Abstract
Due to the poor permeability of the plasma membrane, several strategies are designed to enhance the transfer of therapeutics into cells. Over the last 20 years, small peptides called Cell-Penetrating Peptides (CPPs) have been widely developed to improve the cellular delivery of biomolecules. These small peptides derive from protein transduction domains, chimerical constructs, or model sequences. Several CPPs are primary or secondary amphipathic peptides, depending on whether the distribution of their hydrophobic and hydrophilic domains occurs from their amino-acid sequence or through α-helical folding. Most of the CPPs are able to deliver different therapeutics such as nucleic acids or proteins in vitro and in vivo. Although their mechanisms of internalization are varied and controversial, the understanding of the intrinsic features of CPPs is essential for future developments. This chapter describes several protocols for the investigation of biophysical properties of amphipathic CPPs. Surface physics approaches are specifically applied to characterize the interactions of amphipathic peptides with model membranes. Circular dichroism and infra-red spectroscopy allow the identification of their structural state. These methods are exemplified by the analyses of the main biophysical features of the cell-penetrating peptides MPG, Pep-1, and CADY.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Dietz, G.P. and Bahr, M. (2004) Delivery of bioactive molecules into the cell: the Trojan horse approach. Mol. Cell. Neurosci. 27, 85–131.
Fischer, R., Fotin-Mleczek, M., Hufnagel, H. and Brock, R. (2005) Break on through to the other side-biophysics and cell biology shed light on cell-penetrating peptides. Chem. Biochem. 6, 2126–2142.
Heitz, F., Morris, M.C. and Divita, G. (2009) Twenty years of cell-penetrating peptides: from molecular mechanisms to therapeutics. Br. J. Pharmacol. 157, 195–206.
Langel, Ü. (2006) Preface, Cell-Penetrating Peptides, 2nd edition. (Ed.: Ü. Langel), CRC Press, Boca Raton.
Morris, M.C., Deshayes, S., Heitz, F. and Divita, G. (2008) Cell-penetrating peptides: from molecular mechanisms to therapeutics. Biol. Cell. 100, 201–217.
Mano, M., Teodósio, C., Paiva, A., Simões, S. and Pedroso de Lima, M.C. (2005) On the mechanisms of the internalization of S4(13)-PV cell-penetrating peptide. Biochem. J. 390, 603–612.
Duchardt, F., Fotin-Mleczek, M., Schwarz, H., Fischer, R. and Brock, R. (2007) A comprehensive model for the cellular uptake of cationic cell-penetrating peptides. Traffic 8, 848–866.
Deshayes, S., Morris, M.C., Divita, G. and Heitz, F. (2005) Cell-penetrating peptides: tools for intracellular delivery of therapeutics. Cell. Mol. Life Sci. 62, 1839–1849.
Crombez, L., Morris, M.C., Deshayes, S., Heitz, F. and Divita, G. (2008) Peptide-based nanoparticle for ex vivo and in vivo drug delivery. Curr. Pharm. Des. 14, 3656–3665.
Deshayes, S., Morris, M.C., Heitz, F. and Divita, G. (2008) Delivery of proteins and nucleic acids using a non-covalent peptide-based strategy. Adv. Drug Deliv. Rev. 60, 537–547.
Morris, M.C., Vidal, P., Chaloin, L., Heitz, F. and Divita, G. (1997) A new peptide vector for efficient delivery of oligonucleotides into mammalian cells. Nucleic Acids Res. 25, 2730–2736.
Morris, M.C., Chaloin, L., Méry, J., Heitz, F. and Divita, G. (1999) A novel potent strategy for gene delivery using a single peptide vector as a carrier. Nucleic Acids Res. 27, 3510–3517.
Simeoni, F., Morris, M.C., Heitz, F. and Divita, G. (2003) Insight into the mechanism of the peptide-based gene delivery system MPG: implications for delivery of siRNA into mammalian cells. Nucleic Acids Res. 31, 2717–2724.
Deshayes, S., Gerbal-Chaloin, S., Morris, M.C., Aldrian-Herrada, G., Charnet, P., Divita, G. and Heitz, F. (2004) On the mechanism of non-endosomial peptide-mediated cellular delivery of nucleic acids. Biochim. Biophys. Acta 1667, 141–147.
Morris, M.C., Depollier, J., Méry, J., Heitz, F. and Divita, G. (2001) A peptide carrier for the delivery of biologically active proteins into mammalian cells. Nat. Biotechnol. 19, 1173–1176.
