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1.
In this study, we performed an ultraviolet-visible (UV-Vis) and circular dichroism (CD) spectroscopic analysis of the binary and ternary supramolecular structures formed by self-assembling the following three water-soluble porphyrins with and without a chiral template: the negatively charged, meso-Tetra(4-sulfonatophenyl) porphine (H2TPPS4−); the positively charged meso-trans-(di(N-methyl-4-pyridyl)diphenyl) porphine (trans-DmPyDPP) and meso-cis-(di(N-methyl-4-pyridyl)diphenyl) porphine (cis-DmPyDPP). Polyglutamic acid (both L and D enantiomers) was selected as the chiral template due to its ability to change secondary structure with pH. The propensity for the porphyrins to show an induced CD in the presence of polyglutamic acid is demonstrated. The induced chirality of all supramolecular structures was found to depend on the pH of the solution, the chirality of the polymer, and the order of addition of the positively and negatively charged porphyrins (for ternary complexes). Of particular interest is that the interaction of H2TPPS4− with the chiral scaffold seems to undergo a dynamic rearrangement of the supramolecular structure as evident from the change in the CD spectrum over time. Moreover, experiments with ternary complexes suggest that the preferential interaction of trans-DmPyDPP with the random coil of the polymer shows promise as a sensor of protein secondary structure. 相似文献
2.
Vigen G. Barkhudaryan Gayane V. Ananyan 《Journal of biomolecular structure & dynamics》2020,38(12):3489-3495
AbstractThe present paper is an overview of studying of DNA-porphyrin interactions using viscometry in combination with the spectroscopic methods. It was shown, that when porphyrins interact with DNA as an outside binder, the interaction mode and intensity does not depend on metal center, peripheral substituent’s and their positions on pyridylic ring. In case of planar porphyrins, the binding type is mainly determined by type of peripheral substituent’s and their position on the pyridylic ring. Currently, viscometry is widely used to study the interaction of porphyrins with DNA as an adjunct to other methods. Due to high accuracy and maximum sensitivity to changes in the size and shape of macromolecules, it is recommended to use viscometry as the cogent method for studying the interaction of small molecules with DNA, especially if intercalation is expected using other methods if necessary to confirm the results obtained. Abbreviations DNA deoxyribonucleic acid H2TAllPyP4 meso-tetra-(4N-allylpyridyl) porphyrin H2TAllPyP3 meso-tetra-(3N-allylpyridyl) porphyrin H2THOEtPyP4 meso-tetra-(4N-hydroxyethylpyridyl) porphyrin H2THOEtPyP3 meso-tetra-(3N-hydroxyethylpyridyl) porphyrin UV/VIS spectrophotometry ultraviolet-visible spectrophotometry CD spectroscopy circular dichroism spectroscopy Communicated by Ramaswamy H. Sarma 相似文献
3.
《Mutation Research - Genetic Toxicology and Environmental Mutagenesis》2002,513(1-2):113-120
Previous studies have demonstrated that phenolic compounds, including genistein (4′,5,7-trihydroxyisoflavone) and resveratrol (3,4′,5-trihydroxystilbene), are able to protect against carcinogenesis in animal models. This study was undertaken to examine the ability of genistein and resveratrol to inhibit reactive oxygen species (ROS)-mediated strand breaks in φX-174 plasmid DNA. H2O2/Cu(II) and hydroquinone/Cu(II) were used to cause oxidative DNA strand breaks in the plasmid DNA. We demonstrated that the presence of genistein at micromolar concentrations resulted in a marked inhibition of DNA strand breaks induced by either H2O2/Cu(II) or hydroquinone/Cu(II). Genistein neither affected the Cu(II)/Cu(I) redox cycle nor reacted with H2O2 suggest that genistein may directly scavenge the ROS that participate in the induction of DNA strand breaks. In contrast to the inhibitory effects of genistein, the presence of resveratrol at similar concentrations led to increased DNA strand breaks induced by H2O2/Cu(II). Further studies showed that in the presence of Cu(II), resveratrol, but not genistein was able to cause DNA strand breaks. Moreover, both Cu(II)/Cu(I) redox cycle and H2O2 were shown to be critically involved in resveratrol/copper-mediated DNA strand breaks. The above results indicate that despite their similar in vivo anticarcinogenic effects, genistein and resveratrol appear to exert different effects on oxidative DNA damage in vitro. 相似文献
4.
