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Biochemical Society Transactions Jun 1987
Topics: Animals; Cytosol; Digitonin; Liver; Liver Circulation; Perfusion; Potassium; Proteins; Rats; Sodium
PubMed: 3622915
DOI: 10.1042/bst0150361 -
Vision Research Oct 1993We present experiments that test the effects of agents commonly used in visual pigment investigations, namely hydroxylamine (NH2OH), digitonin and triton X-100, on the...
We present experiments that test the effects of agents commonly used in visual pigment investigations, namely hydroxylamine (NH2OH), digitonin and triton X-100, on the photoreceptor and photoreception of Euglena. Hydroxylamine reacts with free and opsin-bound retinal, in aqueous solution, to form stable oximes, whereas digitonin and triton X-100 are the most common extractants of rhodopsin. Since previous data indicate that the chromophore present in Euglena photoreceptor is retinal, we investigated the influence of these chemicals on this organelle. The effects of these agents were studied by means of phase contrast, fluorescence and transmission electron microscopy and photobehaviour experiments. Hydroxylamine inhibited the formation of the Euglena photoreceptor. Photoaccumulation experiments on hydroxylamine-treated cells showed that they are unable to perceive light. Digitonin solubilized the crystalline structure of the photoreceptor, whereas the triton effect was limited to the membranous structures of the cell, leaving the photoreceptor unimpaired.
Topics: Animals; Digitonin; Euglena gracilis; Hydroxylamine; Hydroxylamines; Microscopy, Electron, Scanning Transmission; Microscopy, Phase-Contrast; Octoxynol; Photoreceptor Cells, Invertebrate
PubMed: 8266645
DOI: 10.1016/0042-6989(93)90002-e -
Biochimica Et Biophysica Acta May 1980Digitonin exerts profound effects on chitosomes (microvesicular structures with chitin synthetase activity isolated from the fungus Mucor rouxii). At low concentrations,... (Comparative Study)
Comparative Study
Digitonin exerts profound effects on chitosomes (microvesicular structures with chitin synthetase activity isolated from the fungus Mucor rouxii). At low concentrations, it stimulates chitin synthetase (UDP-2-acetamido-2-deoxy-D-glucose: chitin 4-beta acetamidodeoxy-D-glucosyltransferase, EC 2.4.1.16) activity; at higher concentrations, it inhibits it. Digitonin also causes disintegration of the chitosome and the release of a homogeneous population of chitosome subunits with chitin synthetase activity. These chitosome subunits have a sedimentation coefficient of 16 S, compared to 105 S for whole chitosomes, as determined by centrifugation in sucrose density gradients, and measure 7--12 nm in diameter. After dissociation, chitin synthetase remains in a zymogenic state, and requires treatment with a protease for activation. No change in sedimentation coefficient of chitosome subunits was observed after proteolytic activation. The product synthesized by the chitosome subunits was characterized by X-ray diffractometry ad alpha-chitin and was by the criterion indistinfuishable from chitin made by preparations of undissociated chitosomes. However, in the electron microscope, the chitin microfibrils made from chitosome subunits were, in general, much shorter than those produced by undissociated chitosomes and often exhibited a needle-like appearance.
Topics: Centrifugation, Density Gradient; Chitin; Chitin Synthase; Digitonin; Glucosyltransferases; Microscopy, Electron; Molecular Conformation; Mucor
PubMed: 6446324
DOI: 10.1016/0304-4165(80)90094-x -
Methods in Molecular Biology (Clifton,... 2023The protozoan parasite, Trypanosoma brucei, offers a simple system to study the growth and duplication of the Golgi. Cell lines stably expressing a photoactivatable GFP...
The protozoan parasite, Trypanosoma brucei, offers a simple system to study the growth and duplication of the Golgi. Cell lines stably expressing a photoactivatable GFP attached to an endogenous Golgi protein are permeabilized using digitonin. Photoactivation followed by imaging can then be used to follow the formation of the new Golgi.
Topics: Animals; Trypanosoma brucei brucei; Golgi Apparatus; Digitonin; Parasites; Protozoan Proteins
PubMed: 36512209
DOI: 10.1007/978-1-0716-2639-9_5 -
Journal of Neuroscience Methods Dec 1993Digitonin is widely used for extracting active neurotransmitter receptors from membranes. However, its low critical micellar concentration has made its removal from...
Digitonin is widely used for extracting active neurotransmitter receptors from membranes. However, its low critical micellar concentration has made its removal from samples problematic. Here we report that digitonin can be efficiently removed (> 90%) from solution using Extracti-Gel D, a detergent-absorbing matrix. Active kappa 1 opioid receptors solubilized from brain survive Extracti-Gel D chromatography with a recovery of 50-55% and 25% dilution by added volume. The loss of receptor and the dilution, however, are compensated for to a large extent by the disinhibition of binding that results from the removal of digitonin. Extracti-Gel D chromatography had little or no effect on the apparent equilibrium dissociation constant for [3H]U-69,593 binding to the kappa 1 receptor. We conclude that Extracti-Gel D column chromatography is a simple, highly efficient and practical method for markedly reducing the concentration of digitonin in biological samples. Application of the procedure should allow characterization of digitonin-solubilized receptors with minimal complications from bound digitonin and extend the usefulness of digitonin to studies going beyond the initial stages of receptor purification.
Topics: Absorption; Analgesics; Animals; Benzeneacetamides; Brain Chemistry; Chromatography, Ion Exchange; Digitonin; Guinea Pigs; Kinetics; Membranes; Nerve Tissue Proteins; Pyrrolidines; Receptors, Opioid, kappa; Solvents
PubMed: 8152245
DOI: 10.1016/0165-0270(93)90041-o -
Biochimica Et Biophysica Acta Jul 1980Artificial membrane vesicles (liposomes) have been used to study the lytic mechanism of the bacterial toxin, streptolysin O, compared to that of the well-known plant...
