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Publications (10)33.56 Total impact

  • A Bründl, K Buff
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    ABSTRACT: A protein capable of specifically binding polychlorinated biphenyls (PCB) was partially purified from rat liver cytosol. After labeling with [3H]2,2',4,4',5,5'-hexachlorobiphenyl (6-CB), protein enrichment was guided by monitoring the protein-bound radioactivity through a sequence of purification steps including ion exchange chromatography and preparative gel electrophoresis. In addition, specific binding tests of individual fractions were carried out. An average 100-fold enrichment of the 40 kDa protein was achieved. A variety of ligands were tested in competitive binding studies with 6-CB. Whereas penta- and hexachloro-PCB congeners are high affinity competitors, the 3,3',4,4'-tetrachlorobiphenyl congener does not compete for 6-CB binding. Studies on the species and tissue distribution suggest a prevalence of the binding protein in tissues of the rat. Since the natural physiological ligand of the protein has not yet been identified, the function of the protein can only be speculated on.
    Biochemical Pharmacology 03/1993; 45(4):885-91. · 4.58 Impact Factor
  • K Buff, A Bründl
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    ABSTRACT: Specific binding of polychlorinated biphenyls (PCBs) to rat liver cytosol protein has been detected using the 3H-labeled PCB probe 2,2',4,4',5,5'-hexachlorobiphenyl (6-CB). Binding of 6-CB to cytosol protein is displaced by its non-radioactive congener, is of high affinity (Kd approximately 3 nM) and is saturable (maximal binding capacity Bmax approximately 600 pmol/mg protein). 6-CB binding is not found in liver cytosol of animals fed a PCB-supplemented diet (500 ppm PCB for 5 days). Binding is also in vitro inhibited by high concentrations of triglyceride. PCB congeners such as 3,3',4,4',5-pentachlorobiphenyl as well as the thyroid hormones 3,5,3',5'-tetraiodothyronine and 3,5,3'-triiodothyronine (the latter hormone with an order of magnitude lower affinity) compete for the PCB binding site. On the other hand, a number of biochemically important compounds including the PCB core compound biphenyl and the hormone ligands dexamethasone and estradiol, as well as 2,3,7,8-tetrachlorodibenzo-p-dioxin, do not compete for the 6-CB binding site. The data provide the first evidence of specific binding of unmetabolized PCB congeners to distinct binding sites in rat liver cytosol.
    Biochemical Pharmacology 04/1992; 43(5):965-70. · 4.58 Impact Factor
  • K Buff, M Wegenke, A Bründl
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    ABSTRACT: Association of the PCB congener 2,2',4,4',5,5'-hexachlorobiphenyl (6-CB) with cell nuclei has been studied in cultured monolayer human Chang liver cells. Photo-induced formation of covalent bonds determined 6-CB binding to protein of cell nuclei and to DNA. Nuclear binding of 6-CB approached equilibrium after approximately 30 min of incubation. Photo-induced binding in vitro to purified Chang liver cell DNA substantiated direct interaction of the PCB congener with DNA. In monolayer cells, low levels of photo-induced 6-CB DNA adducts could be detected using the very sensitive 32P-postlabeling method. Adduct formation was dependent on 6-CB concentration as well as on incubation time. Highest adduct levels were in the range of 2 X 10(-8). Model reactions in vitro showed photo-induced binding of 6-CB to individual purine deoxyribonucleotide-3'-phosphates. The results demonstrate rapid intracellular movement of the PCB congener into the cell nucleus. The vast majority is associated with nuclear protein, minute amounts of 6-CB are found proximate to the DNA helix as evidenced by photo-induced adducts of purine nucleotides.
    Biochemical Pharmacology 10/1989; 38(17):2773-9. · 4.58 Impact Factor
  • A Bründl, K Buff
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    ABSTRACT: The polychlorinated biphenyl congener 2,2',4,4',5,5'-hexachlorobiphenyl can be photoactivated by brief high-intensity ultraviolet irradiation. Photoactivated intermediates are bound to neighboring biological macromolecules. Properties and stability of hexachlorobiphenyl photobinding were examined with bovine serum albumin, a protein known to strongly bind lipophilic compounds. Photobinding to cultured human Chang liver cells was a function of ligand and cell protein concentration as well as of irradiation time. Binding increased with incubation time, in support of the time course of uptake previously measured in the same system by alternative methods. Separation of cell proteins by gel electrophoresis showed that the distribution pattern of photobinding changed at different rates for different proteins. Photobinding to major cell lipid groups and to individual phospholipids likewise reflected uptake of the compound. Notably, photobinding to phosphatidyl choline was elevated relative to phosphatidyl ethanolamine. Thus, the presented method is suitable to follow up transport and intracellular equilibrium distribution of photoactivatable ligands. As a particular advantage, artefactual redistribution of persistent lipophilic compounds during cell fractionation can be avoided.