Sandrine Pereira Isabelle Cavalie Virginie Camilleri Rodolphe Gilbin Christelle Adam-Guillermin 《Mutation Research - Genetic Toxicology and Environmental Mutagenesis》2013,750(1-2):19-26
Aluminium is a toxic metal whose genotoxicity has been scarcely studied in aquatic species and more generally in mammals. Recently, human and ecological disaster caused by the discharge of red mud in Hungary has revived questions about the toxicity of this metal particularly for the environment. On the contrary, cadmium is a highly toxic metal whose genotoxicity has been well characterized in various mammalian cells. However on non-human cells, little is known about its impact on DNA damage and repair.In this study, the genotoxic potential of both metals on embryonic zebrafish cells ZF4 was analyzed and particularly the impairment of the major DNA double strand breaks (DSB)-repair pathway, i.e. non-homologous end-joining (NHEJ).To this aim, DNA single strand breaks (SSB) and DSB were evaluated using the comet assay and the immunodetection of γ-H2AX proteins, respectively, in AlCl3 or CdCl2 exposed ZF4 cells. These exposures result in the production of DSBs a few hours after incubation. The DNA-PK kinase activity, essential for NHEJ, is more affected by the presence of aluminium than cadmium. Altogether our data provide evidence of the high toxicity induced by aluminium in zebrafish and indicates the pertinence of genotoxicity evaluation in organisms living in contaminated water. 相似文献
5.
Effects of growth phase and repair capacity on rejoining of ethyl methanesulfonate-induced DNA breaks in Escherichia coli 总被引:1,自引:0,他引:1
The effects of growth phase and DNA repair capacity on the production and rejoining of ethyl methanesulfonate (EMS)-induced single-strand breaks were studied in 4 strains of E. coli. DNAs from logarithmic and stationary phase cells of the DNA polymerase I deficient mutant, P3478 polA, a recombination deficient mutant, DZ417recA, and from the respective parental strains, W3110pol+ and AB253rec+ were examined by sedimentation in alkaline sucrose gradients.In both parental strains, stationary phase cells exhibited enhanced strand rejoining. In the mutants, alkylated DNA was repaired to some extent in both growth phases, but it contained a greater proportion of small DNA fragments compared to the parental strains. Some DNA breakdown occured in all four strains but this was most extensive in stationary phase cells of the repair-deficient mutants.These results indicate that the four strains can rejoin EMS-induced DNA strand breaks with varying efficiency depending on the physiological state and the genetic capacity for repair. 相似文献
6.
《The International journal of biochemistry》1994,26(5):631-637
- 1.1. The interaction of haemopexin and albumin with TPPS4 was studied by measuring the absorption and fluorescence spectra. Haemopexin was found to have one strong TPPS4 binding center (Ka = 3 × 107M−1).
- 2.2. Haem-haemopexin complex appears to have no specific binding site for TPPS4. Occupation of the specific binding center of haemopexin molecule by a haem abolishes TPPS4 binding.
- 3.3. Albumin was found to possess one strong TPPS4 binding center (Ka = 3 × 106M−1) besides two or three weak binding sites (Ka = 2 × 105M−1).
- 4.4. Haern-albumin complex possesses only one weak TPPS4 binding site (Ka = 7 × lO5M−1). These observations suggest identity of primary binding sites of TPPS4 and haem on albumin molecule.
7.
