Abolition of the thermotropic transition of charged phospholipids induced by a cardiotoxin from Naja mossambica mossambica as detected by fluorescence polarization, differential scanning calorimetry, and Raman spectroscopy

Abstract

The effects of N. mossambica mossambica cardiotoxin on the thermotropic properties of charged phospholipids were studied by fluorescence polarization, differential scanning calorimetry and Raman spectroscopy. The binding of the toxin was only governed by the net charge at the interface and was not affected by the polar head group structure of the phospholipids or by the acyl chain physical state, degree of unsaturation or length. The effect of the toxin on the phospholipid structure was drastic. In all cases, the gel to liquid-crystalline phase transition monitored by fluorescence and Raman spectroscopies was progressively abolished without notable shift in temperature as the proportion of toxin was increased. The endothermic peaks detected by differential scanning calorimetry decreased in intensity as the toxin content was increased but always remained sharp. All the techniques gave complementary results, and none of them revealed the presence of secondary transitions at higher or lower temperatures. The lipid molecules that were perturbed by the toxin, .apprx. 10 .+-. 2 molecules, did not undergo a phase transition. Raman results demonstrated that these boundary lipids displayed a population of gauche rotamers that was as high as the one found in the liquid-crystalline phase of the pure phospholipid and this was well below the phase transition temperature. Fluorescence results were interpreted as due to a partial immobilization of the lipids in contact with the toxin above the transition temperature. Even though the interaction was governed by electrostatic forces, the toxin penetrated at least partially into the bilayers, inducing a disorganization of the aliphatic chains and changes in their mobility; this could explain their lytic activity.