Direct translocation of cell penetrating peptides in liposomes: a combined mass spectrometry quantification and fluorescence detection study
| Autor: | Sandrine Sagan, Sophie Cribier, Marie-Lise Jobin, Lucrèce Matheron, Isabel D. Alves, Stéphane Chaignepain, Astrid Walrant |
|---|---|
| Přispěvatelé: | Biologie intégrative (FRBI), Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Pierre et Marie Curie - Paris 6 (UPMC), Université Pierre et Marie Curie - Paris 6 (UPMC), Institut Européen de Chimie et Biologie, Chimie et Biologie des Membranes et des Nanoobjets (CBMN), Université de Bordeaux (UB)-École Nationale d'Ingénieurs des Travaux Agricoles - Bordeaux (ENITAB)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Institut de Biologie Intégrative (IFR-BI), Université Pierre et Marie Curie - Paris 6 (UPMC)-Ecole Superieure de Physique et de Chimie Industrielles de la Ville de Paris (ESPCI Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS) |
| Jazyk: | angličtina |
| Rok vydání: | 2013 |
| Předmět: |
Biophysics
Peptide CHO Cells Cell-Penetrating Peptides 010402 general chemistry Endocytosis 01 natural sciences Biochemistry Cell membrane 03 medical and health sciences Cricetulus Cricetinae Fluorescence microscope medicine Animals Amino Acid Sequence Molecular Biology Peptide sequence 030304 developmental biology chemistry.chemical_classification 0303 health sciences Liposome Chemistry [CHIM.ORGA]Chemical Sciences/Organic chemistry Cell Membrane Cell Biology 0104 chemical sciences Matrix-assisted laser desorption/ionization Protein Transport medicine.anatomical_structure Membrane Spectrometry Fluorescence Spectrometry Mass Matrix-Assisted Laser Desorption-Ionization Liposomes |
| Zdroj: | Analytical Biochemistry Analytical Biochemistry, Elsevier Masson, 2013, 438 (1), pp.1-10. ⟨10.1016/j.ab.2013.03.009⟩ Analytical Biochemistry, 2013, 438 (1), pp.1-10. ⟨10.1016/j.ab.2013.03.009⟩ |
| ISSN: | 0003-2697 1096-0309 |
| Popis: | Cell-penetrating peptides (CPPs) can cross cell membranes in a receptor-independent manner. Two main routes for their cellular uptake have been proposed: endocytosis and direct translocation through the cell membrane. The ability of a peptide to enter cells through direct translocation can be assessed by evaluating the amount of peptide crossing the membrane of liposomes. Most methods reported so far rely on the use of fluorescent probes, which, when attached to a CPP, often alter its physical/chemical properties. Herein, a matrix-assisted laser desorption/ionization time-of-flight MS-based method is described to quantify the amount of CPP taken up into lipid vesicles and to distinguish it from the amount that is bound or inserted in the membrane. For comparison, visualization of the uptake of the same, but fluorophore-labeled, peptides into giant vesicles and cells by fluorescence microscopy is also reported. We show that membrane charge density is an important factor for direct translocation. We also show that fluorophore-labeled peptides have a different translocation behavior and that they are more toxic to cells. Alternative methods to fluorescence, such as the one reported herein, should be favored when investigating the uptake mechanism of CPPs, as fluorescent dyes can alter short peptides’ physical/chemical properties and their internalization capacities. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|
Full Text from ScienceDirect