Kenzo Yamanaka Akira Hasegawa Ryoji Sawamura Shoji Okada 《Biological trace element research》1989,21(1):413-417
DNA damage induced by administration of dimethylarsinic acid (DMAA) to rats and mice was investigated. At 12 h after administration of DMAA, DNA single-strand breaks were induced markedly in lung. The majority of dimethylarsine, one of the main metabolites, in the expired air was excreted within 6–18 h after administration of DMAA to rats. In vitro experiments using nuclei isolated from lung of mice indicated that DNA strand breaks were caused by dimethylarsine. Furthermore, the strand breaks after exposure to dimethylarsine were reduced in the presence of catalase and/or superoxide dismutase. These results strongly suggest that the strand breaks are induced not by dimethylarsine itself but by active oxygen, e.g., O 2 ? and ·OH, produced both by dimethylarsine and molecular oxygen. When DNA was exposed to dimethylarsine, thiobarbituric acid (TBA)-reactive intermediates andcis-thymine glycol were produced. Dimethylarsine appears to induce DNA damage by the mechanism similar to the damage produced by ionizing radiation. 相似文献
8.
Masayasu Sugiyama 《Biological trace element research》1989,21(1):399-404
The induction of DNA single strand breaks by carcinogenic chromate compounds has been found to be altered by vitamin E and vitamin B2. Pretreatment with vitamin E for 24 h prior to exposure to Na2CrO4 resulted in a decrease of chromate-induced DNA single strand breaks, whereas similar treatment with vitamin B2 enhanced levels of breaks induced by chromate. In contrast, levels of DNA protein crosslinks, the other major lesion induced, were not affected by vitamin E or vitamin B2. The uptake of Na2CrO4 was not affected by pretreatment with these vitamins. The role of vitamins in chromate-induced DNA damages is discussed. 相似文献
9.
The nuclear and mitochondrial DNA of the amoebae of the cellular slime mold Dictyostelium discoideum have been labeled with [methyl-3H]thymidine by allowing them to grow on Escherichia coli 15T? containing this label in its DNA. Neutral CsCl gradients were used to identify the labeled molecules. Alkaline sucrose sedimentation profiles of cells lysed directly on the gradients revealed two high molecular weight species, one of about 90 S (single-strand mol wt = 1.4 × 108) identified by alkaline CsCl rebanding as nuclear DNA, and another of 43 S (single-strand mol wt = 2.3 × 107), identified as mitochondrial DNA. These alkaline sucrose gradients were used to study the production of single-strand breaks and their rejoining in DNA of a gamma ray-resistant strain (NC-4; 10% survival dose for cell proliferation, D10 = 300 krad) and in two radiation-sensitive daughter mutants (γs-18, D10 = 75 krad; γs-13, D10 = 4 krad). With 60Co gamma rays, breaks were produced in nuclear and mitochondrial DNA at an efficiency of one break per 33 eV in all three strains. At doses up to about 100 krad, these single-strand breaks were closed equally well during post-irradiation incubation of NC-4, γs-18 and γs-13, even though their survivals were widely different, indicating no apparent correlation between parental strand rejoining and survival in the sensitive strains. At higher doses, post-irradiation treatment with 1 mg caffeine/ml sensitized NC-4 and retarded strand-rejoining, suggesting that lethality in this resistant strain may be related to strand breaks. It is concluded that single-strand rejoining is a necessary, but not sufficient, condition for radiation survival in this organism. The nature of the apparently unrepaired lesions leading to lethality in the sensitive strains is not known. 相似文献
10.
DNA damage responses are important for the maintenance of genome stability and the survival of organisms. Such responses are activated in the presence of DNA damage and lead to cell cycle arrest, apoptosis, and DNA repair. In Caenorhabditis elegans, double-strand breaks induced by DNA damaging agents have been detected indirectly by antibodies against DSB recognizing proteins. In this study we used a comet assay to detect DNA strand breaks and to measure the elimination of DNA strand breaks in mitotic germline nuclei of C. elegans. We found that C. elegans brc-1 mutants were more sensitive to ionizing radiation and camptothecin than the N2 wild-type strain and repaired DNA strand breaks less efficiently than N2. This study is the first demonstration of direct measurement of DNA strand breaks in mitotic germline nuclei of C. elegans. This newly developed assay can be applied to detect DNA strand breaks in different C. elegans mutants that are sensitive to DNA damaging agents. 相似文献
11.
