| 1. |
- Mazari, Aslam M. A., et al.
(författare)
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Identification of new inhibitors for human hematopoietic prostaglandin D-2 synthase among FDA-approved drugs and other compounds
- 2015
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Ingår i: Chemico-Biological Interactions. - : Elsevier BV. - 0009-2797 .- 1872-7786. ; 229, s. 91-99
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Tidskriftsartikel (refereegranskat)abstract
- Objective: Hematopoietic prostaglandin D-2 synthase (HPGDS) is a member of the Sigma class glutathione transferases (GSTs) catalyzing the isomerization of prostaglandin H-2 to prostaglandin D-2, a mediator of allergy and inflammation responses. Selective inhibitors of human HPGDS are expected to be of therapeutic importance in relieving symptoms related to allergy and asthma. Hence, a collection of diverse FDA-approved compounds was screened for potential novel applications as inhibitors of HPGDS. Methods: The catalytic activity of purified HPGDS was used for inhibition studies in vitro. Results: Our inhibition studies revealed 23 compounds as effective inhibitors of HPGDS with IC50 values in the low micromolar range. Erythrosine sodium, suramin, tannic acid and sanguinarine sulfate were characterized with IC50 values of 0.2, 0.3, 0.4, and 0.6 mu M, respectively. Kinetic inhibition analysis showed that erythrosine sodium is a nonlinear competitive inhibitor of HPGDS, while suramin, tannic acid and sanguinarine sulfate are linear competitive inhibitors. Conclusion: The results show that certain FDA-approved compounds may have pharmacological effects not previously realized that warrant further consideration in their clinical use.
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| 2. |
- Scian, Michele, et al.
(författare)
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Comparison of epsilon- and delta-class glutathione S-transferases : the crystal structures of the glutathione S-transferases DmGSTE6 and DmGSTE7 from Drosophila melanogaster
- 2015
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Ingår i: Acta Crystallographica Section D. - 0907-4449 .- 1399-0047. ; 71, s. 2089-2098
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Tidskriftsartikel (refereegranskat)abstract
- Cytosolic glutathione transferases (GSTs) comprise a large family of enzymes with canonical structures that diverge functionally and structurally among mammals, invertebrates and plants. Whereas mammalian GSTs have been characterized extensively with regard to their structure and function, invertebrate GSTs remain relatively unstudied. The invertebrate GSTs do, however, represent potentially important drug targets for infectious diseases and agricultural applications. In addition, it is essential to fully understand the structure and function of invertebrate GSTs, which play important roles in basic biological processes. Invertebrates harbor delta-and epsilon-class GSTs, which are not found in other organisms. Drosophila melanogaster GSTs (DmGSTs) are likely to contribute to detoxication or antioxidative stress during development, but they have not been fully characterized. Here, the structures of two epsilon-class GSTs from Drosophila, DmGSTE6 and DmGSTE7, are reported at 2.1 and 1.5 angstrom resolution, respectively, and are compared with other GSTs to identify structural features that might correlate with their biological functions. The structures of DmGSTE6 and DmGSTE7 are remarkably similar; the structures do not reveal obvious sources of the minor functional differences that have been observed. The main structural difference between the epsilon-and delta-class GSTs is the longer helix (A8) at the C-termini of the epsilon-class enzymes.
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| 3. |
- Tzafestas, Kyriakos, et al.
(författare)
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Expression of a Drosophila glutathione transferase in Arabidopsis confers the ability to detoxify the environmental pollutant, and explosive, 2,4,6-trinitrotoluene
- 2017
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Ingår i: New Phytologist. - : Wiley. - 0028-646X .- 1469-8137. ; 214:1, s. 294-303
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Tidskriftsartikel (refereegranskat)abstract
- The explosive 2,4,6-trinitrotoluene (TNT) is a significant, global environmental pollutant that is both toxic and recalcitrant to degradation. Given the sheer scale and inaccessible nature of contaminated areas, phytoremediation may be a viable clean-up approach. Here, we have characterized a Drosophila melanogaster glutathione transferase (DmGSTE6) which has activity towards TNT. Recombinantly expressed, purified DmGSTE6 produces predominantly 2-glutathionyl-4, 6-dinitrotoluene, and has a 2.5-fold higher Maximal Velocity (Vmax), and five-fold lower Michaelis Constant (Km) than previously characterized TNT-active Arabidopsis thaliana (Arabidopsis) GSTs. Expression of DmGSTE6 in Arabidopsis conferred enhanced resistance to TNT, and increased the ability to remove TNT from contaminated soil relative to wild-type plants. Arabidopsis lines overexpressing TNT-active GSTs AtGST-U24 and AtGST-U25 were compromised in biomass production when grown in the absence of TNT. This yield drag was not observed in the DmGSTE6-expressing Arabidopsis lines. We hypothesize that increased levels of endogenous TNT-active GSTs catalyse excessive glutathionylation of endogenous substrates, depleting glutathione pools, an activity that DmGST may lack. In conclusion, DmGSTE6 has activity towards TNT, producing a compound with potential for further biodegradation. Selecting or manipulating plants to confer DmGSTE6-like activity could contribute towards development of phytoremediation strategies to clean up TNT from polluted military sites.