Deshayes, S., Heitz, A., Morris, M.C., Charnet, P., Divita, G. and Heitz, F. (2004) Insight into the mechanism of internalization of the cell-penetrating carrier peptide Pep-1 through conformational analysis. Biochemistry 43, 1449–1457.
Rittner, K., Benavente, A., Bompard-Sorlet, A., Heitz, F., Divita, G., Brasseur, R. and Jacobs, E. (2002) New basic membrane-destabilizing peptides for plasmid-based gene delivery in vitro and in vivo. Mol. Ther. 5, 104–114.
Crombez, L., Aldrian-Herrada, G., Konate, K., Nguyen, Q.N., McMaster, G.K., Brasseur, R., Heitz, F. and Divita, G. (2009) A new potent secondary amphipathic cell-penetrating peptide for siRNA delivery into mammalian cells. Mol. Ther. 17, 95–103.
Konate, K., Crombez, L., Deshayes, S., Decaffmeyer, M., Thomas, A., Brasseur, R., Aldrian, G., Heitz, F. and Divita, G. (2010) Insight into the cellular uptake mechanism of a secondary amphipathic cell penetrating peptide for siRNA delivery. Biochemistry 49, 3393–3402.
Méry, J., Granier, C., Juin, M. and Brugidou, J. (1993) Disulfide linkage to polyacrylic resin for automated Fmoc peptide synthesis. Immunochemical applications of peptide resins and mercaptoamide peptides. Int. J. Pept. Protein Res. 42, 44–52.
Vidal, P., Chaloin, L., Méry, J., Lamb, N., Lautredou, N., Bennes, R. and Heitz, F. (1996) Solid-phase synthesis and cellular localization of a C- and/or N-terminal labelled peptide. J. Pept. Sci. 2, 125–133.
Maget-Dana, R. (1999) The monolayer technique: a potent tool for studying the interfacial properties of antimicrobial and membrane-lytic peptides and their interactions with lipid membranes. Biochim. Biophys. Acta 1462, 109–140.
Brockman, H. (1999) Lipid monolayers: why use half a membrane to characterize protein-membrane interactions? Curr. Opin. Struct. Biol. 9, 438–443.
Calvez, P., Bussières, S., Demers, E. and Salesse, C. (2009) Parameters modulating the maximum insertion pressure of proteins and peptides in lipid monolayers. Biochimie 91, 718–733.
Greenfield, N. and Fasman, G.D. (1969) Computed circular dichroism spectra for the evaluation of protein conformation. Biochemistry 8, 4108–4116
Chen, Y.H., Yang, J.T. and Chau, K.H. (1974) Determination of the helix and beta form of proteins in aqueous solution by circular dichroism. Biochemistry 13, 3350–3359.
Yang, J.T., Wu, C.S. and Martinez, H.M. (1986) Calculation of protein conformation from circular dichroism. Methods Enzymol. 130, 208–269.
Griffiths, P.R. and De Haseth, J.A. (1986) Fourier transform infrared spectrometry in Chemical Analysis. John Wiley and Sons Inc., New York.
Surewicz, W.K. and Mantsch, H.H. (1988) New insight into protein secondary structure from resolution-enhanced infrared spectra. Biochim. Biophys. Acta. 952, 115–130.
Jackson, M. and Mantsch, H.H. (1995) The use and misuse of FTIR spectroscopy in the determination of protein structure. Crit. Rev. Biochem. Mol. Biol. 30, 95–120.
Acknowledgments
This work was supported in part by the Centre National de la Recherche Scientifique (CNRS) and by grants from the ANR (Agence Nationale de la Recherche, ANR-06-BLAN-0071), and the European Community (QLK2-CT-2001-01451). We thank May C. Morris (CRBM-UMR5237-CNRS) for critical reading of the manuscript and all members of the laboratory and our collaborators for fruitful discussions.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2011 Springer Science+Business Media, LLC
About this protocol
Cite this protocol
Deshayes, S., Konate, K., Aldrian, G., Heitz, F., Divita, G. (2011). Interactions of Amphipathic CPPs with Model Membranes. In: Langel, Ü. (eds) Cell-Penetrating Peptides. Methods in Molecular Biology, vol 683. Humana Press. https://doi.org/10.1007/978-1-60761-919-2_4
Download citation
DOI: https://doi.org/10.1007/978-1-60761-919-2_4
Published:
Publisher Name: Humana Press
Print ISBN: 978-1-60761-918-5
Online ISBN: 978-1-60761-919-2
eBook Packages: Springer Protocols