Binding of antibiotic amphotericin B to lipid membranes: monomolecular layer technique and linear dichroism-FTIR studies

Abstract

Amphotericin B (AmB) is one of the main antibiotics applied in treatment of deep-seated mycotic infections. Tensiometric technique has been applied to monitor binding of AmB, from the water subphase, to the lipid monomolecular layers, formed with dipalmitoylphosphatidylcholine at the air-water interface. Time dependencies of surface pressure in the monolayers demonstrate strong enhancement of AmB binding to monolayers brought about by sterols present in the membranes. The monolayers have been deposited to a solid support and examined by means of FTIR spectroscopy. FTIR measurements show that majority of the AmB molecules which bind to the membranes are localized in the polar headgroup region. The results of the linear dichroism-FTIR measurements are consistent with the microscopic picture according to which the molecules of the membrane-bound AmB are distributed among two orientational fractions: one horizontal and one vertical with respect to the plane of the membrane (59% versus 41% respectively, in the case of the membrane formed with the pure lipid without sterols). The presence of cholesterol in the membranes (50 mol% with respect to lipid) slightly affects such a distribution (53% horizontal versus 47% vertical) but the presence of ergosterol has a pronounced effect in the increase in population of the fraction of horizontally bound AmB (85% horizontal vs. 15% vertical). The results of the measurements indicate that mode of action of the AmB consists in disruption of the polar headgroup region of biomembranes, brought about by the AmB molecules bound horizontally with respect to the plane of the membrane.

Molecular organization of antifungal antibiotic amphotericin B in lipid monolayers studied by means of Fluorescence Lifetime Imaging Microscopy

“…Such a protocol yields efficient binding of AmB to the lipid membranes as described in our previous work [13]. the systems studied is highly heterogeneous.…”

“…It has been also proposed that selectivity toward cells of fungi is based upon a difference of the radii of porous structures of AmB binding ergosterol and cholesterol [6,9]. On the other hand, very recent reports show that alternatively both the biological action of the drug as well as toxic side effects may be directly related to the effect of AmB on physical properties of the membranes [10][11][12][13]. The 1 H-NMR technique studies demonstrated that the polar headgroup region of the membranes, rather than the hydrophobic core, is a predominant site of binding of AmB from the water phase [12].…”

“…The 1 H-NMR technique studies demonstrated that the polar headgroup region of the membranes, rather than the hydrophobic core, is a predominant site of binding of AmB from the water phase [12]. Analysis of the effect of AmB on structural and dynamic properties of lipid membranes carried out with application of SANS (small angle neutron scattering), 1 H-NMR and FTIR techniques showed that AmB molecules, even bound to the lipid polar groups, influence motional freedom of acyl lipid chains [11][12][13]. This effect is particularly pronounced in the case of the lipid…”

“…one oriented parallel and the other one perpendicular with respect to the plane of the membrane [13][14][15]. It appeared that a laterally-oriented pool was particularly large in the membranes containing ergosterol and that such orientation of the drug had strongest effect on the membrane organization [13].…”

“…It appeared that a laterally-oriented pool was particularly large in the membranes containing ergosterol and that such orientation of the drug had strongest effect on the membrane organization [13]. According to our earlier studies, AmB is able to form hydrophilic pores which can disturb transmembrane ion transport [16], indeed, but the results of the most recent studies, outlined above, seem to suggest that molecular mode of action of AmB with respect to biomembranes is primarily based on interference with the structural and dynamic properties of the lipid bilayers.…”