Effect of alkylated and intercalated DNA on the generation of superoxide anion by riboflavin (original) (raw)
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Effect of riboflavin and light on the secondary structure of DNA
Chemico-Biological Interactions, 1984
S, nuclease hydrolysis and benzoylated naphthoylated DEAE cellulose (BND-cellulose) chromatography have been used to study the effect of riboflavin and visible light on DNA. Native calf thymus DNA was incubated with nboflawn In the presence of fluorescent light for various time periods and subjected to S, nuclease hydrolysis. An Increasing degree of DNA degradation was seen suggesting a destabflization of the secondary structure. A decrease in melting temperature was also observed Incubation with riboflavin and illumination caused adherence to BND-cellulose indmatmg the production of single stranded regions or breaks in the native double stranded molecules. However, when incubation was done in dark and in the presence of triplet excited state quencher, potassium iodide, a reduced adherence of DNA to BND-cellulose was seen. Plasmid pBR322 DNA was also treated with nboflavm under these conditions and subjected to agarose gel electrophoresm. No degradation could be seen in dark incubated and potassmm iodide treated samples. These results indicate that the adherence of DNA to BND-cellulose in dark is possibly due to the binding of aromatic residues to the resin suggesting the formation of a complex between riboflavin and DNA.
Journal of photochemistry and photobiology. B, Biology, 2015
Determination of the superoxide dismutase activity is an important issue in the fields of biochemistry and the medical sciences. In the riboflavin/nitro blue tetrazolium (B2/NBT) method, the light sources used for generating superoxide anion radicals from light-excited riboflavin are normally fluorescent lamps. However, the conditions of B2/NBT experiments vary. This study investigated the effect of the light source on the light-excitation of riboflavin. The effectiveness of the photolysis was controlled by the wavelength of the light source. The spectra of fluorescent lamps are composed of multiple colour lights, and the emission spectra of fluorescent lamps made by different manufacturers may vary. Blue light was determined to be the most efficient for the photochemical reaction of riboflavin in visible region. The quality of the blue light in fluorescent lamps is critical to the photo-decomposition of riboflavin. A blue light is better than a fluorescent lamp for the photo-decomp...
Riboflavin (Vitamin B 2), one of the most important hydrosoluble vitamins, is a constitutive part of two coenzymes: flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD), being involved in redox processes occuring in humans. Using chemilluminescence system luminol -hydrogen peroxide, in Tris-HCl buffer, pH=8.5, in the present study the action of different concentrations of Cys, Arg, Lys and His upon the antioxidative activity of the riboflavin has been evidenced. It was found that Cys, Arg, Lys increase while, Arg decreases the riboflavin antioxidative activity. The effect of these amino acids upon the riboflavin fluorescence has also been investigated. A reaction mechanism is also proposed and the results are discussed with relevance to the redox processes of riboflavin.
Biochemistry, 1982
The multiplicity of damages induced in cellular DNA by treatment with ultraviolet light or with some chemical agents interferes with the assignment of specific types of damages in the DNA to the various biological effects expressed in the treated cells. This problem was approached by developing a series of light-induced alkylation reactions with 2-propanol which led to a single-type chemically defined product of each of the bases in nucleic acids. In the work described here, these reactions were applied to synthetic deoxyribopolynucleotides and to the various forms of 4X 174 DNA in order to study the effect of sequence and secondary structure on the reactivity of the bases in these reactions. When a series of deoxyribodinucleoside monophosphates and ordered homo-and heterodeoxyribopolynucleotides was used, it was shown that purines having 5'-neighboring pyrimidines are more reactive than those having 5'-neighboring purines. It was thus concluded that the middle purines in the sequences Py-Pu-Py or Py-Pu-Pu are more reactive than those in the sequences Pu-Pu-Py or Pu-Pu-Pu. The effect of the secondary structure of DNA on the photoalkylation reaction has been studied by comparing the reactivities of the bases in the following 4x1 74 DNA substrates: single-stranded DNA, whole phage particles, and double-stranded replicative forms (RFI E x p o s u r e of living cells to radiation or some chemical agents has deleterious consequences which are caused mainly by damage induced in their DNA. This damage may cause local destruction or local alterations of the genetic information, which may then be expressed as mutations, initiation of carcinogenesis, or inactivation . One of the main factors that determines these biological effects is the type of the induced damage and its location along the genome. However, attempts to elucidate the chemical basis of these biological effects are complicated by the multiplicity of products which are induced in the DNA subsequent to its exposure to radiation or chemical agents (Kittler & L o k r , 1977;. Under these circumstances, an assignment of a given chemical change to the consequent biological effect is made impossible.
