CONFORMATIONAL AND ASSOCIATIVE BEHAVIORS OF THE 3RD HELIX OF ANTENNAPEDIA HOMEODOMAIN IN MEMBRANE-MIMETIC ENVIRONMENTS

The third helix of antennapedia homeodomain pAntp-(43-58) can transloc ate through cell membrane and has been used as an intracellular vehicl e for delivering peptides and oligonucleotides. The conformational and associative behaviour of two peptidic vectors pAntp-(43-58) and [Pro5 0]pAntp-(43-58) has been analyzed by different biophysical methods. pA ntp-(43-58) adopts an amphipathic helical structure in 30% (by vol.) h exafluoroisopropanol, in perfluoro-tert-butanol and in the presence of SDS micelles. CD spectra indicate that the conformation of [Pro50]pAn tp-(43-58) in contrast to pAntp-(43-58) is independent of the media us ed. H-1-NMR spectroscopy in SDS micelles or in perfluoro-tert-butanol allows detection of aggregated peptides probably in a ribbon 2, type c onformation. These conformations became the predominant structure when Gln50 was replaced by Pro50. Interproton-distance restraints derived from NOE measurements have been classified in two groups corresponding to two types of structures: alpha-helix and essentially extended stru ctures. Consecutive CH alpha(i)/CH alpha(i+1) NOEs are only compatible with aggregates. Simulated annealing calculation of dimeric structure agrees with phi and psi angles in the beta-sheet and gamma-turn regio ns. Fluorescence spectroscopy analysis has shown that the indole group s of both peptides penetrate into SDS micelles; both peptides also ind uce the formation of micelles at very low concentration of SDS (20 mu M) Similar interaction was observed with reverse-phase micelles made o f bis(2-ethyhexyl) sodium sulfosuccinate and small unilamellar vesicle s (SW) made of a mixture of phosphatidylcholine/phosphatidylserine. P- 31-NMR of vesicles (SUV and large unilamellar vesicles) indicated that the addition of pAntp analogues did not affect the size of phosphatid ylcholine/phosphatidylserine vesicles. The addition of pAntp analogues to lipidic dispersions modulates lipid polymorphism in different ways depending on the mixtures of acidic lipids.