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| 2. |
- Castro, Victor M, et al.
(författare)
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Differences among human tumor cell lines in the expression of glutathione transferases and other glutathione-linked enzymes
- 1990
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Ingår i: Carcinogenesis. - : Oxford University Press. - 0143-3334 .- 1460-2180. ; 11:9, s. 1569-1576
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Tidskriftsartikel (refereegranskat)abstract
- A large number of human tumor cell lines of various origins have been investigated with respect to expression of glutathione-linked enzymes in the cytosol fraction. The amounts of the different enzymes were estimated by use of activity measurements and by silver staining or immunoblot analysis after electrophoresis of cytosol fractions purified by affinity chromatography on S-hexylglutathione Sepharose. Class Pi glutathione transferase was the most abundant enzyme in most tumor cells; the cell lines HepG2 and Raji were exceptions in not expressing significant amounts of this enzyme. HepG2 cells derive from hepatocytes, which normally do not express the class Pi enzyme, whereas Raji cells originate from B-lymphocytes, which normally do express a class Pi glutathione transferase. The highest level of the class Pi transferase, in terms of protein reacting with antibodies as well as enzyme activity, was noted in the colon carcinoma cell line LS174T. Hu549Pat cells, EBV-transformed B-lymphocytes, also expressed high levels of a protein reacting with antibodies specific for class Pi glutathione transferases, but did not display any significant activity with ethacrynic acid, a substrate characteristic for this class. Class Alpha and class Mu glutathione transferases, in cell lines expressing these isoenzymes, were present in significantly lower concentrations than the class Pi enzyme. Most of the tumor cells contained a class Alpha transferase composed of 27.5 kd subunits, which has the physicochemical and immunological properties of the most basic glutathione transferase found in human skin. In several cell lines, a protein was detected with an apparent subunit Mr value of 30 kd that was tentatively identified as an additional class Alpha glutathione transferase not previously described. In addition, other glutathione-linked enzyme activities, namely glutathione peroxidase, glutathione reductase and glyoxalase I, were assayed with specific substrates in the cytosolic fraction of the tumor cells; glyoxalase I could also be estimated semiquantitatively by silver staining of SDS-PAGE cells after affinity chromatography. Like the glutathione transferases, these enzymes displayed distinctly different levels of expression in the various cell lines. Thus, virtually every cell line was found to have a unique pattern of glutathione-linked enzymes, suggesting that the resistance phenotypes of the cells differ accordingly.
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| 3. |
- Kim, Seog K., et al.
(författare)
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ENHANCEMENT OF BINDING RATE OF RECA PROTEIN TO DNA BY CARCINOGENIC BENZO A PYRENE DERIVATIVES AND SELECTIVE CHANGE OF ADDUCT CONFORMATION
- 1993
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Ingår i: Carcinogenesis. - : Oxford University Press (OUP). - 0143-3334 .- 1460-2180. ; 14:2, s. 311-313
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Tidskriftsartikel (refereegranskat)abstract
- The association kinetics of RecA protein from Escherichia coli to DNA is strongly enhanced if even a minor fraction of DNA bases has been modified by a carcinogenic (+)-anti metabolite of benzo[a]pyrene (BPDE). The enhancement is much smaller with the less carcinogenic (-)-anti enantiomer of BPDE suggesting the possibility that the RecA protein binds selectively to the proto-oncogenic target. Most importantly, the binding of RecA to DNA modified with the latter enantiomer is found to give rise to a reorganization of this BPDE adduct from an intercalation site into a minor groove site. This indicates that the binding mechanism of RecA is via intercalation of some amino acid moiety, a discovery that could explain the approximately 50% contour length increase of the DNA within its fibrous complex with RecA.
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| 4. |
- Ponten, I., et al.
(författare)
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SPECTROSCOPIC STUDIES OF THE TRANS ADDUCTS DERIVED FROM (+)-ANTI-BENZO A PYRENE-7,8-DIHYDRODIOL-9,10-EPOXIDE AND (-)-ANTI-BENZO A PYRENE-7,8-DIHYDRODIOL-9,10-EPOXIDE AND THE OLIGONUCLEOTIDE 5'-D(CCTATAGATATCC)
- 1994
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Ingår i: Carcinogenesis. - : Oxford University Press (OUP). - 0143-3334 .- 1460-2180. ; 15:10, s. 2207-2213
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Tidskriftsartikel (refereegranskat)abstract
- The oligonucleotide 5'-d(CCTATAGATATCC) has been reacted with the (+)- or (-)-enantiomers of trans-7,8-dihydroxy-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene [(+)-and (-)-anti-BPDE respectively]. Consistent with previous studies employing single-stranded oligonucleotides, adduct formation of both anti-BPDE enantiomers preferentially involved trans-addition of the C10 position of the diol-epoxide to the exocyclic nitrogen of deoxyguanosine [in the following abbreviated as (+)-BPDE(t)-N-2-G and (-)-BPDE(t)-N-2-G adducts respectively]. The unmodified or (+)-BPDE(t)-N-2-G-modified oligonucleotide was allowed to form duplexes with the complementary sequence 5'-d(GGATATCTATAGG) or sequences in which C has been replaced with T, G or A and analysed with regard to thermal stability. The presence of a (+)-BPDE(t)-N-2-G adduct in oligonucleotide duplexes substantially decreased the value of the melting point relative to the corresponding unmodified duplex. In mismatched complexes containing the (+)-BPDE(t)-N-2-G adduct, a further decrease in thermal stability was observed. The presence of a (+)-BPDE(t)-N-2-G adduct did not seem to change the extent of hyperchromicity (approximate to 20%) upon melting. 5'-d(GGATATCTATAGG) or strands in which C was replaced with T,G or A were gradually added to (+)- or (-)-BPDE(t)-N-2-G-modified oligonucleotides and the fluorescence emission intensity was determined. In all cases with (+)-BPDE(t)-N-2-G, except when C was replaced with A in the complement, the fluorescence intensity steadily decreased and became constant at equal strand concentrations. When a strand containing A in place of C was gradually added to the (+)-BPDE(t)-N-2-G oligonucleotide, a marked increase in the fluorescence intensity was observed (>3-fold). In contrast, addition of strands containing A, T or G to the (-)-BPDE(t)-N-2-G-modified oligonucleotide increased tbe fluorescence intensity from 1.5- to >5-fold. Addition of the fully complementary sequence to the (-)-BPDE(t)-N-2-G-containing oligonucleotide resulted in reduced fluorescence, however less pronounced than with the (+)-BPDE(t)-N-2-G-modified analogue. Significant changes in spectral properties of the adducts were observed in the duplexes. The absorption and fluorescence excitation maxima of the single-stranded (+)-BPDE(t)-N-2-G-modified oligonucleotide were at 353 nm. Insertion of C or A opposite the adduct caused a significant shift of these maxima to shorter wavelengths (347-348 nm). Addition of acrylamide, a fluorescence quencher, reduced the fluorescence intensity in all cases, but to variable extents. The adducts not quenchable by acrylamide demonstrate spectral properties similar to those of the single-stranded (+)-BPDE(t)-N-2-G-modified oligonucleotide.
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