Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “SSB”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 865 records · Page 48Linked to original sources
PubMed1992

Inhibition of repair of bleomycin-induced DNA strand breaks by 2'-deoxycoformycin and its effect on antitumor activity in L5178Y lymphoblasts.

We have observed previously that treatment of plateau-phase L5178Y murine lymphoblasts in vitro with 2'-deoxycoformycin plus deoxyadenosine (dCF/dAdo) can inhibit the repair of X-irradiation-induced DNA single-strand breaks (SSB) in these cells and that this effect is associated with synergistic cell kill. In this study we examined the effect of a combination treatment of plateau-phase L5178Y cells with bleomycin (BLM) plus dCF/dAdo. Incubation of BLM-treated cells with dCF/dAdo resulted in significant inhibition of the repair of BLM-induced DNA SSB. However, an additive, but not a synergistic, increase in cell kill was observed when cells were treated with a combination of BLM plus dCF/dAdo.

Animals↗
PubMed1990

The role of hydroxyl radicals in tetrachlorohydroquinone induced DNA strand break formation in PM2 DNA and human fibroblasts.

Tetrachlorohydroquinone (TCHQ), which has previously been identified as a metabolite of pentachlorophenol, induces DNA strand breaks in isolated DNA and in human fibroblasts. Strand break formation in PM2 DNA is prevented by the addition of catalase and the hydroxyl radical scavengers DMSO, ethanol and mannitol, whereas addition of SOD reduced SSB only slightly. Oxygen radicals are formed by the autoxidation of TCHQ to the tetrachlorosemiquinone radical. Desferrioxamine (0.2 mM) completely abolished strand break formation, whereas the metal chelator DETAPAC (1 mM) reduced SSB by only 8.5%. The formation of the semiquinone radical at physiological conditions is shown by ESR spectroscopy. Exposure of human fibroblasts to TCHQ also leads to DNA single strand breaks measured by the alkaline elution assay. These were reduced by addition of 5% DMSO. This indicates that at least part of the strand break formation in human cells is also due to the action of hydroxyl radicals.

Cells, Cultured↗
PubMed1991

2-Chlorodeoxyadenosine inhibits the repair of DNA double-strand breaks and does not inhibit the repair of DNA single-strand breaks in X-irradiated Chinese hamster V79 cells.

The exposure of log-phase Chinese hamster V79 cells to 2-chlorodeoxyadenosine (CdA) for 3 h after X irradiation enhanced the lethal effects of X-rays in a concentration-dependent manner. The enhancement of the killing efficiency of X-rays by CdA was mainly observed in the reduction of quasi-threshold doses (Dq) of the dose-response curves. When the ability of CdA to inhibit the repair of X-ray-induced double- and single-strand breaks (dsb and ssb) of DNA was investigated by neutral- and alkaline-filter elution techniques, respectively, it was observed that 90% of dsb were rejoined in the absence of CdA within 30 min after X irradiation and 15-40% of dsb rejoining was suppressed by co-incubation of the cells with 5-10 microM of CdA for 3 h after X irradiation, whereas almost 100% of ssb were rejoined within 15 min regardless of the presence or absence of CdA. From these results it was concluded that CdA interfered exclusively with the repair of DNA dsb in X-irradiated Chinese hamster V79 cells and thereby increased the lethality of X-rays.

2-Chloroadenosine↗
PubMed1991

DNA strand break metabolism in human lymphocytes: a reevaluation.

Recent findings concerning the presumed existence of single-strand breaks (SSB) in quiescent human peripheral blood lymphocytes (PBL) are discussed in relation to the role of poly(ADP-ribosyl)ation in DNA strand break metabolism. It is argued that the activation of poly(ADP-ribose)polymerase (ADPRP) by a DNA-damaging agent is not indicative of an obligatory role of poly(ADP-ribosyl)ation in DNA repair. From this it follows that SSB induced by different strand-breaking agents might be removed by either ADPRP-dependent or ADPRP-independent DNA repair pathways.

DNA Damage↗
PubMed1990

Energy transfer in complexes of E. coli single-stranded DNA-binding protein with single-stranded poly-(2-thiouridylic acid).