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| 4. |
- Kjellander, Marcus, et al.
(författare)
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Glutathione transferases immobilized on nanoporous alumina : Flow system kinetics, screening, and stability
- 2014
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Ingår i: Analytical Biochemistry. - : Elsevier BV. - 0003-2697 .- 1096-0309. ; 446, s. 59-63
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Tidskriftsartikel (refereegranskat)abstract
- The previously uncharacterized Drosophila melanogaster Epsilon-class glutathione transferases E6 and E7 were immobilized on nanoporous alumina. The nanoporous anodized alumina membranes were derivatized with 3-aminopropyl-triethoxysilane, and the amino groups were activated with carbonyldiimidazole to allow coupling of the enzymes via c-amino groups. Kinetic analyses of the immobilized enzymes were carried out in a circulating flow system using CDNB (1-chloro-2,4-dinitrobenzene) as substrate, followed by specificity screening with alternative substrates. A good correlation was observed between the substrate screening data for immobilized enzyme and corresponding data for the enzyme in solution. A limited kinetic study was also carried out on immobilized human GST S1-1 (also known as hematopoietic prostaglandin D synthase). The stability of the immobilized enzymes was virtually identical to that of enzymes in solution, and no leakage of enzyme from the matrix could be observed.
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| 5. |
- Lindström, Helena, et al.
(författare)
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Inhibition characteristics of equine steroid isomerase EcaGST A3-3
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- Equine glutathione transferase A3-3 (EcaGST A3-3) belongs to the superfamily of detoxifying enzymes found in all organisms. However it is also the most efficient steroid double-bond isomerase known in mammals. In contrast to the rodents, Equus ferus caballus shares the steroidogenic pathway with Homo sapiens, which makes it a more suitable model for human steroidogenesis than the murine one. Inhibition of EcaGST A3-3 might help treat reproductive and neurodegenerative disorders. We screened an FDA-approved library of 1040 compounds for the ability as novel inhibitors of EcaGST A3-3. Our results revealed anthralin, sennoside A, tannic acid and ethacrynic acid as the most potent, submicromolar-range inhibitors of EcaGST A3-3 with the natural substrate Δ5-androstene-3,17-dione.
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| 6. |
- Lindström, Helena, et al.
(författare)
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Potent inhibitors of equine steroid isomerase EcaGST A3-3
- 2019
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Ingår i: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 14:3
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Tidskriftsartikel (refereegranskat)abstract
- Equine glutathione transferase A3-3 (EcaGST A3-3) belongs to the superfamily of detoxication enzymes found in all higher organisms. However, it is also the most efficient steroid double-bond isomerase known in mammals. Equus ferus caballus shares the steroidogenic pathway with Homo sapiens, which makes the horse a suitable animal model for investigations of human steroidogenesis. Inhibition of the enzyme has potential for treatment of steroid-hormone-dependent disorders. Screening of a library of FDA-approved drugs identified 16 out of 1040 compounds, which at 10 mu M concentration afforded at least 50% inhibition of EcaGST A3-3. The most potent inhibitors, anthralin, sennoside A, tannic acid, and ethacrynic acid, were characterized by IC50 values in the submicromolar range when assayed with the natural substrate Delta(5)-androstene-3,17-dione.
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| 7. |
- Mazari, Aslam M. A., et al.
(författare)
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Drosophila GSTs display outstanding catalytic efficiencies with the environmental pollutants 2,4,6-trinitrotoluene and 2,4-dinitrotoluene
- 2016
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Ingår i: Biochemistry and Biophysics Reports. - : Elsevier BV. - 2405-5808. ; 5, s. 141-145
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Tidskriftsartikel (refereegranskat)abstract
- The nitroaromatic explosive 2,4,6-trinitrotoluene (TNT) and the related 2,4-dinitrotoluene (DNT) aretoxic environmental pollutants. The biotransformation and detoxication of these persistent compoundsin higher organisms are of great significance from a health perspective as well as for the biotechnological challenge of bioremediation of contaminated soil. We demonstrate that different human glutathionetransferases (GSTs) and GSTs from the fruit fly Drosophila melanogaster are catalysts of the biotransformationof TNT and DNT. The human GSTs had significant but modest catalytic activities with both DNT and TNT. However, D. melanogaster GSTE6 and GSTE7 displayed outstanding high activities withboth substrates.
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| 8. |
- Mazari, Aslam M. A., et al.
