title: Energy-independent translocation of cell-penetrating peptides occurs without formation of pores. A biophysical study with pep-1.
abstract: Pep-1 is a cell-penetrating peptide (CPP) with the ability to translocate across biological membranes and introduce active proteins inside cells. The uptake mechanism used by this CPP is, as yet, unknown in detail. Previous results show that such a mechanism is endocytosis-independent and suggests that physical-chemical interactions between the peptide and lipid bilayers govern the translocation mechanism. Formation of a transmembrane pore has been proposed but this issue has always remained controversial. In this work the secondary structure of pep-1 in the absence/presence of lipidic bilayers was determined by CD and ATR-FTIR spectroscopies and the occurrence of pore formation was evaluated through electrophysiological measurements with planar lipid membranes and by confocal microscopy using giant unilamellar vesicles. Despite pep-1 hydrophobic domain tendency for amphipathic alpha-helix conformation in the presence of lipidic bilayers, there was no evidence for membrane pores in the presence of pep-1. Furthermore, alterations in membrane permeability only occurred for high peptide/lipid ratios, which induced the complete membrane disintegration. Such observations indicate that electrostatic interactions are of first importance in the pep-1-membrane interactions and show that pores are not formed. A peptide-lipid structure is probably formed during peptide partition, which favours peptide translocation.
authors.affiliation: Centro de Química e Bioquímica, Faculdade de Ciências da Universidade de Lisboa, Campo Grande, Lisboa, Portugal.
authors.forename: Sónia Troeira
authors.initials: ST
authors.lastname: Henriques
authors.forename: Alexandre
authors.initials: A
authors.lastname: Quintas
authors.forename: Luis A
authors.initials: LA
authors.lastname: Bagatolli
authors.forename: Fabrice
authors.initials: F
authors.lastname: Homblé
authors.forename: Miguel A R B
authors.initials: MA
authors.lastname: Castanho
mesh_terms: D002463:Cell Membrane Permeability; D004705:Endocytosis; D004734:Energy Metabolism; D008051:Lipid Bilayers / Q000378:metabolism; D008081:Liposomes; D010455:Peptides / Q000737:chemistry / Q000493:pharmacokinetics*; D016062:Porosity; D017433:Protein Structure, Secondary; D055672:Static Electricity
chemical_list: D008051:Lipid Bilayers; D008081:Liposomes; D010455:Peptides