A new paramagnetic analogue of cholesterol as a tool for studying molecular interactions of genuine cholesterol

The synthesis of a new paramagnetic (nitroxide) analogue of cholesterol is described. This compound (called CNO) contains a doxyl group in the lateral chain at position 25. Our results show that CNO retains three molecular interactions which characterize authentic cholesterol: It assumes an orientation perpendicular to the phospholipid bilayer with the doxyl group buried in the membrane core, as seen by ESR spectroscopy. It widens the transition temperature of dimyristoylphosphatidylcholine, to the same extent as cholesterol, as measured by Raman and ESR spectroscopies. It interacts with polyene antibiotics, such as amphotericin B and filipin, in the same manner as its model. This was proved on the one hand by the change in fluorescence of self quenched vesicle-entrapped calcein, after dilution in the external medium, provoked by filipin, and on the other hand by fluorescence quenching provoked by cobalt ions entering the vesicles under the influence of amphotericin B. We concluded that CNO, although it has a side chain different from genuine cholesterol, can help to solve many physiologically meaningful questions related to the distribution and movement rate of cholesterol itself.