and treatment of depression. Methods: A narrative review of empirical and theoretical literature on the effect of folic acid supplementation on the se-verity and treatment outcomes of depression. Result: Many enzymes and neurotransmitters depend on folic acid for optimal functions. The monoamine hypothesis of depression confirms the depletion of serotonin, dopamine, and norepinephrine. The systemic level of methyl folate is dependent upon the enzyme methylenetetrahydrofolate reductase [MTHFR], which is encoded by a polymorphic gene [C677T-MTHFR], as well as being dependent on dietary folic acid intake. However, folate has procarcinogenic properties because its coenzymes are involved in de novo purine and thymine nucleotide biosynthesis. Besides, folate deficiency in normal tissues may also predispose to neoplastic transformation, while folate supplementation may suppress the development of tumors in normal tissues. Epidemiological studies revealed that low folate status is associated with depression; especially in terms of severity and the outcome of antidepressant use. Conclusion: The role of folic acid in the etiology and the management of depression cannot be overemphasized. Folate supplementation has been yielding positive results in the management of depression.

Trustees of Columbia Univ. in City of New York v. Illumina, Inc. United States Court of Appeals of the Federal Circuit, 2021 No. 2019-2302, 2021 WL 317661 (nonprecedential)

Trustees of Columbia Univ. in City of New York v. Illumina, Inc. United States Court of Appeals of the Federal Circuit, 2021 No. 2019-2302, 2021 WL 317661 (nonprecedential)

Hg(II) Binding to Thymine Bases in DNA.

The compounds of mercury can be highly toxic and can interfere with a range of biological processes, although many aspects of the mechanism of toxicity are still obscure or unknown. One especially intriguing property of Hg(II) is its ability to bind DNA directly, making interstrand cross-links between thymine nucleobases in AT-rich sequences. We have used a combination of small molecule X-ray diffraction, X-ray spectroscopies, and computational chemistry to study the interactions of Hg(II) with thymine. We find that the energetically preferred mode of thymine binding in DNA is to the N3 and predict only minor distortions of the DNA structure on binding one Hg(II) to two cross-adjacent thymine nucleotides. The preferred geometry is predicted to be twisted away from coplanar through a torsion angle of between 32 and 43°. Using 1-methylthymine as a model, the bis-thymine coordination of Hg(II) is found to give a highly characteristic X-ray spectroscopic signature that is quite distinct from other previously described biological modes of binding of Hg(II). This work enlarges and deepens our view of significant biological targets of Hg(II) and demonstrates tools that can provide a characteristic signature for the binding of Hg(II) to DNA in more complex matrices including intact cells and tissues, laying the foundation for future studies of mechanisms of mercury toxicity.