-
International Journal of Molecular... May 2023When modified uridine derivatives are incorporated into DNA, radical species may form that cause DNA damage. This category of molecules has been proposed as...
When modified uridine derivatives are incorporated into DNA, radical species may form that cause DNA damage. This category of molecules has been proposed as radiosensitizers and is currently being researched. Here, we study electron attachment to 5-bromo-4-thiouracil (BrSU), a uracil derivative, and 5-bromo-4-thio-2'-deoxyuridine (BrSdU), with an attached deoxyribose moiety via the N-glycosidic (N1-C) bond. Quadrupole mass spectrometry was used to detect the anionic products of dissociative electron attachment (DEA), and the experimental results were supported by quantum chemical calculations performed at the M062X/aug-cc-pVTZ level of theory. Experimentally, we found that BrSU predominantly captures low-energy electrons with kinetic energies near 0 eV, though the abundance of bromine anions was rather low compared to a similar experiment with bromouracil. We suggest that, for this reaction channel, proton-transfer reactions in the transient negative ions limit the release of bromine anions.
Topics: Electrons; Deoxyribose; Bromine; Anions; Bromodeoxyuridine
PubMed: 37240053
DOI: 10.3390/ijms24108706 -
Zeitschrift Fur Naturforschung. C,... 1992Bifilarly BU-substituted ColE 1 plasmid and monofilarly BU-substituted M 13 phage DNA were irradiated with UV light of 313 nm. Using agarose gel electrophoresis and...
Bifilarly BU-substituted ColE 1 plasmid and monofilarly BU-substituted M 13 phage DNA were irradiated with UV light of 313 nm. Using agarose gel electrophoresis and "reversed phase" HPLC technique ssb, dsb induction and uracil formation, respectively, could be detected in the irradiated DNA in dependence on the UV fluence. The analysis of the strandbreaks in bifilar ColE 1 DNA shows a significant part of directly induced dsb. Cross sections of ssb induction from 4.1 m2/J x 10(7) in 28%, 3.9 m2/J x 10(7) in 55% and 3.1 m2/J x 10(7) in 85-90% BU-substituted DNA were calculated. The cross section for dsb induction was found to be 0.04 m2/J x 10(7), estimated from the linear part of the fluence effect curve. In monofilar M 13 DNA a linear fluence effect curve for dsb induction was obtained. Excluding other than the direct production of dsb by using an in vitro approach for M 13 DNA, the results strongly support the hypothesis that dsb can be induced by one photochemical absorption event. The cross section for ssb was 3.8 m2/J x 10(7) and for dsb 0.05 m2/J x 10(7) in 41.5% monofilarly BU-substituted M 13 DNA. The comparison of ssb, dsb, and uracil production in bifilar and monofilar DNA with similar BU substitution showed no significant difference between the two DNA systems (ColE 1, M 13), indicating that the location of BU molecules in one or in both DNA strands will not lead to a different number of lesions after UV313 exposure.
Topics: Bromouracil; Coliphages; DNA Damage; DNA, Viral; Dose-Response Relationship, Radiation; Kinetics; Mathematics; Plasmids; Ultraviolet Rays
PubMed: 1590888
DOI: 10.1515/znc-1992-3-415 -
Bioorganic & Medicinal Chemistry Jan 2019Electron transfer through π-stacked arrays of double-stranded DNA contributes to the redox chemistry of bases, including guanine oxidation and thymine-thymine dimer...
Electron transfer through π-stacked arrays of double-stranded DNA contributes to the redox chemistry of bases, including guanine oxidation and thymine-thymine dimer repair by photolyase. 5-Bromouracil is an attractive photoreactive thymine analogue that can be used to investigate electron transfer in DNA, and is a useful probe for protein-DNA interaction analysis. In the present study using U we found that UV irradiation facilitated electron injection from mitochondrial transcription factor A into DNA. We also observed that this electron injection could lead to repair of a thymine-thymine dimer.
Topics: Base Sequence; Bromouracil; DNA; DNA Repair; DNA-Binding Proteins; Electrons; Humans; Mitochondrial Proteins; Promoter Regions, Genetic; Protein Binding; Pyrimidine Dimers; Transcription Factors; Ultraviolet Rays
PubMed: 30552005
DOI: 10.1016/j.bmc.2018.11.044 -
Yakugaku Zasshi : Journal of the... Aug 2002In 1993, there were 18 acute deaths in Japanese patients who had the viral disease herpes zoster and were treated with the new antiviral drug sorivudine (SRV,... (Review)
Review
[Molecular toxicological mechanism of the lethal interactions of the new antiviral drug, sorivudine, with 5-fluorouracil prodrugs and genetic deficiency of dihydropyrimidine dehydrogenase].