Photochemistry and Photobiology, 1978
Superoxide radicals, singlet oxygen and hydroxyl radicals are individually or in combination involved in radiation or photochemical processes and in various enzymatic reactions. The reactivity and the mechanism of reaction of these oxygen species with some biologically significant DNA components were investigated through the characterization of the final oxidation products. Superoxide radicals appear to be unreactive with purine and pyrimidine 2'-deoxyribonucleosides. However, the autoxidation reaction of 6-hydroxydopamine leads to extensive degradation of thymine through the intermediary of hydroxyl radicals. Chemically and microwave-discharge generated singlet oxygen oxidation is specific to 2'-deoxyguanosine. The main oxidized products of these reactions were also characterized as well as an as yet unidentified nucleoside in the methylene-blue photooxydation of 2'-deoxyguanosine. These results, in addition to specific deuterium effect experiments. lend support to the involvment of singlet oxygen (type I1 mechanism) in the methylene-blue photosentization. No singlet oxygen effect was observed in aqueous irradiated system.
Photochemistry and Photobiology, 1997
The acridine and phenanthridine hydroperoxides 3 and 7 were synthesized as photochemical hydroxyl radical sources for oxidative DNA damage studies. The generation of hydroxyl radicals upon UVA irradiation (A = 350 nm) was verified by trapping experiments with 5,5-dimethyl-1-pyrroline N-oxide and benzene. The enzymatic assays of the damage in cell-free DNA from bacteriophage PM2 caused by the acridine and phenanthridine hydroperoxides 3 and 7 under near-UVA irradiation revealed a wide range of DNA modifications. Particularly, extensive single-strand break formation and DNA base modifications sensitive to formamidopyrimidine DNA glycosylase (Fpg protein) were observed. In the photooxidation of calf thymus DNA, up to 0.69 * 0.03% 8-0x0-7,8dihydroguanine was formed by the hydroperoxides 3 and 7 on irradiation, whose yield was reduced up to 40% in the presence of the hydroxyl radical scavengers mannitol and tert-butanol. The acridine and phenanthridine hydroperoxides 3 and 7 also induce DNA damage through the type I photooxidation process, for which photoinduced electron transfer from 2'-deoxyguanosine to the singlet states of 3 and 7 was estimated by the Rehm-Weller equation to possess a negative Gibb's free energy of ca-5 kcaY mol. Control experiments with the sensitizers acridine 1 and the acridine alcohol 4 in calf thymus and PM2 DNA confirmed the photosensitizing propensity of the UVA-absorbing chromophores. The present study emphasizes that for the development of selective and efficient photochemical hydroxyl radical sources, chromophores with low photosensitizing ability must be chosen to avoid type I and type I1 photooxidation processes.
Photochemistry and Photobiology, 2006
Products of riboflavin-mediated photosensitization of 2'deoxyguanosine (dG) and thymidylyl-(3'-5')-2'-deoxyguanosine (TpdG) by 350-nm light in oxygen-saturated aqueous solution have been isolated and identified as 1-(2-deoxy-p-~erythro-pentofuranosyl) oxaluric acid (P-dOx) and thymidylyl-(3'-5')-1-(2-deoxy-~-~-erythro-pentofuranosy1) oxaluric acid (Tpp-dOx), respectively. In aqueous solution the modified pdeoxyribonucleoside is slowly converted to the a-anomer, generating a-dOx and Tpa-dOx. These modified nucleosides and dinucleoside monophosphates have been isolated by HPLC and characterized by proton and carbon NMR spectroscopy, fast atom bombardment mass spectrometry, and enzymatic analyses. Both a-dOx and Tpa-dOx slowly convert back into the modified p-deoxyribonucleoside, indicating that the furanosidic anomers are in dynamic equilibrium. Relative to TpdG, the rate of hydrolysis of Tpp-dOx and Tpa-dOx by spleen phosphodiesterase is greatly reduced. Hot piperidine (1.0 M , 90°C, 30 min) destroys Tpp-dOx and Tpa-dOx. Riboflavin-mediated photosensitization of TpdG in D20 instead of H 2 0 has no detectable effect on the yield of Tpp-dOx, suggesting that oxaluric acid is generated through a Type-I reaction mechanism, likely through the intermediary on initially generated 8-oxo-7,8-dihydro-2'-deoxyguanosine.
