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Cholesterol and ergosterol influence nystatin surface aggregation: relation to pore formation.
Ana Coutinho1, Liana Silva, Alexander Fedorov
1Centro de Química-Física Molecular, Instituto Superior Técnico, P-1049-001 Lisbon, Portugal. pcacountinho@mail.ist.utl.pt
Biophysical Journal
|August 19, 2004
Summary
Nystatin exhibits a two-stage mechanism against fungal membranes, involving initial membrane perturbation followed by pore formation. Its interaction with cholesterol-rich membranes differs, suggesting a less defined interaction.
Area of Science:
- Biochemistry
- Pharmacology
- Membrane Biophysics
Background:
- Nystatin is an antifungal antibiotic.
- Its mechanism of action involves interaction with membrane sterols.
- Understanding nystatin's interaction with fungal versus mammalian membranes is crucial for its therapeutic application.
Purpose of the Study:
- To investigate the interaction of nystatin with liposomes mimicking fungal (ergosterol) and mammalian (cholesterol) membranes.
- To elucidate the mechanism of nystatin's antibiotic activity.
- To compare nystatin's behavior in ergosterol-containing versus cholesterol-containing lipid bilayers.
Main Methods:
- Fluorescence spectroscopy was used to study nystatin's interaction with large unilamellar vesicles (LUVs).
- Liposomes were composed of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) with varying concentrations of ergosterol or cholesterol.
- A pyranine fluorescence-detected K+/H+ exchange assay measured nystatin's activity and pore formation kinetics.
Main Results:
- Nystatin's fluorescence lifetime and spectral properties changed significantly with ergosterol content, indicating oligomerization and excitonic coupling.
- A sharp increase in nystatin mean fluorescence lifetime (5 to 37 ns) and pore formation rate was observed above a critical threshold (100 molecules per liposome).
- In cholesterol-containing liposomes, nystatin's spectroscopic properties and K+ efflux kinetics remained largely unchanged, suggesting a different interaction mechanism.
Conclusions:
- Nystatin acts via a two-stage mechanism in ergosterol-rich membranes: initial perturbation by monomers, followed by transmembrane channel formation upon oligomerization.
- The critical threshold for pore formation is dependent on ergosterol concentration.
- Nystatin's interaction with cholesterol-rich membranes is distinct, possibly involving looser assemblies or predominantly the initial perturbation stage.