In 1993, there were 18 acute deaths in Japanese patients who had the viral disease herpes zoster and were treated with the new antiviral drug sorivudine (SRV, 1-beta-D-arabinofuranosyl-(E)-5-(2-bromovinyl)uracil). All the dead patients had received a 5-fluorouracil (5-FU) prodrug as anticancer chemotherapy concomitant with SRV administration. Studies on toxicokinetics in rats and on hepatic dihydropyrimidine dehydrogenase (DPD), a rate-limiting enzyme for 5-FU catabolism in rats and humans, strongly suggested that in the patients who received both SRV and the 5-FU prodrug, tissue levels of highly toxic 5-FU markedly increased as a result of irreversible inactivation of DPD in the presence of NADPH by 5-(2-bromovinyl)uracil (BVU), a metabolite formed from SRV by gut flora in rats and humans. Recombinant human (h) DPD was also irreversibly inactivated by [14C] BVU in the presence of NADPH. MALDI-TOF MS analysis of radioactive tryptic fragments from the radiolabeled and inactivated hDPD demonstrated that a Cys residue located at position 671 in the pyrimidine-binding domain of hDPD was modified with an allyl bromide type of reactive metabolite, dihydro-BVU. Thus artificial DPD deficiency caused by BVU from SRV led to patient deaths when coadministered with the 5-FU prodrug. Human population studies using healthy volunteers have demonstrated that there are poor and extensive 5-FU metabolizers who have very low and high DPD activities, respectively. Administration of a clinical dose of 5-FU or its prodrug to poor 5-FU metabolizers may cause death unless DPD activity is determined using their peripheral blood mononuclear cells prior to the administration of the anticancer drug.
Topics: Animals; Antiviral Agents; Arabinofuranosyluracil; Bromouracil; Dihydrouracil Dehydrogenase (NADP); Drug Interactions; Drug Therapy, Combination; Fluorouracil; Humans; NADP; Oxidoreductases; Prodrugs; Rats
PubMed: 12187768
DOI: 10.1248/yakushi.122.527 -
European Journal of Histochemistry : EJH 2003In order to localize at EM level the sites of transcription of both pre-mRNA and pre-rRNA, we have detected the DNA/RNA hybrid molecules and m3Gcapped structures by...
In order to localize at EM level the sites of transcription of both pre-mRNA and pre-rRNA, we have detected the DNA/RNA hybrid molecules and m3Gcapped structures by means of specific antibodies after short bromo-uridine (BrU) incorporation. In addition, the sections have been stained by a selective RNA stain, terbium citrate. Our data indicate that perichromatin fibrils incorporate BrU and are labeled by the anti-hybrid probe; this supports the idea that they are the pre-mRNA transcription sites. On the contrary, interchromatin granules do not incorporate BrU after short pulses and are not labeled by the anti-hybrid probe. Concerning the nucleolus, anti-hybrid and anti-BrdU antibodies colocalize only on the dense fibrillar component, suggesting that this is the site of rRNA transcription. Interestingly, the dense fibrillar component and the granular component, after specific RNA staining, show remarkable structural similarities, both containing fibrogranular RNA structures.
Topics: Animals; Antibodies; Bromodeoxyuridine; Bromouracil; Cells, Cultured; DNA; Fibroblasts; Humans; Microscopy, Electron; RNA; Rats; Transcription, Genetic; Uridine
PubMed: 14514409
DOI: 10.4081/827 -
Methods (San Diego, Calif.) May 2014Gene expression studies commonly examine total cellular RNA, which only provides information about its steady-state pool of RNA. It remains unclear whether differences...
Gene expression studies commonly examine total cellular RNA, which only provides information about its steady-state pool of RNA. It remains unclear whether differences in the steady-state reflects variable rates of transcription or RNA degradation. To specifically monitor RNA synthesis and degradation genome-wide, we developed Bru-Seq and BruChase-Seq. These assays are based on metabolic pulse-chase labeling of RNA using bromouridine (Bru). In Bru-Seq, recently labeled RNAs are sequenced to reveal spans of nascent transcription in the genome. In BruChase-Seq, cells are chased in uridine for different periods of time following Bru-labeling, allowing for the isolation of RNA populations of specific ages. Here we describe these methodologies in detail and highlight their usefulness in assessing RNA synthesis and stability as well as splicing kinetics with examples of specific genes from different human cell lines.
Topics: Animals; Bromouracil; Codon, Nonsense; DNA, Complementary; Frameshift Mutation; Genome, Human; HeLa Cells; High-Throughput Nucleotide Sequencing; Humans; Hyaluronan Receptors; K562 Cells; Kinetics; Molecular Sequence Annotation; RNA Splicing; RNA Stability; RNA, Messenger; Retinoblastoma Protein; Sequence Analysis, RNA; Staining and Labeling; Tumor Suppressor Protein p53; Uridine
PubMed: 23973811
DOI: 10.1016/j.ymeth.2013.08.015 -
Proceedings of the National Academy of... Oct 1970Synchronously replicating chromosomes in germinating spores of Bacillus subtilis were labeled near the origin with [(3)H]bromouracil. The label appeared in heavy-light...
