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| 2. |
- Sundberg, Kathrin, et al.
(författare)
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Differences in the catalytic efficiencies of allelic variants of glutathione transferase P1-1 towards carcinogenic diol epoxides of polycyclic aromatic hydrocarbons
- 1998
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Ingår i: Carcinogenesis. - : Oxford University Press (OUP). - 0143-3334 .- 1460-2180. ; 19:3, s. 433-436
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Tidskriftsartikel (refereegranskat)abstract
- Previous studies have identified allelic variants of the human glutathione transferase (GST) Pi gene and showed that the two different encoded proteins with isoleucine (GSTP1-1/I-105) or valine (GSTP1-1/V-105) at position 105, respectively, differ significantly in their catalytic activities with model substrates. Moreover, recent epidemiological studies have demonstrated that individuals differing in the expression of these allelic variants also differ in susceptibility to tumour formation in certain organs, including such in which polycyclic aromatic hydrocarbons (PAH) may be etiological factors. In the present study the catalytic efficiencies (kcat/Km) of these GSTP1-1 variants were determined with a number of stereoisomeric bay-region diol epoxides, known as the ultimate mutagenic and carcinogenic metabolites of PAH, including those from chrysene, benzo[a]pyrene and dibenz[a,h]anthracene. In addition, GSTP1-1 mutants in which amino residue 105 is alanine (GSTP1-1/A-105) or tryptophan (GSTP1-1/W-105) have been constructed and characterized. GSTP1-1/V-105 was found to be more active than GSTP1-1/I-105 in conjugation reactions with the bulky diol epoxides of PAH, being up to 3-fold as active towards the anti- and syn-diol epoxide enantiomers with R-absolute configuration at the benzylic oxiranyl carbon. Comparing the four enzyme variants, GSTP1-1/A-105 generally demonstrated the highest kcat/Km value and GSTP1-1/W-105 the lowest with the anti-diol epoxides. A close correlation was observed between the volume occupied by the amino acid residue at position 105 and the value of kcat/Km. With the syn-diol epoxides, such a correlation was observed with alanine, valine and isoleucine, whereas tryptophan was associated with increased kcat/Km values. The mutational replacement of isoleucine with alanine or tryptophan at position 105 did not alter the enantio selectivity of the GSTP1-1 variants compared with the naturally occurring allelic variants GSTP1-1/I-105 and GSTP1-1/V-105. Since the amino acid at position 105 forms part of the substrate binding site (H-site) the effect of increasing bulkiness is expected to cause restricted access of the diol epoxide and proper alignment of the two reactants for efficient glutathionylation. In conclusion, the present study indicates that individuals who are homozygous for the allele GSTP1* B (coding for GSTP1-1/V-105) display a higher susceptibility to malignancy because of other factors than a decreased catalytic efficiency of GSTP1-1/V-105 in the detoxication of carcinogenic diol epoxides of benzo[a]pyrene or structurally related PAH.
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| 4. |
- Dreij, Kristian, et al.
(författare)
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In silico Modeling of Intracellular Diffusion and Reaction of Benzo[a]pyrene Diol Epoxide
- 2012
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Rapport (övrigt vetenskapligt/konstnärligt)abstract
- Several studies has suggested that glutathione conjugation of polycyclicaromatic hydrocarbons (PAHs) catalyzed by glutathione transferases (GSTs)are important factors in protecting cells against toxicity and DNA damagederived from these compounds. To further characterize the intracellular dynamicsof PAH DEs and the role of GSTs in protection against DNA damage,we recently developed a PDE model using techniques for mathematicalhomogenization (Dreij K et al. PLoS One 6(8), 2011). In this study, wewanted to further develop our model by benchmarking against results fromfour V79 cell lines; control cells and cells overexpressing human GSTs A1-1, M1-1 and P1-1. We used an approach of global optimization of the parametersdescribing the diffusion and reaction of the ultimate carcinogenic PAHmetabolite benzo[a]pyrene diol epoxide to fit measured values from the fourV79 cell lines. Numerical results concerning the formation of glutathioneconjugates and hydrolysis were in good agreement with results from measurementsin V79 cell culture. Cellular results showed significant protectionby GST expression against formation of DNA adducts with more than 10-fold reduced levels compared to control cells. Results from the model usingglobally optimized parameters showed that the model cannot predict theprotective effects of GSTs. Extending the model to also include effects fromprotein interactions and GST localization showed the same discrepancy. Insummary, the results show that we have an incomplete understanding of theintracellular dynamics of the interaction between BPDE and GST that warrantsfurther investigation and development of the model.
