Lipid phase separations induced by the association of cholera toxin to phospholipid membranes containing ganglioside GM1.
The interactions of cholera toxin and their isolated binding and active subunits with phospholipid bilayers containing the toxin receptor ganglioside GM1 have been studied by using high-sensitivity differential scanning calorimetry and steady-state and time-resolved fluorescence and phosphorescence spectroscopy. The results of this investigation indicate that cholera toxin associates with phospholipid bilayers containing ganglioside GM1, independent of the physical state of the membrane. In the absence of Ca2+, calorimetric scans of intact cholera toxin bound to dipalmitoylphosphatidylcholine (DPPC) large unilamellar vesicles containing ganglioside GM1 result in a broadening of the lipid phase transition peak and a slight decrease (less than 5%) in the transition enthalpy. In the presence of Ca2+ concentrations sufficient to cause ganglioside phase separation, the association of the intact toxin to the membrane results in a significant decrease of enthalpy change for the lipid transition, indicating that under these conditions the toxin molecule perturbs the hydrophobic core of the bilayer. Calorimetric scans using isolated binding subunits lacking the hydrophobic toxic subunit did not exhibit a decrease in the phospholipid transition enthalpy even in the presence of Ca2+, indicating that the binding subunits per se do not perturb the hydrophobic core of the bilayer. On the other hand, the hydrophobic A1 subunit by itself was able to reduce the phospholipid transition enthalpy when reconstituted into DPPC vesicles. These calorimetric observations were confirmed by fluorescence experiments using pyrene phospholipids.(ABSTRACT TRUNCATED AT 250 WORDS)
D001665:Binding Sites; D002118:Calcium; D002152:Calorimetry, Differential Scanning; D002772:Cholera Toxin; D005677:G(M1) Ganglioside; D005732:Gangliosides; D008081:Liposomes; D008968:Molecular Conformation; D011663:Pulmonary Surfactants; D011956:Receptors, Cell Surface; D011971:Receptors, Immunologic; D013050:Spectrometry, Fluorescence; D013816:Thermodynamics; D014364:Tryptophan
D005732:Gangliosides; D008081:Liposomes; D011663:Pulmonary Surfactants; D011956:Receptors, Cell Surface; D011971:Receptors, Immunologic; C023552:choleragen receptor; D005677:G(M1) Ganglioside; D014364:Tryptophan; D002772:Cholera Toxin; D002118:Calcium