Tryptophan within basic peptide sequences triggers glycosaminoglycan-dependent endocytosis.
Deciphering the structural requirements and mechanisms for internalization of cell-penetrating peptides (CPPs) is required to improve their delivery efficiency. Herein, a unique role of tryptophan (Trp) residues in the interaction and structuring of cationic CPP sequences with glycosaminoglycans (GAGs) has been characterized, in relation with cell internalization. Using isothermal titration calorimetry, circular dichroism, NMR, mass spectrometry, and phase-contrast microscopy, we compared the interaction of 7 basic CPPs with 5 classes of GAGs. We found that the affinity of CPPs for GAGs increases linearly with the number of Trp residues, from 30 nM for a penetratin analog with 1 Trp residue to 1.5 nM for a penetratin analog with 6 Trp residues for heparin (HI); peptides with Trp residues adopt a predominantly β-strand structure in complex with HI and form large, stable β-sheet aggregates with GAGs; and in the absence of any cytotoxicity effect, the quantity of peptide internalized into CHO cells increased 2 times with 1 Trp residue, 10 times with 2 Trp residues, and 20 times with 3 Trp residues, compared with +6 peptides with no Trp residues. Therefore, Trp residues represent molecular determinants in basic peptide sequences not only for direct membrane translocation but also for efficient endocytosis through GAGs.
Université Pierre et Marie Curie, Université Paris 6, Unité Mixte de Recherche 7203, Paris, France.
Chérine
C
Bechara
Manjula
M
Pallerla
Yefim
Y
Zaltsman
Fabienne
F
Burlina
Isabel D
ID
Alves
Olivier
O
Lequin
Sandrine
S
Sagan
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D001693:Biological Transport, Active; D016466:CHO Cells; D002462:Cell Membrane / Q000378:metabolism; D057846:Cell-Penetrating Peptides / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006224:Cricetinae; D003412:Cricetulus; D004705:Endocytosis / Q000502:physiology*; D006025:Glycosaminoglycans / Q000737:chemistry / Q000378:metabolism*; D008954:Models, Biological; D008969:Molecular Sequence Data; D015394:Molecular Structure; D054730:Protein Interaction Domains and Motifs; D055503:Protein Multimerization; D017433:Protein Structure, Secondary; D013816:Thermodynamics; D014364:Tryptophan / Q000737:chemistry
D057846:Cell-Penetrating Peptides; D006025:Glycosaminoglycans; D014364:Tryptophan