The complexes of point-mutated Escherichia coli single-stranded DNA-binding protein (Eco SSB) with poly-(2-thiouridylic acid) (poly S2U) have been studied by optical detection of magnetic resonance spectroscopy (ODMR). Previous work has determined that two of four tryptophan (Trp) residues in Eco SSB undergo stacking interactions with nucleic acid bases. Selective photoexcitation of S2U bases was performed and subsequent triplet----triplet energy transfer from S2U to nearby Trp residues in the protein took place. The zero-field splitting (ZFS) parameters and sublevel kinetics were determined for each Trp residue sensitized by S2U. The sublevel lifetimes of the two sensitized residues are similar to those of normal Trp. The ZFS parameters, on the other hand, show a dramatic reduction relative to those of the uncomplexed protein, implying a more polarizable environment for the sensitized Trp residues and/or charge transfer interactions with the S2U bases.

DNA-Binding Proteins↗
PubMed1986

Conformation of the DNA undecamer 5'd(A-A-G-T-G-T-G-A-T-A-T) bound to the single-stranded DNA binding protein of Escherichia coli. A time-dependent transferred nuclear Overhauser enhancement study.

A time-dependent transferred nuclear Overhauser enhancement study of the conformation of the single-stranded DNA 11mer 5'd(A-A-G-T-G-T-G-A-T-A-T) bound to the single-stranded DNA binding protein of Escherichia coli (SSB) is presented. It is shown that the conformation of the bound 11mer is that of a right-handed B-type helix similar to that of the free 11mer. The observation of internucleotide transferred nuclear Overhauser enhancements for every base step excludes the possibility of intercalation by aromatic protein residues. In addition, it is shown that the effective correlation time of the bases (80 ns) corresponds to that of a complex of molecular weight approximately 170,000, containing two SSB tetramers. The sugars, on the other hand, exhibit a shorter effective correlation time (40 ns), indicating the presence of internal motion. This suggests that the bases are anchored to the protein surface, possibly by hydrophobic interactions, whereas the sugar-phosphate groups are directed outwards towards the solvent.

Bacterial Proteins↗
PubMed1989

Biochemical events essential to the recombination activity of Escherichia coli RecA protein. I. Properties of the mutant RecA142 protein.

We have characterized the biochemical properties of Escherichia coli RecA142 protein, the product of a recA allele that is phenotypically defective in genetic recombination. In vitro, this mutant RecA protein is totally defective in DNA heteroduplex formation. Despite this defect, RecA142 protein is not deficient in all other biochemical activities. RecA142 protein is proficient in single-strand (ss) DNA binding ability, ssDNA-dependent ATPase activity, and DNA-free self-association (although the first 2 properties show a greater sensitivity to NaCl concentration than does the wild-type protein). However, RecA142 protein is deficient in four properties: (1) its ssDNA-dependent ATPase activity is completely inhibited by ssDNA binding (SSB) protein, demonstrating that RecA142 protein is unable to compete effectively with SSB protein for ssDNA binding sites; (2) it is unable to promote the coaggregation of ssDNA and double-strand (ds) DNA; (3) its M13 dsDNA-dependent ATPase activity is attenuated to approximately 5% of the level of the wild-type protein; (4) it is unable fully to develop characteristics of the high-affinity ssDNA-binding state that is normally induced by ATP. The first three deficiencies correspond to defects in the presynaptic, synaptic and postsynaptic steps of the in vitro DNA strand exchange reaction, respectively; the fourth is the likely fundamental basis for defects 1 and 3. Therefore, one or more of these properties must be important to both the in vitro and in vivo processes.

Adenosine Triphosphatases↗
PubMed1989

Plasticity of DNA conformation around the Drosophila melanogaster alcohol dehydrogenase gene under torsional stress.

Genomic DNA of eukaryotes is thought to be organized into multiple topological domains whose conformation can be independently regulated by torsional stress. We have demonstrated the formation of altered DNA structures around the Drosophila melanogaster alcohol dehydrogenase (Adh) gene by sensitivity to endonucleases and by binding single-strand binding (SSB) protein. Several altered DNA structures were detected only on torsionally stressed DNA at specific sites. Some corresponded to the two initiation cap sites and the poly(A) addition sites and others were found in the 5'-flanking regions of both the adult and larval cap sites and in the 3'-flanking region of the Adh gene. In particular, the 5'-flanking regions both exhibited a plasticity of DNA conformation according to the strength of torsional stress and the concentration of Mg2+. SSB protein bound preferentially to the non-coding regions of the Adh gene only on torsionally stressed DNA and not on relaxed or linear DNA. The observed binding preference appeared to correspond to the thermodynamic stability of the base-pairs involved. These results suggest that DNA conformation is specifically organized around the Adh gene for gene function. The plasticity of DNA may play a role in the regulation of transcriptional activation.

Alcohol Dehydrogenase↗
PubMed1991

The effects on strand exchange of 5' versus 3' ends of single-stranded DNA in RecA nucleoprotein filaments.