Telomeric C-rich strands can form a noncanonical intercalated DNA structure known as an i-motif. We have studied the interactions of the cationic porphyrin 5,10,15,20-tetra-(N-methyl-4-pyridyl)porphine (TMPyP4) with the i-motif forms of several oligonucleotides containing telomeric sequences. TMPyP4 was found to promote the formation of the i-motif DNA structure. On the basis of (1)H NMR studies, we have created a model of the i-motif-TMPyP4 complex that is consistent with all the available experimental data. Two-dimensional NOESY data prompted us to conclude that TMPyP4 binds specifically to the edge of the intercalated DNA core by a nonintercalative mechanism. Since we have shown that TMPyP4 binds to and stabilizes the G-quadruplex form of the complementary G-rich telomeric strand, this study raises the intriguing possibility that TMPyP4 can trigger the formation of unusual DNA structures in both strands of the telomeres, which may in turn explain the recently documented biological effects of TMPyP4 in cancer cells. 相似文献
12.
Early events in the cellular response to DNA damage, such as double strand breaks, rely on lesion recognition and activation of proteins involved in maintenance of genomic stability. One important component of this process is the phosphorylation of the histone variant H2AX. To investigate factors explaining the variation in carcinogenic potency between different categories of polycyclic aromatic hydrocarbons (PAHs), we have studied the phosphorylation of H2AX (H2AXγ). A549 cells were exposed to benzo[a]pyrene diol epoxide [(+)-anti-BPDE] (a bay-region PAH) and dibenzo[a,l]pyrene diol epoxide [(−)-anti-DBPDE] (a fjord-region PAH) and H2AXγ was studied using immunocytochemistry and Western blot. Hydrogen peroxide (H2O2) was used to induce oxidative DNA damage and strand breaks. As showed with single cell gel electrophoresis, neither of the diol epoxides resulted in DNA strand breaks relative to H2O2. Visualisation of H2AXγ formation demonstrated that the proportion of cells exhibiting H2AXγ staining at 1 h differed between BPDE, 40% followed by a decline, and DBPDE, <10% followed by an increase. With H2O2 treatment, almost all cells demonstrated H2AXγ at 1 h. Western blot analysis of the H2AXγ formation also showed concentration and time-dependent response patterns. The kinetics of H2AXγ formation correlated with the previously observed kinetics of elimination of BPDE and DBPDE adducts. Thus, the extent of H2AXγ formation and persistence was related to both the number of adducts and their structural features. 相似文献
13.
《Biochimica et Biophysica Acta (BBA) - Gene Structure and Expression》1997,1354(3):252-260
A series of meso-5,10,15-tris(N-methyl-4-pyridiniumyl)-20-(4-alkylamidophenyl) porphyrins were synthesized by derivatizing the amino group on the phenyl ring with the following hydrophobic groups: –C(O)C7F15, –C(O)CHCH2, C(O)CH3, –C(O)C7H15, and –C(O)C15H31. The cationic tris-pyridiumyl porphyrin core serves as a DNA binding motif and a photosensitizer to photomodify DNA molecules. The changes of the UV–Vis absorption spectra during the titration of these porphyrins with calf thymus DNA revealed a large bathochromic shift (up to 14 nm) and a hypochromicity (up to 55%) of the porphyrins Soret bands, usually considered as proof of porphyrin intercalation into DNA. Association constants (K) calculated according to the McGhee and von Hippel model, were in the range of 106–107 M−1. An increase in hydrophobicity of the substituents at the 20−meso-position produced higher binding affinity. These porphyrins caused photomodification of the supercoiled plasmid DNA when a green laser beam at 532 nm was applied. Those with higher surface activity acted more efficiently as DNA photomodifiers. The porphyrin with a perfluorinated alkyl chain (–COC7F15) at the meso-20-position inhibited the growth of gram-positive bacteria (S. aureus, or S. epidermidis). Other porphyrins exhibited moderate activity against both gram-negative and gram-positive organisms. 相似文献
14.
