American Chemical Society
ja9b09563_si_001.pdf (1.28 MB)

Fluorescence Monitoring of Peptide Transport Pathways into Large and Giant Vesicles by Supramolecular Host–Dye Reporter Pairs

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journal contribution
posted on 2019-12-02, 15:45 authored by Andrea Barba-Bon, Yu-Chen Pan, Frank Biedermann, Dong-Sheng Guo, Werner M. Nau, Andreas Hennig
The membrane transport mechanisms of cell-penetrating peptides (CPPs) are still controversial, and reliable assays to report on their internalization in model membranes are required. Herein, we introduce a label-free, fluorescence-based method to monitor membrane transport of peptides in real time. For this purpose, a macrocyclic host and a fluorescent dye forming a host–dye reporter pair are encapsulated inside phospholipid vesicles. Internalization of peptides, which can bind to the supramolecular host, leads to displacement of the dye from the host, resulting in a fluorescence change that signals the peptide uptake and, thus, provides unambiguous evidence for their transport through the membrane. The method was successfully validated with various established CPPs, including the elusive peptide TP2, in the presence of counterion activators of CPPs, and with a calixarene-based supramolecular membrane transport system. In addition, transport experiments with encapsulated host–dye reporter pairs are not limited to large unilamellar vesicles (LUVs) but can also be used with giant unilamellar vesicles (GUVs) and fluorescence microscopy imaging.