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| 5. |
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| 6. |
- Jarvis, Ian W H, et al.
(författare)
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Interactions between polycyclic aromatic hydrocarbons in complex mixtures and implications for cancer risk assessment
- 2014
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Ingår i: Toxicology. - Stockholm : Karolinska Institutet, Institute of Environmental Medicine. - 0300-483X .- 1879-3185.
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Tidskriftsartikel (refereegranskat)abstract
- In this review we discuss the effects of exposure to complex PAH mixtures in vitro and in vivo on mechanisms related to carcinogenesis. Of particular concern regarding exposure to complex PAH mixtures is how interactions between different constituents can affect the carcinogenic response and how these might be included in risk assessment. Overall the findings suggest that the responses resulting from exposure to complex PAH mixtures is varied and complicated. More- and less-than additive effects on bioactivation and DNA damage formation have been observed depending on the various mixtures studied, and equally dependent on the different test systems that are used. Furthermore, the findings show that the commonly used biological end-point of DNA damage formation is insufficient for studying mixture effects. At present the assessment of the risk of exposure to complex PAH mixtures involves comparison to individual compounds using either a surrogate or a component-based potency approach. We discuss how future risk assessment strategies for complex PAH mixtures should be based around whole mixture assessment in order to account for interaction effects. Inherent to this is the need to incorporate different experimental approaches using robust and sensitive biological endpoints. Furthermore, the emphasis on future research should be placed on studying real life mixtures that better represent the complex PAH mixtures that humans are exposed to.
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| 7. |
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| 8. |
- Lagerqvist, Anne, et al.
(författare)
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Both replication bypass fidelity and repair efficiency influence the yield of mutations per target dose in intact mammalian cells induced by benzo(a)pyrene-diol-epoxide and dibenzo(a,l)-pyrene-diol-epoxide.
- 2008
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Ingår i: DNA Repair. - : Elsevier. - 1568-7864 .- 1568-7856. ; 7:8, s. 1202-1012
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Tidskriftsartikel (refereegranskat)abstract
- Mutations induced by polycyclic aromatic hydrocarbons (PAH) are expected to be produced when error-prone DNA replication occurs across unrepaired DNA lesions formed by reactive PAH metabolites such as diol epoxides. The mutagenicity of the two PAH-diol epoxides (+)-anti-7,8-dihydroxy-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (BPDE) and (±)-anti-11,12-dihydroxy-13,14-epoxy-11,12,13,14-tetrahydrodibenzo[a,l]pyrene (DBPDE) was compared in nucleotide excision repair (NER) proficient and deficient hamster cell lines. We applied the 32P-postlabelling assay to analyze adduct levels and the hprt gene mutation assay for monitoring mutations. It was found that the mutagenicity per target dose was 4 times higher for DBPDE compared to BPDE in NER proficient cells while in NER deficient cells, the mutagenicity per target dose was 1.4 times higher for BPDE. In order to investigate to what extent the mutagenicity of the different adducts in NER proficient cells was influenced by repair or replication bypass, we measured the overall NER incision rate, the rate of adduct removal, the rate of replication bypass and the frequency of induced recombination in the hprt gene. The results suggest that NER of BPDE lesions are 5 times more efficient than for DBPDE lesions, in NER proficient cells. However, DBPDE adducts block replication more efficiently and also induce 6 times more recombination events in the hprt gene than adducts of BPDE, suggesting that DBPDE adducts are, to a larger extent, bypassed by homologous recombination. The results obtained here indicate that the mutagenicity of PAH is influenced not only by NER, but also by replication bypass fidelity. This has been postulated earlier based on results using in vitro enzyme assays, but is now also being recognized in terms of forward mutations in intact mammalian cells.
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| 9. |
- Lagerqvist, Anne, et al.