Artificial membrane vesicles (liposomes) have been used to study the lytic mechanism of the bacterial toxin, streptolysin O, compared to that of the well-known plant glycoside, digitonin. Two types of vesicle were prepared: large unilamellar vesicles and multilamellar liposomes. The vesicles were prepared with varying molar ratios of egg lecithin and cholesterol and loaded with the water-soluble spin label, TEMPO-choline chloride. Lysis of the vesicles was registered as release of spin label and monitored by change in the electron spin resonance (ESR) spectrum. In this system digitonin was able to lyse both large unilamellar vesicles and multilamellar liposomes. The effectiveness of lysis increased by increasing the percentage of cholesterol, but even at 0% cholesterol a significant level of lysis was observed by addition of a large enough concentration of digitonin. In contrast, no lysis was detected from multilamellar liposomes after exposure to streptolysin O, even when they consisted of 50 mol% cholesterol. On the other hand, large unilamellar vesicles could be lysed by streptolysin O, provided the cholesterol content was greater than 33%. At 67 mol% cholesterol in the membranes, the degree of lysis was diminished compared to 50%, which appeared to be optimal. This is the first demonstration of liposome lysis by streptolysin O and demonstrates the cholesterol specificity which has previously been shown by inhibition studies.
Topics: Cholesterol; Digitonin; Dose-Response Relationship, Drug; Electron Spin Resonance Spectroscopy; Liposomes; Microscopy, Electron; Phosphatidylcholines; Spin Labels; Streptolysins
PubMed: 6249363
DOI: 10.1016/0005-2736(80)90414-9 -
The Journal of Eukaryotic Microbiology 2001Respiration, oxidative phosphorylation. and the corresponding changes in membrane potential (deltapsi) of Trypanosoma cruzi epimastigotes grown either in liver...
Respiration, oxidative phosphorylation. and the corresponding changes in membrane potential (deltapsi) of Trypanosoma cruzi epimastigotes grown either in liver infusion-tryptose (LIT) or brain heart infusion (BHI) culture medium were assayed in situ using digitonin to render their plasma membrane permeable to succinate, ADP, safranine O, and other small molecules. When the cells were permeabilized with 64 microM digitonin, a concentration previously used with epimastigotes, the ability of the cells grown in LIT medium to sustain oxidative phosphorylation was demonstrated by the detection of an oligomycin-sensitive decrease in mitochondrial membrane potential induced by ADP. In contrast, the cells grown in BHI medium were not able to sustain a stable membrane potential and did not respond to ADP addition. Analyses of oxygen consumption by these permeabilized cells indicated that the rate of basal respiration, which was similar in both cell types, was significantly decreased by 64 microM digitonin. Addition of ADP to the permeabilized cells grown in LIT medium promoted an oligomycin-sensitive transition from resting to phosphorylating respiration in contrast to the cells grown in BHI medium, whose respiration decreased steadily and did not respond either to ADP or CCCP. Titration of the cells grown in BHI medium with different digitonin concentrations indicated that their mitochondria have higher sensitivity to digitonin than those grown in LIT medium. Analysis of the sterol composition of epimastigotes grown in the two different media showed a higher percentage of cholesterol in total and mitochondrial extracts of epimastigotes grown in BHI medium as compared to those grown in LIT medium, suggesting the involvement of this sterol in their increased sensitivity to digitonin-permeabilization.
Topics: Animals; Cell Membrane Permeability; Cholesterol; Culture Media; Digitonin; Membrane Potentials; Mitochondria; Oxygen Consumption; Phosphorylation; Sterols; Trypanosoma cruzi
PubMed: 11596924
DOI: 10.1111/j.1550-7408.2001.tb00195.x -
Biochimica Et Biophysica Acta Aug 1957
Topics: Digitonin; Humans; Mitochondria; Phosphorylation
PubMed: 13471565
DOI: 10.1016/0006-3002(57)90469-9 -
The Biochemical Journal May 1958
Topics: Digitalis; Digitonin; Electron Transport Complex IV; Ion Transport; Mitochondria; Oxidation-Reduction; Oxidoreductases; Plants
PubMed: 13535585
DOI: 10.1042/bj0690067 -
Acta Biochimica Polonica 1996Preincubation of Ehrlich ascites tumour cells with millimolar concentrations of pantothenic acid, pantothenol or pantethine, but not with homopantothenic acid, at 22...
Preincubation of Ehrlich ascites tumour cells with millimolar concentrations of pantothenic acid, pantothenol or pantethine, but not with homopantothenic acid, at 22 degrees C or 32 degrees C, but not at 0 degrees C, makes the plasma membrane more resistant to the damaging effect of submillimolar concentrations of digitonin. It is proposed that this increased resistance is due to the increased rate of cholesterol biosynthesis. In fact, incorporation of [14C]acetate into cholesterol is by 45% increased in the cells preincubated with pantothenic acid; this probably reflects elevation of the content of CoA in such cells [Slyshenkov, V.S., Rakowska, M., Moiseenok, A.G. & Wojtczak, L. (1995) Free Radical Biol. Med. 19, 767-772].
Topics: Acetic Acid; Animals; Carcinoma, Ehrlich Tumor; Cell Membrane Permeability; Cholesterol; Digitonin; Female; Mice; Pantothenic Acid
PubMed: 8862188
DOI: No ID Found