Since the ends of DNA chains are thought to be important in homologous recombination, the way in which RecA protein and similar recombination enzymes process ends is important. We analyzed the effects of ends both on the formation of joints, and the progression of strand exchange. When the only homologous end was provided by a single strand, there was no significant difference between the formation of joints at a 5' end or a 3' end; but in agreement with the report of Konforti & Davis, Escherichia coli single-stranded DNA binding protein (SSB) selectively inhibited the activity of 5' ends. Complete strand exchange, assessed by study of linear single-stranded and double-stranded substrates, took place only in the 5' to 3' direction relative to DNA in the nucleoprotein filament. These observations pose a paradox: in the presence of SSB, of which there are about 800 tetramers per cell, the formation of homologous joints by RecA protein is favored at a 3' end, from which, however, authentic strand exchange appears not to occur. Since observations reported here and elsewhere show that joints have different properties when formed at a 5' versus a 3' end, we suggest that they may be processed differently in vivo.

DNA, Circular↗
PubMed1989

RecA protein inhibits in vitro replication of single-stranded DNA with DNA polymerase III holoenzyme of Escherichia coli.

Purified RecA protein from Escherichia coli inhibited 5-10-fold the rate of in vitro replication of both unirradiated and UV-irradiated single-stranded DNA (ssDNA) with DNA polymerase III holoenzyme. Maximal inhibition occurred at a ratio of 1 molecule of RecA per 2-4 nucleotides of DNA, and it affected primarily the initiation of elongation on primed ssDNA. Adding single-strand DNA-binding protein (SSB) caused a relief of inhibition. Under conditions when there was enough SSB to cover the ssDNA completely, RecA protein had no effect on initiation, elongation or dissociation steps of replication. These observations together with data from in vivo studies suggest a role for RecA protein in the arrest of DNA replication observed in cells exposed to UV-radiation and a variety of chemical carcinogens.

Autoradiography↗
PubMed1990

Effects of trypsin on X-ray-induced cell killing, chromosome abnormalities and kinetics of DNA repair in mammalian cells.

When cells are trypsinized before irradiation a potentiation of X-ray damage may occur. This is known as the 'trypsin effect'. Potentiation of X-ray damage on cell killing was seen in V79 Chinese hamster cells but was marginal in Chinese hamster ovary (CHO K1) cells and not evident in murine Ehrlich ascites tumour (EAT) cells. Trypsinization did however increase the number of X-ray-induced chromosomal abnormalities in all 3 lines. To investigate the possibility that trypsin acts by digestion of proteins in chromatin, further experiments were performed to monitor DNA damage and repair. Induction of DNA breaks by X-rays was unaffected by trypsin but trypsinized EAT (suspension) cells repaired single-strand breaks (ssb) less rapidly than controls indicating an inhibitory effect of trypsin on ssb repair. However double-strand break (dsb) repair was unaffected by trypsin. It was also found that the EDTA solution in which the trypsin was dissolved also contributes to the inhibition of dsb repair. The results show that trypsinization can enhance X-ray-induced cell killing, chromosomal damage and DNA repair, the effect varying between cell lines.

Animals↗
PubMed1993

Photobiological activities of 1,6-dioxapyrene in pro- and eukaryotic cells.

The photobiological effect of a new pyrene derivative, 1,6-dioxapyrene (1,6-DP), was studied in Salmonella typhimurium (strain TA100) and in the diploid strain D7 of the yeast Saccharomyces cerevisiae. In Salmonella, 1,6-DP shows little mutagenicity in the dark in comparison to benzo[a]pyrene (B[a]P). This mutagenic activity decreases in the presence of liver S9 homogenates from Aroclor induced XVIInc/Z mice. However, in combination with 365 nm (UVA) radiation and in the absence of S9 mix, 1,6-DP behaves as an effective photodynamic compound inducing lethal and mutagenic effects in both organisms. In yeast, its activity, like that of B[a]P, is highly dependent on the presence of oxygen. For the same incident dose of UVA, 1,6-DP is, however, at least 6 times more effective than B[a]P in inducing cytotoxic and mutagenic effects. At equitoxic doses, 1,6-DP is as photomutagenic as B[a]P, suggesting that in both cases mutagenicity is due to similar mechanisms. Spectrophotometric measurements indicate physical interaction of 1,6-DP with DNA in the dark. Laser flash photolysis experiments show that 1,6-DP generates singlet oxygen with a quantum yield of 0.17. In vitro 1,6-DP produces oxidative damage to guanine bases specific for singlet oxygen mediated reactions. Alkaline step elution analysis of 1,6-DP plus UVA treated yeast cells indicates a decrease in average molecular weights in DNA and an induction of single strand breaks (ssb) originating from alkali labile sites. This effect is enhanced by D2O and is thus likely to be due to the production of singlet oxygen. The strand breaks appear to differ from those induced by gamma-rays because little, if any, repair of these ssb occurs during 30 min of post-treatment incubation in complete growth medium. These results suggest that the photobiological effects of 1,6-DP are due to oxidative damage in DNA mostly induced by singlet oxygen.

