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One-sided action of amphotericin B on cholesterol-containing membranes is determined by its self-association in the
1Laboratoire de Physique et Chimie Biomoléculaires (U.A. C.N.R.S. 198), Université Pierre et Marie Curie, Paris, France.
Abstract:
The inducement of K+ permeability through membranes by the polyene antibiotic amphotericin B (AmB) has been analyzed as a measure of the antibiotic activity. Dose-response curves have been obtained with cholesterol- and ergosterol-containing egg yolk phosphatidylcholine large unilamellar vesicles (LUVs), human erythrocytes, and Saccharomyces cerevisiae cells. Conductance changes induced by AmB in sterol-containing planar bilayer membranes have also been studied. AmB self-association in aqueous buffer was determined by circular dichroism (CD) as a function of the antibiotic concentration. Electronic absorption and CD spectra of AmB were recorded in the presence of LUVs. For given AmB concentrations, the extent of permeability inducement is dependent on the lipid concentration. On the other hand, for cholesterol-containing LUVs or erythrocytes, a critical AmB concentration had to be reached before any permeability is observed. Independent of lipid concentration, this concentration was directly related to antibiotic self-association in the aqueous buffer. The same observation was made for erythrocytes and nystatin. The AmB absorption and CD spectra were totally different for ergosterol- and cholesterol-containing LUVs. Formation of single channels by one-sided addition of AmB could be observed only in ergosterol-containing membranes. These data lead us to propose that the permeability pathways induced by amphotericin B or nystatin, in ergosterol- and in cholesterol-containing membranes, are of different natures. In the latter case the antibiotics are only active, by single-sided addition, in the self-associated form. These findings offer important clues for the design of less toxic derivatives of AmB: they should have a low degree of self-association in water.
Insights
Amphotericin B (AmB) antibiotic activity was measured by inducing potassium (K+) permeability in membranes. Different mechanisms of AmB action were observed in cholesterol- versus ergosterol-containing membranes, suggesting pathways for designing less toxic antibiotics.
Area of Science:
- Biochemistry
- Membrane Biophysics
- Pharmacology
Background:
- The polyene antibiotic amphotericin B (AmB) is known to induce potassium (K+) permeability in biological membranes.
- AmB's interaction with sterols, such as cholesterol and ergosterol, is crucial for its membrane activity.
- Understanding the mechanism of AmB-induced permeability is key to developing safer antifungal agents.
Purpose of the Study:
- To analyze the induction of K+ permeability by amphotericin B (AmB) as a measure of its antibiotic activity.
- To investigate the role of sterol type (cholesterol vs. ergosterol) in AmB-mediated membrane permeabilization.
- To explore the relationship between AmB self-association and its activity in different membrane systems.
Main Methods:
- Dose-response studies using large unilamellar vesicles (LUVs) with cholesterol or ergosterol, human erythrocytes, and Saccharomyces cerevisiae cells.
- Measurement of conductance changes in sterol-containing planar bilayer membranes induced by AmB.
- Circular dichroism (CD) and electronic absorption spectroscopy to study AmB self-association and spectral changes in the presence of LUVs.
Main Results:
- AmB-induced permeability was dependent on lipid concentration in LUVs.
- A critical AmB concentration, linked to self-association, was required for permeability in cholesterol-containing membranes (LUVs, erythrocytes).
- Distinct spectral differences and channel formation patterns were observed between ergosterol- and cholesterol-containing membranes, suggesting different permeability pathways.
Conclusions:
- Amphotericin B and nystatin induce permeability through different mechanisms in ergosterol- versus cholesterol-containing membranes.
- In cholesterol-containing membranes, AmB is active only in its self-associated form upon single-sided addition.
- Designing less toxic AmB derivatives requires minimizing their self-association tendency in aqueous environments.