Interactions of the Cell Penetrating Peptide Pep-1 with Model Membranes

The objective of this study is to measure the effect of the cell penetrating peptide Pep-1 on vesicles from single lipids, binary mixtures of lipids and Escherichia coli (E. coli) membranes mimicking model membranes using infrared (IR), UV visible, Raman, fluorescence, thermogravimetric analysis (TGA) and differential scanning calorimetry (DSC) techniques. The interaction was performed in liquid and solid phases. The single lipids are dipalmitoylphosphocholine (DPPC), dimyristoylglycerophosphorylglycerol (DPPG), dimyristoylphosphotidylethanolamine (DPPE), and cardiolipin (CL) while the binary lipid mixtures are DPPE-DPPG, DPPE- CL, and CL-DPPG. The most abundant lipids that are cholesterol (CHOL) and sphingomyelin (SPM) were also studied. The E. coli mimicking membrane used in this study is a ternary mixture of CL/DPPE/DPPG. Upon addition of E. coli membrane, the tryptophan fluorescence intensity of Pep-1 decreased in a non-concentration-manner. Fluorescence data of Pep-1 showed a concomitant decrease and blue shift in the emission intensity of the tryptophan residue through the entire range of the peptide-lipid concentration ratio. These results suggest that Pep-1 is located deep inside the hydrophobic core of the membrane. UV visible spectra demonstrate that the tryptophan and arginine residues are involved in the bonding. IR spectra indicate that Pep-1 adopts a mixture of helix, turn and sheet structures in the absence of the membranes. In the presence of the membranes, Pep-1 adopts a major helix structure. Low frequency Raman spectra revealed that the phosphate groups are involved in the bonding, Pep-1 and E coli membrane interact through CN group of choline, and Pep-1 induces ordering of the acyl chain of the E. coli membrane. TGA and DSC data revealed that the peptide Pep-1 stabilizes the E. coli membrane; E. coli and Pep-1-E.coli complex have a different moisture level, confirming the fluorescence data showing that Pep-1 is deep inside E. coli membrane in a hydrophobic environment. DSC showed that Pep-1 induced phase separation between peptide-rich and peptide-poor regions. Overall, these results show that the structure and stability of Pep-1 in an E. coli mimicking environment are substantially different from those of Pep-1 in solution, being stabilized in a specific conformation that could be important to eliciting its biological activity