INTRAMEMBRANE MOLECULAR DIPOLES AFFECT THE MEMBRANE INSERTION AND FOLDING OF A MODEL AMPHIPHILIC PEPTIDE

The relationship between the dipole potential and the interaction of t he mitochondrial amphipathic signal sequence known as p25 with model m embranes has been studied using yl)-4-[beta[2-(di-n-octyl-amino)-6-nap hthyl]vinyl] pyridinium betaine (di-8-ANEPPS) as a fluorescent probe. The dipole potential of phosphatidylcholine membranes was modified by incorporating into the bilayer the sterols phloretin and 6-ketocholest anol (KC), which decrease and increase the dipole potential, respectiv ely. The results derived from the application of a dual-wavelength rat iometric fluorescence method for following the variation of the membra ne dipole potential have shown that when p25 inserts into the lipidic bilayer, a decrease in the dipole potential takes place. The magnitude of this decrease depends on the initial value of the dipole potential , i.e., before interaction with the peptide. Thus, when KC was incorpo rated into the bilayer, the decrease caused by the membrane insertion of p25 was larger than that caused in PC membranes. Alternatively, in the presence of phloretin, the decrease in the potential caused by the peptide insertion was smaller. Complementary studies involving attenu ated total reflectance-fourier transform infrared spectroscopy of the peptide membrane interactions have shown that modification of the dipo le potential affects the conformation of the peptide during the course of its interaction with the membrane. The presence of KC induces a hi gher amount of helicoidal structure. The presence of phloretin, howeve r, does not appear to affect the secondary structure of the peptide. T he differences observed in the dipole potential decreases caused by th e presence of the peptide with the PC membranes and phloretin-PC membr anes, therefore, must involve differences in the tertiary and, perhaps , quaternary conformations of p25.