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Interaction of the polyene antibiotic amphotericin B with model membranes: differences between small and large
J Milhaud1, M A Hartmann, J Bolard
1Laboratoire de Physique et Chimie Biomoléculaire (CNRS UA198), Université Pierre-et-Marie-Curie, Paris, France.
Abstract:
The interaction of the polyene antibiotic amphotericin B (AmB) (Fig. 1) with large unilamellar vesicles (LUV) was monitored by circular dichroism (CD) and carboxyfluorescein (CF) release. LUV afford a far better model for biological membranes than small unilamellar vesicles (SUV) which have been used until now. With dimyristoyl phosphatidyl choline (DMPC) LUV (i.e., containing saturated acyl chains), a strong and not saturable binding for AmB/lipid ratios up to 0.5 was observed both above and below the phase transition temperature. Incorporation of cholesterol into the vesicles did not significantly change the interaction. With egg PC (EPC) LUV (i.e., containing unsaturated acyl chains), quite a different picture emerged: the binding reached saturation for AmB/lipid ratios of about 5 x 10(-3), a result not observed with EPC SUV. When sterols were introduced into membranes, the CD spectral features obtained in the presence of ergosterol were different from those obtained in the presence of cholesterol. Such a different behavior was not observed with SUV. We suggest that species whose CD spectrum was observed after 15 min in the presence of ergosterol-containing EPC LUV is the particular one which forms wide channels and induces a Ca2+ release. (H. Ramos, A. Attias, B.E. Cohen and J. Bolard, submitted for publication). The CF release from EPC LUV induced by AmB was very low, even at very high concentrations of the antibiotic (3 x 10(-4)M). In contrast, an important release of the fluorescent dye was observed with DMPC LUV at concentrations of approximately 10(-5)M.(ABSTRACT TRUNCATED AT 250 WORDS)
Insights
Large unilamellar vesicles (LUVs) provide a better model for studying amphotericin B (AmB) interactions. AmB binding differs significantly between saturated (DMPC) and unsaturated (EPC) lipid chains in LUVs, impacting membrane permeability.
Area of Science:
- Biochemistry
- Membrane Biophysics
- Pharmacology
Background:
- Amphotericin B (AmB) is a polyene antibiotic used to treat fungal infections.
- Previous studies on AmB-membrane interactions utilized small unilamellar vesicles (SUV), which are less representative of biological membranes than large unilamellar vesicles (LUV).
Purpose of the Study:
- To investigate the interaction of AmB with LUVs composed of different lipid compositions.
- To compare AmB binding and membrane permeabilization in LUVs versus SUVs.
- To elucidate the role of sterols in modulating AmB-membrane interactions.
Main Methods:
- Circular dichroism (CD) spectroscopy to monitor AmB-lipid interactions.
- Carboxyfluorescein (CF) release assay to assess membrane permeability.
- Utilized dimyristoyl phosphatidyl choline (DMPC) and egg phosphatidyl choline (EPC) LUVs.
Main Results:
- AmB exhibited strong, non-saturable binding to DMPC LUVs, independent of temperature.
- AmB binding to EPC LUVs showed saturation at lower AmB/lipid ratios compared to DMPC LUVs.
- Ergosterol incorporation into EPC LUVs altered CD spectral features, suggesting specific AmB-sterol complex formation and potential channel formation.
- Significant CF release was observed from DMPC LUVs at lower AmB concentrations than from EPC LUVs.
Conclusions:
- LUVs offer a more biologically relevant model for studying AmB interactions.
- The lipid acyl chain unsaturation significantly influences AmB binding and membrane disruption.
- Sterols, particularly ergosterol, play a crucial role in modulating AmB's interaction with membranes and its ability to form pores.