A Coiled-Coil Peptide Shaping Lipid Bilayers upon Fusion

Abstract

A system based on two designed peptides, namely the cationic peptide K, (KIAALKE) 3 , and its complementary anionic counterpart called peptide E, (EIAALEK) 3 , has been used as a minimal model for membrane fusion, inspired by SNARE proteins. Although the fact that docking of separate vesicle populations via the formation of a dimeric E/K coiled-coil complex can be rationalized, the reasons for the peptides promoting fusion of vesicles cannot be fully explained. Therefore it is of significant interest to determine how the peptides aid in overcoming energetic barriers during lipid rearrangements leading to fusion. In this study, investigations of the peptides' interactions with neutral PC/PE/cholesterol membranes by fluorescence spectroscopy show that tryptophan-labeled K* binds to the membrane (K K*~6 .2 10 3 M À1 ), whereas E* remains fully water-solvated. 15 N-NMR spectroscopy, depth-dependent fluorescence quenching, CD-spectroscopy experiments, and MD simulations indicate a helix orientation of K* parallel to the membrane surface. Solid-state 31 P-NMR of oriented lipid membranes was used to study the impact of peptide incorporation on lipid headgroup alignment. The membrane-immersed K* is found to locally alter the bilayer curvature, accompanied by a change of headgroup orientation relative to the membrane normal and of the lipid composition in the vicinity of the bound peptide. The NMR results were supported by molecular dynamics simulations, which showed that K reorganizes the membrane composition in its vicinity, induces positive membrane curvature, and enhances the lipid tail protrusion probability. These effects are known to be fusion relevant. The combined results support the hypothesis for a twofold role of K in the mechanism of membrane fusion: 1) to bring opposing membranes into close proximity via coiled-coil formation and 2) to destabilize both membranes thereby promoting fusion.

Controlled Peptide-Mediated Vesicle Fusion Assessed by Simultaneous Dual-Colour Time-Lapsed Fluorescence Microscopy

“…These control experiments reveal that K 4 has a strong interaction with the membrane of GUVs, resulting in non-specific interactions with LUVs. This interaction has been previously reported in lipid monolayer studies combined with surface sensitive infrared reflection absorption spectroscopy (IRRAS) and liposomal assays 48,49 . Therefore, for this system to be highly specific, it is important that the target membrane is functionalized with CP 4 E 4 and LUVs contain CP 4 K 4 .…”

The Multifaceted Role of SNARE Proteins in Membrane Fusion

“…Also the α-helical SNARE TM anchors were shown to be associated with bilayer destabilization (Risselada et al, 2011) and lipid mixing, discussed in the context of their oblique orientation in membranes. An increased number of lipid acyl chain protrusion events was recently also observed in curved membranes (Rabe et al, 2016; Tahir et al, 2016). Additionally, SNARE complexes were reported to induce local membrane protrusions in the target membrane (Bharat et al, 2014).…”

Benchmarking of different molecular docking methods for protein-peptide docking

“…Coiled peptide might adapt better conformational change during docking to find the near-native pose. Also, in previous studies, it has shown that the formation of coiled-coil in peptide help in better docking with the receptor molecule [68,69]. Similarly, in another study, it was shown that the docking method performed better on coiled peptides [70].…”