[Investigation of the mechanism of cell membrane-active cyclic lipodepsipeptides compounds].
The cyclic lipodepsipeptides produced by Pseudomonas syringae pv. syringae possess fungicide properties. They inhibit much of the cell functions, presumably on the basis of their pore-forming activity. The syringomycin E, studied earlier by our research group, formed pores on human red blood cells (RBC), and caused also partially hemolysis. To reach selective toxicity, knowledge of the relationship between the structure and function of the CLPs is required. To fulfill this requirement, the pore forming properties of two other CLPs, the syringopeptin22A (SP22A) and the syringotoxin (ST), were investigated on RBCs. On the basis of our transport kinetic measurements on RBC, the SP22A and the ST formed pores built up by few monomers. ST pores inactivated at 37 degrees C, and at 20 degrees C, however, they remained stable at 8 degrees C, similarly to the SRE. Inactivation of the SRE and ST pores was faster at 37 degrees C than at 20 degrees C. SP22A pores did not inactivate even at 37 degrees C. On the basis of our results, we concluded that fluidity has a crucial role in the process and the dimension of the hydrophobic part and the number of the positive charges of the polar headgroup influence the stability of the pores. To study the interactions of CLPs with membranes on molecular level we applied EPR spectroscopy and spectral simulation methods for liposomes of different composition. CLPs decreased the fluidity at all depths of the membrane: the motional and rotational freedom of the lipid molecules decreased, while their ordering increased. These observations in conjunction with the concentration dependence of the CLP's action suggested that CLPs form pores involving the lipids, too. Investigation of the temperature dependence of the CLPs' action showed that to get their complete effect a given temperature range is required. It corroborated that the fluidity has a crucial role in the interactions between CLPs and membrane lipids. From the results, obtained with DPPC-DOPC or DPPC-cholesterol liposomes we concluded that the membrane-inhomogeneity decreases the effectiveness of the CLPs. Dynamic light scattering measurements were used to determine the size-distribution of the liposomes. According to our experiences CLPs provoke fusion and/or aggregation, which can have a role in the cell-killing properties of the CLPs.
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Semmelweis Egyetem, Biofizikai és Sugárbiológiai Intézet, Budapest, Puskin u. 9.-1088.
D002462:Cell Membrane / Q000737:chemistry* / Q000648:ultrastructure; D008070:Lipopolysaccharides / Q000737:chemistry*; D008563:Membrane Lipids / Q000032:analysis*; D008958:Models, Molecular
D008070:Lipopolysaccharides; D008563:Membrane Lipids; C023023:lipid-linked oligosaccharides