Benzo(a)pyrene↗
PubMed1995

Induction of 8-hydroxy-2-deoxyguanosine and single-strand breaks in DNA of V79 cells by tetrachloro-p-hydroquinone.

The biocide pentachlorophenol (PCP) is in part metabolized, by microsomal enzymes, to tetrachloro-p-hydroquinone (TCHQ), which easily oxidizes to its semiquinone radical. Redox cycling of this compound produces reactive oxygen species (ROS) which ultimately may damage cellular DNA. Here, we report on DNA damage generated by TCHQ in hamster lung fibroblasts (V79 cells) using 8-hydroxy-2-deoxyguanosine (8-OH-dG) as a marker of a major oxidative genetic lesion and measuring the induction of DNA single-strand breaks (DNA SSB) with the aid of the alkaline elution assay. TCHQ was administered to cell cultures in concentrations of 6.25, 12.5, 25, and 50 microM for 1 h. 6.25 and 12.5 microM had no significant effect on both parameters, whereas the higher concentrations resulted in increases of the level of 8-OH-dG (2.3- and 2.0-fold, respectively) and induced DNA SSB. The latter lesion may arise from (i) direct attack of OH., (ii) repair of hydroxylated DNA bases, or (iii) cytotoxic effects. Metabolic transformation of PCP to TCHQ and/or other metabolites with quinoid structures and the subsequent generation of ROS, producing oxidative DNA damage, may play a role in PCP-induced carcinogenicity observed in mice.

8-Hydroxy-2'-Deoxyguanosine↗
PubMed1995

The biological activity of hydrogen peroxide. VI. Mechanism of the enhancing effects of L-histidine: the role of the formation of a histidine-peroxide adduct and membrane transport.

Further details of the mechanism of the enhancing effects of L-histidine (L-His) on the clastogenic activities of hydrogen peroxide (H2O2) were investigated. The L-His-H2O2 adduct was prepared and its physicochemical properties and biological activities were compared with those of a mixture of L-His plus H2O2 and of H2O2 alone. When the stabilities of the three test samples against glucose were determined in terms of residual H2O2 content in solutions of various pH values over the course of 11 days, the adduct was found to be more stable than H2O2 alone and very similar in terms of stability to the mixture. The almost equivalent stability of the adduct and the mixture suggested formation of the adduct in the mixture even though the interaction between L-His and H2O2 in solution seems, from 13C-NMR analysis, to be rather weak. In cell-free DNA after lysis of cell membranes, the induction of single-strand breaks (SSB) by the adduct and by the mixture was less effective than by H2O2 alone. These results contrast with previous results obtained in intact cells (Oya et al., 1992) and demonstrate the indispensability of the cell membrane for the enhancing effects of L-His. In the presence of inhibitors of the active transport of L-His, namely, 10 different neutral amino acids, effective suppression of the clastogenic activity of the adduct and of the mixture was observed, whereas four acidic and basic amino acids had no effect. Thus, the participation of active transport in the enhancing effects of L-His was apparent. The formation of the adduct of L-His with H2O2 brings about the stabilization or reduces the reactivity of H2O2 and, as a result, the induction of SSB is prevented to some extent in cell-free DNA systems. By contrast, in a cellular system, the accumulation of the adduct in cells by active transport is potentiated by the enhancing effect of L-His, although the mediation of some factors that can generate hydroxyl radicals (*OH) from the adduct in cells must be postulated.

Biological Transport↗
PubMed1987

Age-correlated DNA damage in human muscle tissue.

