Use of Spontaneous Membrane Translocating Peptide for Cytosolic Delivery of Biologically Active Polar Peptide

Abstract

The hydrophobic nature of cell membranes prevents the cytoplasmic entry of potentially useful polar cargoes. To evade this issue, the spontaneous membrane translocating peptides (SMTPs) were previously selected via orthogonal high-throughput screening for their ability to spontaneously translocate through synthetic lipid bilayers. These SMTPs are also able to translocate through the plasma membranes of cells, carrying polar dyes as cargo. Based on this preliminary data, we hypothesize that SMTPs can be used for cellular delivery of some biologically active, membrane-impermeant polar cargoes. To test this hypothesis, we use a bicyclic polar peptide phalloidin, a toxin isolated from the deadly Amanita phalloides “death cap” mushroom, as a cargo and linked it to one of the SMTPs, TP2. Phalloidin inhibits actin polymerization, therefore we perform assays of cell mobility and viability to determine the ability of TP2 to translocate phalloidin through cell membrane into the cytoplasm. Our results verify the cytoplasmic entry of this peptide-cargo conjugate, showing that SMTPs can deliver useful cargoes to living cells.