    Biochemical Pharmacology 05/1988; 37(8):1601-8. · 4.58 Impact Factor
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    ABSTRACT: Interaction of the insecticide 1,1,1-trichloro-2,2-bis-(p-chlorophenyl)ethane (DDT) with β-receptor binding and adenylate cyclase activity of biological membranes has been studied. Following exposure of cultured Chang liver cells to DDT, maximal binding of the catecholamine antagonist [125I]iodohydroxybenzylpindolol (HYP) to isolated cell membranes was decreased by 30% whereas the dissociation constant remained unchanged. Both basal activity and maximal isoproterenol-stimulated activity of adenylate cyclase were not altered. The isoproterenol concentration required for half-maximal stimulation of the enzyme was increased about 2-fold as was the agonist concentration required for half-maximal displacement of the antagonist HYP at the β-receptor binding site. Thus, coupling efficiency of hormone-stimulated adenylate cyclase activity was not influenced by the presence of DDT in these membranes.The data show that interaction of DDT with the β-receptor adenylate cyclase complex is restricted to the receptor component. Enzyme activity is directly linked to changes of agonist binding at the β-receptor. Interference of DDT with signal transuction via ‘fluidization’ of membrane lipids has not been detected.
    Chemico-Biological Interactions 01/1984; · 2.97 Impact Factor
  • K. Buff, A. Bründl
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    ABSTRACT: A variety of membrane-specific parameters was examined in both intact cells and isolated plasma membranes following exposure of cultured human liver cells to the insecticide 1,1-(2,2,2-trichloroethylidene)bis(4-chloro)benzene (DDT). Uptake of DDT was at equilibrium within 6 hr. In contrast, a decrease in the number of β-adrenergic hormone receptors first became significant after 48 hr of cell exposure. Whereas the uptake was largely reversible, the loss in the number of β receptors did not recover after DDT-exposed cells were cultured in fresh medium lacking the insecticide. Experiments in vitro substantiated the time lag of the biological effect. The decrease in receptor proteins was persistent in membranes with increased phospholipid unsaturation. Temperature-activity profiles (“Arrhenius plots”) of and 5′-nucleotidase were unchanged. Endogenous tryptophan fluorescence of membrane proteins was lower in membranes from DDT-exposed cells. These selective alterations in membrane parameters suggest a specific interaction of DDT with membrane proteins; interference with cellular protein synthesis is possible. The results indicate that membrane lipid “fluidization” does not play a physiologically important role in the mechanism of DDT action in biomembranes.
    Pesticide Biochemistry and Physiology. 01/1984;
  • K Buff, A Bründl, J Berndt
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    ABSTRACT: The thermal dependence of the fluorescence polarization of 1,6-diphenylhexatriene was recorded upon interaction of 1,1,1-trichloro-2,2-bis(p-chlorophenyl)ethane (DDT) and some other pesticides with dipalmitoylphosphatidylcholine liposomes. Differential effects on the gel-crystalline phase were observed. Most substances decreased probe polarization; pentachlorophenol caused an increase of this parameter. The DDT-induced change of polarization was also dependent on the vesicle concentration thus indicating the influence of light scattering. The amount of DDT and pentachlorophenol residing in the lipid bilayer was determined to confirm the localization in the membrane. Correlation with the effects on probe polarization was obtained. The difference in response of the fluorescent probe to the presence of foreign molecules in the lipid bilayer may reflect different modes of interaction.
    Biochimica et Biophysica Acta 06/1982; 688(1):93-100. · 4.66 Impact Factor
  • E Pöschl, K Buff, J Berndt
    Chemico-Biological Interactions 05/1982; 39(3):369-74. · 2.97 Impact Factor
  • K Buff, J Berndt
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    ABSTRACT: The influence of 1,1,1-trichloro-2,2-bis(p-chlorophenyl)ethane (DDT) and several other pesticides on the physical state of membrane phospholipids was investigated using model lipids. The thermal dependence of fluorescence intensity of the probe parinaric acid in dipalmitoylphosphatidylcholine liposomes and lipid vesicles of mixed composition were recorded. DDT was incorporated into the liposomal bilayer. The insecticide lowered the phase transition temperature and broadened the temperature range of the transition. The effects were concentration-dependent. The results may be interpreted as a sort of blurred and facilitated phase transition of bilayer lipids caused by intercalation of DDT between fatty acyl chains of membrane phospholipids.
    Biochimica et Biophysica Acta 05/1981; 643(1):205-12. · 4.66 Impact Factor
  • K. Buff, J. Berndt
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    ABSTRACT: The influence of (DDT) and several other pesticides on the physical state of membrane phospholipids was investigated using model lipids. The thermal dependence of fluorescence intensity of the probe parinaric acid in dipalmitoylphosphatidylcholine liposomes and lipid vesicles of mixed composition were recorded. DDT was incorporated into the liposomal bilayer. The insecticide lowered the phase transition temperature and broadened the temperature range of the transition. The effects were concentration-dependent.The results may be interpreted as a sort of blurred and facilitated phase transition of bilayer lipids caused by intercalation of DDT between fatty acyl chains of membrane phospholipids.
    Biochimica et Biophysica Acta (BBA) - Biomembranes. 01/1981;