The amino acid histidine was found to increase the toxicity of H2O2 in cultured mammalian cells. Histidine also augmented the level of DNA single strand breaks (SSB) detectable in cells exposed to the oxidant and, in addition, resulted in the appearance of DNA double strand breaks (DSB), a lesion which is not produced by H2O2 alone. 相似文献
15.
Christoph Krettler Carville G. Bevans Christoph Reinhart Matthias Watzka Johannes Oldenburg 《Analytical biochemistry》2015
Use of the reductant dithiothreitol (DTT) as a substrate for measuring vitamin K 2,3-epoxide reductase (VKOR) activity in vitro has been reported to be problematic because it enables side reactions involving the vitamin K1 2,3-epoxide (K1>O) substrate. Here we characterize specific problems when using DTT and show that tris(3-hydroxypropyl)phosphine (THPP) is a reliable alternative to DTT for in vitro assessment of VKOR enzymatic activity. In addition, the pH buffering compound imidazole was found to be problematic in enhancing DTT-dependent non-enzymatic side reactions. Using THPP and phosphate-based pH buffering, we measured apparent Michaelis–Menten constants of 1.20 μM for K1>O and 260 μM for the active neutral form of THPP. The Km value for K1>O is in agreement with the value that we previously obtained using DTT (1.24 μM). Using THPP, we successfully eliminated non-enzymatic production of 3-hydroxyvitamin K1 and its previously reported base-catalyzed conversion to K1, both of which were shown to occur when DTT and imidazole are used as the reductant and pH buffer, respectively, in the in vitro VKOR assay. Accordingly, substitution of THPP for DTT in the in vitro VKOR assay will ensure more accurate enzymatic measurements and assessment of warfarin and other 4-hydroxycoumarin inhibition constants. 相似文献
16.
The cytotoxic and DNA-damaging effects of cyclophosphamide (CP) and its ‘activated’ derivative 4-OOH-CP were studied using a series of strains of S. cerevisiae which allow a phenotypical classification of genotoxic characteristics as well as direct physicochemical demonstration of key DNA lesions. The concurring results of biological and biochemical experiments indicate that (i) non-activated CP has a weak but detectable monofunctional alkylating potency, leading to DNA strand breaks and (ii) 4-OOH-CP has the ability to induce both DNA strand breaks and interstrand cross-links. The activity of CP is probably due to spontaneous decomposition in aqueous solution. 相似文献
17.
Non homologous end joining (NHEJ) is an important process that repairs double strand DNA breaks (DSBs) in eukaryotic cells. Cells defective in NHEJ are unable to join chromosomal breaks. Two different NHEJ assays are typically used to determine the efficiency of NHEJ. One requires NHEJ of linearized plasmid DNA transformed into the test organism; the other requires NHEJ of a single chromosomal break induced either by HO endonuclease or the I-SceI restriction enzyme. These two assays are generally considered equivalent and rely on the same set of NHEJ genes. PC4 is an abundant DNA binding protein that has been suggested to stimulate NHEJ. Here we tested the role of PC4''s yeast homolog SUB1 in repair of DNA double strand breaks using different assays. We found SUB1 is required for NHEJ repair of DSBs in plasmid DNA, but not in chromosomal DNA. Our results suggest that these two assays, while similar are not equivalent and that repair of plasmid DNA requires additional factor(s) that are not required for NHEJ repair of chromosomal double-strand DNA breaks. Possible roles for Sub1 proteins in NHEJ of plasmid DNA are discussed. 相似文献
18.
