Aggregation and organization of pardaxin in phospholipid membranes. A fluorescence energy transfer study.

Fluorescence resonance energy transfer has been used to study the aggregation and organization of pardaxin and its analogues within lipid membranes. Peptide molecules labeled with 5- (and 6-) carboxyfluorescein at their N-terminal amino acid served as donors in these energy transfer measurements, whereas peptides labeled with 5- (and 6-) carboxytetramethylrhodamine at either their N- or C-terminal amino acid, served as acceptors. The membrane-permeating activity of the native molecule was maintained in the labeled peptides. Upon aggregation of the labeled peptides, fluorescence energy transfer was detected as a quenching of the donor fluorescence (520 nm), as well as an enhancement of the acceptor fluorescence (575 nm). Correlation exists between self-aggregation of the different analogues within membranes and their poreforming abilities. A comparison of the degrees of fluorescence energy transfer from N1-donor-labeled pardaxin to N1-acceptor-labeled pardaxin with the transfer efficiency observed in the interaction between the same donor and C1-acceptor-labeled pardaxin suggests that aggregates are formed in an ordered manner, with a preferentially parallel orientation of monomers within the aggregate. The extent of hetero-oligomer formation, i.e. complexes composed of two different analogue species, revealed that complementary charges contribute to peptide-peptide recognition within the lipid bilayer. Taken together, these results provide further support for the barrel stave model, involving parallel organization of monomers within the aggregate, as a description of the pore formation mechanism of pardaxin and its analogues.