LATERAL DIFFUSION AND CONDUCTANCE PROPERTIES OF A FLUORESCEIN-LABELEDALAMETHICIN IN PLANAR LIPID BILAYERS

In order to follow alamethicin diffusion within membranes under condit ions of pore-formation, a fluorescein isothiocyanate (FITC) analogue w as synthesized. To test the influence of the fluorescent probe additio n on the pore-forming activity of the new analogue, macroscopic and si ngle-channel experiments into planar lipid bilayers were performed. Al though the apparent mean number of monomers per conducting aggregate w as equivalent, the voltage-dependence of the new analogue was slightly reduced and hysteresses were broader, in agreement with the much long er duration of the open single-channels. Thus, the conducting aggregat es seem to be stabilized by the introduction of the probe, presumably through the interaction of the conjugated cycles with the lipid headgr oups, while the added steric hindrance may account for the slightly hi gher conductances of the open substates. Lateral diffusion of the labe lled peptide associated with the bilayer was then investigated by the fluorescence recovery after photobleaching technique. Under applied vo ltage, associated with high conductance, D, the lateral diffusion coef ficient, was reduced by 50% when compared to peptide at rest. These re sults provide new independent experimental evidence for a voltage-driv en insertion of the highly mobile surface-associated peptide into the bilayer as a prominent step in pore formation. (C) 1997 Elsevier Scien ce B.V.