(författare)
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DNA repair and replication influence the number of mutations per adduct of polycyclic aromatic hydrocarbons in mammalian cells
- 2011
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Ingår i: DNA Repair. - : Elsevier BV. - 1568-7864 .- 1568-7856. ; 10:8, s. 877-886
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Tidskriftsartikel (refereegranskat)abstract
- Polycyclic aromatic hydrocarbons (PAH) are an important class of environmental contaminants many of which require metabolic activation to DNA-reactive bay or fjord region diolepoxides (DE) in order to exert their mutagenic and carcinogenic effects. In this study, the mutagenicity of the bay region diolepoxides (+)-anti-7,8-dihydroxy-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (BPDE) and ()-anti-1,2-dihydroxy-3,4-epoxy-1,2,3,4-tetrahydrodibenzo[a,h]anthracene (DBADE) and the fjord region diolepoxides ()-anti-11,12-dihydroxy-13,14-epoxy-11,12,13,14-tetrahydrodibenzo[a,l]-pyrene (DBPDE) and (+/-)-anti-3,4-dihydroxy-1,2-epoxy-1,2,3,4-tetrahydrobenzo[c]-phenanthrene (BPhDE) was compared in nucleotide excision repair (NER) proficient and deficient hamster cell lines. The (32)P-postlabelling assay was applied to analyze DNA adduct levels and the Hprt gene mutation assay for monitoring mutations. Previously, we found that the mutagenicity per adduct was four times higher for DBPDE compared to BPDE in NER proficient cells. In these same cells, the mutagenicity of DBADE and BPhDE adducts was now found to be significantly lower compared to that of BPDE. In NER deficient cells the highest mutagenicity per adduct was found for BPDE and there was a tenfold and fivefold difference when comparing the BPDE data with the DBADE and BPhDE data, respectively. In order to investigate to what extent the mutagenicity of the different adducts in NER proficient cells was influenced by repair or replication bypass, we measured the overall NER incision rate, the rate of adduct removal, the rate of replication bypass and the frequency of induced recombination in the Hprt gene. Since NER turned out to be an important pathway for the yield of mutations, we further analyzed the role of transcription coupled NER versus global genome NER. However, our data demonstrate that neither of these pathways seems to be the sole factor determining the mutation frequency of the four PAH-DE and that the differences in the repair efficiency of these compounds could not be related to the presence of a bay or fjord region in the parent PAH.
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| 10. |
- Pradhan, P., et al.
(författare)
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Induced Circular Dichroism of Benzo(a)pyrene-7,8-dihydrodiol 9,10-Epoxide Stereoisomers Covalently Bound to Deoxyribooligonucleotides Used to Probe Equilibrium Distribution between Groove Binding and Intercalative Adduct Conformations
- 1998
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Ingår i: Biochemistry. - : American Chemical Society (ACS). - 1520-4995 .- 0006-2960. ; 37:13, s. 4664-4673
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Tidskriftsartikel (refereegranskat)abstract
- Binding conformations of single anti-BPDE-N-2-dG adducts in oligonucleotides of varying base composition have been studied by induced circular dichroism (ICD). The sign of the ICD around 350 nm of single-stranded oligonucleotide adducts and the sign of an exciton type of CD component at 260 nm in both single strand and duplex farms of adducts correlate with the absolute configuration of the cyclohexyl moiety of the adduct. Changes in magnitude and sign of the ICD around 350 nm were observed upon duplex formation. The results show that adducts displaying external (minor groove) binding characteristics are associated with a significant positive ICD. Conversely, adducts displaying intercalation binding characteristics were found to have a positive or negative ICD. The magnitude of the ICD is dependent on the sequence context and the particular adduct isomer studied. Duplexes with (+)-trans-anti-BPDE-N-2-dG in 5'-d(CCTATCGCTATCC) or 5'-d(CCTATAGATATCC) exhibit a relatively strong positive ICD. In contrast, the duplexes with (+)-trans-anti-BPDE-N-2-dG in 5'-d(CCTATTGCTATCC) and 5'-d(CCTATTGTTATCC) display a small positive and negative ICD, respectively, in both cases suggesting conformational heterogeneity. Partially complementary duplexes (dA, dT, or do) localized opposite the (+)-trans-anti-BPDE-N-2-dG adduct in 5'-d(CCTATCGCTATCC) or 5'-d(CCTATAGATATCC) also demonstrated negative ICD. These results together with light absorption characteristics suggest a preferred conformation of intercalation for the mismatched duplexes. Evidence of an equilibrium between the external and intercalative adduct conformation is provided by the results from the temperature dependence of the near-UV absorption and ICD characteristics of (+)-trans-anti-BPDE-N-2-dG complex in a 5'-d(CCTATAGATATCC) duplex.
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