This investigation represents the largest study so far published on human DNA damage and aging. The subject of this investigation is damage, determined as DNA alterations which give rise to complete molecular breaks in the course of treatment of purified DNA solutions with single-strand-specific nucleases. The DNA is derived from milligram samples of human muscle of individuals mostly undergoing surgical treatment. Care has been taken to bring the muscle samples, once shut off from blood circulation to liquid nitrogen temperatures within few seconds. The DNA is prepared by a procedure keeping breaks by handling and by DNAase attack as low as possible, however pushing DNA purity, especially with respect to protein as high as possible. Highly purified DNA treated in this way has some sites which are susceptible to single-strand (ss) specific DNAase splitting (ss-events). Three different deoxyribonucleases have been used: Nuclease S1, Nuclease BAL31 and Pea Endo-Nuclease. They give very similar results, i.e. splitting of the DNA so as to yield DNA pieces of given distribution. The lengths of these double-strand (ds) pieces have been determined from their electron microscopical pictures, either by following the image contours with a magnetostrictive stylo of the projected photo on a pad, by following the contours with a mileage ruler, or by integrating the silver grains on the photo. The molecular weight averages of the ds DNA threads between two ss-events for each individual have been determined from 20 to 200 molecules. The 470 individuals contributing their data were from age groups from 1 to 91 years. The molecular weights show a considerable scatter with an average molecular weight of the DNA ds pieces between two ss-events of 43.93 MDa and a standard deviation of 17.99 MDa. Among the single-strand breaks (ssb) that split the DNA into such pieces is a fraction, the number of which increases in a highly significant fashion with the age of the donor. From this derives the fact that the average molecular weight of the DNA strand pieces between two ssb decreases with age. It is remarkable that the standard deviation of the molecular weights of such pieces increases with age significantly, too. On the basis of additional information mainly supplied by the DNA donor himself or by his parents the 470 members of the main group M where grouped according to their life-style, into: (1) abstinent people, essentially non-smokers and refraining from use of licit or illicit drugs, sub-group N.(ABSTRACT TRUNCATED AT 400 WORDS)

Adolescent↗
PubMed1992

The translation machinery and 70 kd heat shock protein cooperate in protein synthesis.

The function of the yeast SSB 70 kd heatshock proteins (hsp70s) was investigated by a variety of approaches. The SSB hsp70s (Ssb1/2p) are associated with translating ribosomes. This association is disrupted by puromycin, suggesting that Ssb1/2p may bind directly to the nascent polypeptide. Mutant ssb1 ssb2 strains grow slowly, contain a low number of translating ribosomes, and are hypersensitive to several inhibitors of protein synthesis. The slow growth phenotype of ssb1 ssb2 mutants is suppressed by increased copy number of a gene encoding a novel translation elongation factor 1 alpha (EF-1 alpha)-like protein. We suggest that cytosolic hsp70 aids in the passage of the nascent polypeptide chain through the ribosome in a manner analogous to the role played by organelle-localized hsp70 in the transport of proteins across membranes.

Amino Acid Sequence↗
PubMed1995

Cytogenetic biomonitoring of a population of children allegedly exposed to environmental pollutants. Phase 2: Results of a three-year longitudinal study.

Our previous cytogenetic biomonitoring of a group of inhabitants in a village (Mellery, Belgium) where exposure to a mixture of toxic environmental pollutants, (probably originating from a neighbouring chemical waste disposal site) was suspected, showed that difference in the SCE and HFC bioassays was more pronounced for children. The results of follow-up study in 1992 confirmed this surprising conclusion by an even higher incidence. As very few studies have been performed on the levels of children's biomarkers, this group of exposed populations needed to be explored further. Do children residing in the vicinity of hazardous waste sites indeed represent a population at higher risk? In the present study, we compare the performance of various bioassays (SCE, HFC, SSB and MN) in extended exposed and reference children's groups. Simultaneously, in the exposed group, we followed variation in the lymphocyte SCE frequencies as a function of time. Reversibility of the latter biomarker was ascertained subsequent to a preliminary technical remediation of the disposal site. We compared these data with those obtained from a synchronous cross-sectional study on a group of children living near a similar chemical disposal site. The two exposed populations did not differ from the reference population regarding to the SCE and HFC mean levels. Comparisons of the mean levels of the two other biomarkers, SSB and MN, showed no difference between the Mellery exposed children and the reference group from Wavre whereas significant differences appeared when the Hensies group is compared either to the Mellery or to the Wavre reference group.

Adolescent↗
PubMed1988

Overproduction of single-stranded DNA-binding protein increases UV-induced mutagenesis in Escherichia coli.

UV-induced mutagenesis was investigated in the uvrB strain and its isogenic counterpart overproducing the single-stranded DNA-binding protein (SSB). It was demonstrated that overproduction of SSB significantly increases the frequency of mutation. Our results indicate that such an increase might be due to certain abnormalities in induction of the SOS response (untimely and prolonged activation of the RecA protein).

DNA, Bacterial↗