title: The role of tryptophans on the cellular uptake and membrane interaction of arginine-rich cell penetrating peptides.
abstract: Cell-penetrating peptides (CPP) are able to efficiently transport cargos across cell membranes without being cytotoxic to cells, thus present a great potential in drug delivery and diagnosis. While the role of cationic residues in CPPs has been well studied, that of Trp is still not clear. Herein 7 peptide analogs of RW9 (RRWWRRWRR, an efficient CPP) were synthesized in which Trp were systematically replaced by Phe residues. Quantification of cellular uptake reveals that substitution of Trp by Phe strongly reduces the internalization of all peptides despite the fact that they strongly accumulate in the cell membrane. Cellular internalization and biophysical studies show that not only the number of Trp residues but also their positioning in the helix and the size of the hydrophobic face they form are important for their internalization efficacy, the highest uptake occurring for the analog with 3 Trp residues. Using CD and ATR-FTIR spectroscopy we observe that all peptides became structured in contact with lipids, mainly in α-helix. Intrinsic tryptophan fluorescence studies indicate that all peptides partition in the membrane in about the same manner (Kp~10(5)) and that they are located just below the lipid headgroups (~10 Å) with slightly different insertion depths for the different analogs. Plasmon Waveguide Resonance studies reveal a direct correlation between the number of Trp residues and the reversibility of the interaction following membrane washing. Thus a more interfacial location of the CPP renders the interaction with the membrane more adjustable and transitory enhancing its internalization ability.
authors.affiliation: CBMN-Univ Bordeaux, UMR 5248, Allée Geoffroy St Hilaire, 33600 Pessac, France.
authors.forename: Marie-Lise
authors.initials: ML
authors.lastname: Jobin
authors.affiliation: CBMN-Univ Bordeaux, UMR 5248, Allée Geoffroy St Hilaire, 33600 Pessac, France.
authors.forename: Marine
authors.initials: M
authors.lastname: Blanchet
authors.affiliation: CBMN-Univ Bordeaux, UMR 5248, Allée Geoffroy St Hilaire, 33600 Pessac, France.
authors.forename: Sarah
authors.initials: S
authors.lastname: Henry
authors.affiliation: CBMN-Univ Bordeaux, UMR 5248, Allée Geoffroy St Hilaire, 33600 Pessac, France.
authors.forename: Stéphane
authors.initials: S
authors.lastname: Chaignepain
authors.affiliation: CBMN-Univ Bordeaux, UMR 5248, Allée Geoffroy St Hilaire, 33600 Pessac, France.
authors.forename: Claude
authors.initials: C
authors.lastname: Manigand
authors.affiliation: CBMN-Univ Bordeaux, UMR 5248, Allée Geoffroy St Hilaire, 33600 Pessac, France.
authors.forename: Sabine
authors.initials: S
authors.lastname: Castano
authors.affiliation: CBMN-Univ Bordeaux, UMR 5248, Allée Geoffroy St Hilaire, 33600 Pessac, France.
authors.forename: Sophie
authors.initials: S
authors.lastname: Lecomte
authors.affiliation: Sorbonne Universités - UPMC Univ Paris 06, École Normale Supérieure-PSL Research University, Département de Chimie, CNRS UMR 7203 LBM, 4 Place Jussieu, F-75005 Paris, France.
authors.forename: Fabienne
authors.initials: F
authors.lastname: Burlina
authors.affiliation: Sorbonne Universités - UPMC Univ Paris 06, École Normale Supérieure-PSL Research University, Département de Chimie, CNRS UMR 7203 LBM, 4 Place Jussieu, F-75005 Paris, France.
authors.forename: Sandrine
authors.initials: S
authors.lastname: Sagan
authors.affiliation: CBMN-Univ Bordeaux, UMR 5248, Allée Geoffroy St Hilaire, 33600 Pessac, France. Electronic address: i.alves@cbmn.u-bordeaux.fr.
authors.forename: Isabel D
authors.initials: ID
authors.lastname: Alves
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D016466:CHO Cells; D002462:Cell Membrane / Q000737:chemistry / Q000187:drug effects*; D002463:Cell Membrane Permeability / Q000187:drug effects*; D002470:Cell Survival / Q000187:drug effects; D057846:Cell-Penetrating Peptides / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology; D003412:Cricetulus; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D008969:Molecular Sequence Data; D010649:Phenylalanine / Q000737:chemistry; D010713:Phosphatidylcholines / Q000737:chemistry*; D010715:Phosphatidylglycerols / Q000737:chemistry*; D011485:Protein Binding; D017433:Protein Structure, Secondary; D021381:Protein Transport; D055672:Static Electricity; D013329:Structure-Activity Relationship; D014364:Tryptophan / Q000737:chemistry*
chemical_list: D057846:Cell-Penetrating Peptides; D010713:Phosphatidylcholines; D010715:Phosphatidylglycerols; D010649:Phenylalanine; C051388:1,2-dioleoyl-sn-glycero-3-phosphoglycerol; D014364:Tryptophan; C017251:1,2-oleoylphosphatidylcholine
keywords: Cell penetrating peptide; Lipid model systems; Peptide/lipid interaction