Blue light induced free radicals from riboflavin on E. coli DNA damage
Journal of Photochemistry and Photobiology B: Biology, 2013
The micronutrients in many cellular processes, riboflavin (vitamin B 2), FMN, and FAD are photo-sensitive to UV and visible light to generate reactive oxygen species (ROS). The riboflavin photochemical treatment with UV light has been applied for the inactivation of microorganisms to serve as an effective and safe technology. Ultraviolet or high-intensity radiation is, however, considered as a highly risky practice. This study was working on the application of visible LED lights to riboflavin photochemical reactions to development an effective antimicrobial treatment. The photosensitization of bacterial genome with riboflavin was investigated in vitro and in vivo by light quality and irradiation dosage. The riboflavin photochemical treatment with blue LED light was proved to be able to inactivate E. coli by damaging nucleic acids with ROS generated. Riboflavin is capable of intercalating between the bases of bacterial DNA or RNA and absorbs lights in the visible regions. LED light illumination could be a more accessible and safe practice for riboflavin photochemical treatments to achieve hygienic requirements in vitro.
Oxyresveratrol protects against DNA damage induced by photosensitized riboflavin
Natural product communications, 2011
Riboflavin can be photosensitized to produce reactive oxygen species. In the present study, a DNA damage assay was developed based on the photo reaction of riboflavin. In this test system, oxyresveratrol showed higher DNA protective effect than the well-known antioxidants Trolox and ascorbic acid. The results suggest potential applications for oxyresveratrol as an anti-aging agent and a riboflavin stabilizer.
Combination Reactions of Superoxide with 8-Oxo-7,8-dihydroguanine Radicals in DNA
Journal of Biological Chemistry, 2004
One of the major biomarkers of oxidative stress and oxidative damage of cellular DNA is 8-oxo-7,8-dihydroguanine (8-oxoGua), which is more easily oxidized than guanine to diverse oxidative products. In this work, we have investigated further oxidative transformations of 8-oxoGua in single-and double-stranded oligonucleotides to the dehydroguanidinohydantoin, oxaluric acid, and diastereomeric spiroiminodihydantoin lesions. The relative distributions of these end products were explored by a combined kinetic laser spectroscopy and mass spectrometry approach and are shown to depend markedly on the presence of superoxide radical anions. The 8-oxaGua radicals were produced by oneelectron oxidation of 8-oxoGua by 2-aminopurine radicals generated by the two-photon ionization of 2-aminopurine residues site specifically positioned in 5d(CC[2-aminopurine]TC[8-oxoGua]CTACC). The hydrated electrons also formed in the photoionization process were trapped by dissolved molecular oxygen thus producing superoxide. A combination reaction between the 8-oxoGua and superoxide radicals occurs with the rate constant of (1.3 ؎ 0.2) ؋ 10 8 M ؊1 s ؊1 and (1.0 ؎ 0.5) ؋ 10 8 M ؊1 s ؊1 in single-and double-stranded DNA, respectively. The major end products of this reaction are the dehydroguanidinohydantoin lesions that slowly hydrolyze to oxaluric acid residues. In the presence of Cu,Znsuperoxide dismutase, an enzyme that induces the rapid catalytic dismutation of superoxide to the less reactive H 2 O 2 and O 2 , the yields of the dehydroguanidinohydantoin lesions become negligible. Under these conditions, the 8-oxoGua radicals do not exhibit any observable reactivities with oxygen (k < 10 2 M ؊1 s ؊1), decay on the time interval of several seconds, and the major reaction products are the spiroiminodihydantoin lesions. The possible biological implications of the 8-oxoGua oxidation are discussed.