| Popis: |
Cell-penetrating peptides (CPPs) comprise a group of peptides that consist of less than about 40 amino acids and traverse the cell membrane. Most of CPPs are generally cationic because they are enriched in arginine and/or lysine residues. It has been thought that the first step in the biological membrane penetration of CPPs is driven by electrostatic interactions between positive charges in CPPs and negative charges of cell surface glycosaminoglycans (GAGs).In this study, we investigated the interaction of three types of cationic peptides, Rev (TRQARRNRRRRWRERQR), Kev (TKQAKKNKKKKWKEKQK) and R8 (RRRRRRRR) with heparin which was used as a GAG model. Isothermal titration calorimetry demonstrated that the favorable enthalpy in binding of the cationic peptides to heparin mainly arises from non-coulombic interaction. The heparin binding enthalpies of Rev, R8 and Kev were −14 kcal/mole, −10 kcal/mole and −0.5 kcal/mole respectively. Thus, it was indicated that binding of Rev and R8 to heparin was much more exothermic compared to that of Kev to heparin, and that GAGs have higher affinity to arginine-rich peptide than to lysine-rich peptide. 1H NMR spectroscopy showed that proton signals of the Trp residue in Rev, not Kev, disappeared upon binding to heparin, demonstrating that Rev specifically interacts with heparin. The heparin binding induced α-helix structure in Rev whereas not in R8 and Kev. In addition, we demonstrated that Rev penetrates into Chinese hamster ovary cells using a membrane penetration assay of fluorescein labeled-CPPs, while subtle or no penetration was observed in the case of R8 and Kev. These results suggest that the penetration of CPPs across the cellular membrane is regulated by specific interaction of arginine residues in the CPP with GAGs and subsequent α-helix formation of the peptides upon binding to the biological membrane surface. |
