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Differential Effect of Bilayer Thickness on Sticholysin Activity
Juan Palacios-Ortega1,2, Sara García-Linares1,2, Esperanza Rivera-de-Torre1
1Departamento de Bioquímica y Biología Molecular I, Universidad Complutense , Madrid 28040, Spain.
Langmuir : the ACS Journal of Surfaces and Colloids
|September 22, 2017
Summary
Bilayer thickness significantly impacts actinoporin toxin activity, influencing pore formation and membrane binding. Optimal toxin function occurs at specific membrane thicknesses, especially with cholesterol present.
Area of Science:
- Biochemistry
- Membrane Biophysics
- Protein-Lipid Interactions
Background:
- Actinoporins sticholysin I and II (StnI and StnII) are pore-forming toxins.
- These toxins specifically interact with sphingomyelin (SM) in cell membranes.
- Membrane properties, like thickness, can modulate toxin activity.
Purpose of the Study:
- To investigate how bilayer thickness affects the activity of StnI and StnII.
- To determine the role of cholesterol in modulating the interaction between actinoporins and lipid bilayers.
- To understand the structural basis for the observed effects on toxin activity.
Main Methods:
- Utilized large unilamellar vesicles (LUVs) with varying phosphatidylcholine (PC) acyl chain lengths to control bilayer thickness.
- Incorporated sphingomyelin (SM) and cholesterol into LUVs to mimic biological membrane conditions.
- Measured calcein leakage from LUVs to assess toxin-induced pore formation.
- Employed isothermal titration calorimetry (ITC) to quantify the binding affinity of toxins to LUVs.
Main Results:
- Calcein leakage rates showed moderate dependence on bilayer thickness, with peaks at di-16:1 and di-18:1 PC bilayers.
- In the presence of cholesterol, maximum leakage rates were observed with thinner di-14:1 and di-16:1 PC bilayers.
- Bilayer affinity constant (Ka) for Stn toxins peaked with di-18:1 PC bilayers and was reduced with shorter or longer acyl chains.
- Cholesterol increased toxin binding affinity approximately 30-fold at the optimal di-18:1 PC bilayer thickness.
Conclusions:
- Bilayer thickness is a critical factor influencing both the functional activity and conformational aspects of actinoporin binding and pore formation.
- The optimal membrane thickness for Stn toxin pore formation aligns with the length of their N-terminal alpha-helix.
- Cholesterol significantly enhances the binding affinity of actinoporins to membranes of optimal thickness.