Masaki Fukuyo Toshiaki Nakano Yingbiao Zhang Yoshikazu Furuta Ken Ishikawa Miki Watanabe-Matsui Hirokazu Yano Takeshi Hamakawa Hiroshi Ide Ichizo Kobayashi 《Nucleic acids research》2015,43(5):2841-2852
The restriction-modification systems use epigenetic modification to distinguish between self and nonself DNA. A modification enzyme transfers a methyl group to a base in a specific DNA sequence while its cognate restriction enzyme introduces breaks in DNA lacking this methyl group. So far, all the restriction enzymes hydrolyze phosphodiester bonds linking the monomer units of DNA. We recently reported that a restriction enzyme (R.PabI) of the PabI superfamily with half-pipe fold has DNA glycosylase activity that excises an adenine base in the recognition sequence (5′-GTAC). We now found a second activity in this enzyme: at the resulting apurinic/apyrimidinic (AP) (abasic) site (5′-GT#C, # = AP), its AP lyase activity generates an atypical strand break. Although the lyase activity is weak and lacks sequence specificity, its covalent DNA–R.PabI reaction intermediates can be trapped by NaBH4 reduction. The base excision is not coupled with the strand breakage and yet causes restriction because the restriction enzyme action can impair transformation ability of unmethylated DNA even in the absence of strand breaks in vitro. The base excision of R.PabI is inhibited by methylation of the target adenine base. These findings expand our understanding of genetic and epigenetic processes linking those in prokaryotes and eukaryotes. 相似文献
19.
Although organism-specific factors related to individual indicator organisms have hampered the use of bioassays for the evaluation of environmental risk in practice, the importance of understanding organism-specific factors when selecting model organisms has also not yet been fully recognized. In this work, genotoxicity was evaluated in the ciliated protozoa, Euplotes vannus and Pseudokeronopsis rubra, when exposed to graded doses of nitrofurazone for several discrete durations. Genotoxicity was expressed based on the LD50 and was determined by assessing DNA strand breaks (through alkaline comet assay) and DNA–protein crosslinks (DPCs), by means of a KCl–SDS precipitation assay. It was found that E. vannus generally had lower LD50's than P. rubra (P < 0.05), and that the LD50 values decreased in both ciliates as the exposure durations increased. Compared to the control groups, the nitrofurazone treated E. vannus generally produced more DNA strand breaks (P < 0.05), but for DPCs (P > 0.05). The relationship between these parameters was reversed in the case of P. rubra. Biphasic dose–response relationships were generally detected between nitrofurazone and genotoxicity parameters, however, parameters for DNA strand breaks presented significantly positive correlations between each other (P < 0.05), but showed nearly no significant correlations with DPC induction. In brief, our findings confirmed nitrofurazone-induced genotoxicity and the important role of organism-specific factors in the selection of model organisms from ciliated protozoa for environmental monitoring and risk assessment in aquaculture. 相似文献
20.
Catecholestrogens [4-hydroxyestradiol (4-OHE2)] have been implicated in human carcinogenesis, although the mechanism remains unestablished. In this study pUC 18 plasmid DNA was modified with 4-OHE2 and nitric oxide (NO). The modification induced in native DNA exhibited hyperchromicity, single strand breaks, damage to restriction sites, modification of bases, decrease in Tm and change in ellipticity. Modified DNA was found to be highly immunogenic in experimental animal, eliciting high titer antibodies. Circulating cancer autoantibodies showed preferable recognition of 4-OHE2-NO-DNA over native form (p < 0.001) and the oxidative epitopes on the DNA isolates from cancer patients were immunochemically detected by using experimentally induced anti-4-OHE2-NO-DNA antibodies as a probe. Preferential recognition of 4-OHE2-NO-DNA by cancer autoantibodies coupled with enhanced binding of induced antibodies to DNA isolated from cancer patients is an indicative of oxidative stress induced DNA damage in cancer. Possible involvement of unique epitopes on modified DNA in cancer autoantibody induction has been suggested. 相似文献