(författare)
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Overexpression of Glutathione Transferase E7 in Drosophila Differentially Impacts Toxicity of Organic Isothiocyanates in Males and Females
- 2014
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Ingår i: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 9:10
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Tidskriftsartikel (refereegranskat)abstract
- Organic isothiocyanates (ITCs) are allelochemicals produced by plants in order to combat insects and other herbivores. The compounds are toxic electrophiles that can be inactivated and conjugated with intracellular glutathione in reactions catalyzed by glutathione transferases (GSTs). The Drosophila melanogaster GSTE7 was heterologously expressed in Escherichia coli and purified for functional studies. The enzyme showed high catalytic activity with various isothiocyanates including phenethyl isothiocyanate (PEITC) and allyl isothiocyanate (AITC), which in millimolar dietary concentrations conferred toxicity to adult D. melanogaster leading to death or a shortened life-span of the flies. In situ hybridization revealed a maternal contribution of GSTE7 transcripts to embryos, and strongest zygotic expression in the digestive tract. Transgenesis involving the GSTE7 gene controlled by an actin promoter produced viable flies expressing the GSTE7 transcript ubiquitously. Transgenic females show a significantly increased survival when subjected to the same PEITC treatment as the wild-type flies. By contrast, transgenic male flies show a significantly lower survival rate. Oviposition activity was enhanced in transgenic flies. The effect was significant in transgenic females reared in the absence of ITCs as well as in the presence of 0.15 mM PEITC or 1 mM AITC. Thus the GSTE7 transgene elicits responses to exposure to ITC allelochemicals which differentially affect life-span and fecundity of male and female flies.
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| 9. |
- Mazari, Aslam M.A. 1982-
(författare)
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Studies on Human and Drosophila melanogaster Glutathione Transferases of Biomedical and Biotechnological Interest
- 2016
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Glutathione transferases (GSTs, EC.2.5.1.18) are multifunctional enzymes that are universally distributed in all cellular life forms. They play important roles in metabolism and detoxication of endogenously produced toxic compounds and xenobiotics. GSTs have gained considerable interest over the years for biomedical and biotechnological applications due to their involvement in the conjugation of glutathione (GSH) to a vast array of chemical species. Additionally, the emergence of non-detoxifying functions of GSTs has further increased their biological significance. The present work encompasses four scientific studies aimed at investigating human as well as fruit fly Drosophila melanogaster GSTs.Paper I presents the immobilization of GSTs on nanoporous alumina membranes. Kinetic analyses with 1-chloro-2,4-dinitrobenzene followed by specificity screening with alternative substrates showed a good correlation between the data obtained from immobilized enzymes and the enzymes in solution. Furthermore, immobilization showed no adverse effects on the stability of the enzymes. Paper II presents inhibition studies of human hematopoietic prostaglandin D2 synthase (HPGDS), a promising therapeutic target for anti-allergic and anti-inflammatory drugs. Our screening results with an FDA-approved drug library revealed a number of effective inhibitors of HPGDS with IC50 values in the low micromolar range. Paper III concerns the toxicity of organic isothiocyanates (ITCs) that showed high catalytic activities with GSTE7 in vitro. The in vivo results showed that phenethyl isothiocyanate (PEITC) and allyl isothiocyanate in millimolar dietary concentrations conferred toxicity to the adult fruit flies leading to death or shortened life-span. The transgenic female flies overexpressing GSTE7 showed increased tolerance against PEITC toxicity compared to the wild-type. However, the effect was opposite in male flies overexpressing GSTE7 after one week exposure. Notably, the transgene enhanced the oviposition activity of flies with and without ITCs exposure. Paper IV highlights Drosophila GSTs as efficient catalysts of the environmental pollutant and explosive 2,4,6-trinitrotoluene and the related 2,4-dinitrotoluene degradation. This result suggests the potential of GST transgenes in plants for biotransformation and phytoremediation of these persistent environmental pollutants.
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| 10. |
- Xie, Xin, et al.
(författare)
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β-actin regulates a heterochromatin landscape essential for optimal induction of neuronal programs during direct reprograming
- 2018
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Ingår i: PLOS Genetics. - : Public Library of Science (PLoS). - 1553-7390 .- 1553-7404. ; 14:12
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Tidskriftsartikel (refereegranskat)abstract
- During neuronal development, beta-actin serves an important role in growth cone mediated axon guidance. Consistent with this notion, in vivo ablation of the beta-actin gene leads to abnormalities in the nervous system. However, whether beta-actin is involved in the regulation of neuronal gene programs is not known. In this study, we directly reprogramed beta-actin(+/+) WT, beta-actin(+/-) HET and beta-actin(-/-) KO mouse embryonic fibroblast (MEFs) into chemically induced neurons (CiNeurons). Using RNA-seq analysis, we profiled the transcriptome changes among the CiNeurons. We discovered that induction of neuronal gene programs was impaired in KO CiNeurons in comparison to WT ones, whereas HET CiNeurons showed an intermediate levels of induction. ChIP-seq analysis of heterochromatin markers demonstrated that the impaired expression of neuronal gene programs correlated with the elevated H3K9 and H3K27 methylation levels at gene loci in beta-actin deficient MEFs, which is linked to the loss of chromatin association of the BAF complex ATPase subunit Brg1. Together, our study shows that heterochromatin alteration in beta-actin null MEFs impedes the induction of neuronal gene programs during direct reprograming. These findings are in line with the notion that H3K9Me3-based heterochromatin forms a major epigenetic barrier during cell fate change.
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