Synchronously replicating chromosomes in germinating spores of Bacillus subtilis were labeled near the origin with [(3)H]bromouracil. The label appeared in heavy-light DNA. When cell growth and chromosome replication were continued in unlabeled bromouracil, nearly all the tritium label was transferred to heavy-heavy DNA while the terminus remained unreplicated. This implies that both origins of the replicating chromosome can undergo reinitiation. Therefore, during multifork replication the chromosome takes on the symmetric "dichotomous" form rather than the asymmetric configuration predicted by the rolling circle model.
Topics: Bacillus subtilis; Bromouracil; Carbon Isotopes; Chromosomes; DNA Replication; DNA, Bacterial; Models, Biological; Spores, Bacterial; Templates, Genetic; Thymine; Tritium
PubMed: 5002094
DOI: 10.1073/pnas.67.2.717 -
Scientific Reports Aug 20236-Thioguanine is an immunosuppressive drug, an analogue of guanine, applied to treat acute leukemia and inflammatory bowel disease. Excessive use of 6-thioguanine during...
6-Thioguanine is an immunosuppressive drug, an analogue of guanine, applied to treat acute leukemia and inflammatory bowel disease. Excessive use of 6-thioguanine during clinical treatment may cause side effects. Moreover, providing a dose too low will be ineffective. Therefore, there is a critical need for a rapid, selective and routine approach to quantifying 6-thioguanine in body fluids to support a clinical application. A fully validated HPLC method has been developed to determine 6-thioguanine in whole blood samples using 5-bromouracil as an internal standard. 6-Thioguanine nucleotides were released from erythrocytes by perchloric acid, and then hydrolysed at 100 °C to the parent thiopurine, 6-thioguanine. The following validation parameters of the method were determined: specificity/selectivity, linearity range (479-17,118 ng/mL, R > 0.992), limits of detection (150 ng/mL) and quantification (479 ng/mL), accuracy (- 5.6 < Bias < 14.7), repeatability (CV 1.30-3.24%), intermediate precision (CV 4.19-5.78%), extraction recovery (79.1-103.6%) and carryover. Furthermore, the stability of the drug in whole blood samples under various storage conditions was investigated. The suggested method is suitable for determining 6-thioguanine in whole blood erythrocyte samples for drug level monitoring, thus correct dosing.
Topics: Thioguanine; Chromatography, High Pressure Liquid; Erythrocytes; Body Fluids; Bromouracil
PubMed: 37644112
DOI: 10.1038/s41598-023-41426-5 -
Bioorganic & Medicinal Chemistry Jan 2018Given that our knowledge of DNA repair is limited because of the complexity of the DNA system, a technique called UVA micro-irradiation has been developed that can be...
Given that our knowledge of DNA repair is limited because of the complexity of the DNA system, a technique called UVA micro-irradiation has been developed that can be used to visualize the recruitment of DNA repair proteins at double-strand break (DSB) sites. Interestingly, Hoechst 33258 was used under micro-irradiation to sensitize 5-bromouracil (U)-labelled DNA, causing efficient DSBs. However, the molecular basis of DSB formation under UVA micro-irradiation remains unknown. Herein, we investigated the mechanism of DSB formation under UVA micro-irradiation conditions. Our results suggest that the generation of a uracil-5-yl radical through electron transfer from Hoechst 33258 to U caused DNA cleavage preferentially at self-complementary 5'-AAUU-3' sequences to induce DSB. We also investigated the DNA cleavage in the context of the nucleosome to gain a better understanding of UVA micro-irradiation in a cell-like model. We found that DNA cleavage occurred in both core and linker DNA regions although its efficiency reduced in core DNA.
Topics: Bisbenzimidazole; Bromouracil; DNA; DNA Breaks, Double-Stranded; DNA Cleavage; Dose-Response Relationship, Drug; Free Radicals; Molecular Structure; Structure-Activity Relationship; Ultraviolet Rays
PubMed: 29170027
DOI: 10.1016/j.bmc.2017.11.011 -
Journal of Bacteriology Sep 1990An 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...
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.
Topics: Bromodeoxyuridine; Chromatography, High Pressure Liquid; DNA Glycosylases; DNA, Bacterial; Dose-Response Relationship, Radiation; Escherichia coli; Mutation; N-Glycosyl Hydrolases; Phenotype; Ultraviolet Rays; Uracil-DNA Glycosidase
PubMed: 2203748
DOI: 10.1128/jb.172.9.5278-5285.1990