Lipid vesicle formation by encapsulation of SMALPs in surfactant-stabilised droplets

Abstract

Understanding the intricate functions of membrane proteins is pivotal in cell biology and drug discovery. The composition of the cell membrane is highly complex, with different types of membrane proteins and lipid species. Hence, studying cellular membranes in a complexity-reduced context is important to enhance our understanding of the roles of these different elements. However, reconstitution of membrane proteins in an environment that closely mimics the cell, like giant unilamellar vesicles (GUVs), remains challenging, often requiring detergents that compromise protein function. To address this challenge, we present a novel strategy to manufacture GUVs from styrene maleic acid lipid particles (SMALPs) that utilises surfactant-stabilised droplets as a template. As a first step towards the incorporation of membrane proteins, this work focusses on the conversion of pure lipid SMALPs in GUVs. To evaluate the method, we produced a new form of SMA linked to fluorescein, referred to as FSMA. We demonstrate the assembly of SMALPs at the surfactant-stabilised droplet interface, resulting in the formation of GUVs when released upon addition of a demulsifying agent. The released vesicles appear similar to electroformed vesicles imaged with confocal light microscopy, but a fluorescein leakage assay and cryo-TEM imaging reveal their porous nature, potentially as a result of residual interactions of SMA with the lipid bilayer. Our study represents a significant step towards opening new avenues for comprehensive protein research in a complexity-reduced, yet biologically relevant, setting.

This research has received funding from the Research Foundation Flanders with grant No 1S43521N and No G074321N. Additionally, this work received funding from the European Union's Horizon Europe research and EIC grant agreement No 101046894 (SynEry) and No 101130715 (ArTCell). Views and opinions expressed are however those of the author(s) only and do not necessarily reflect those of the European Union or the granting authority European Union's Horizon Europe research and innovation program. Neither the European Union nor the granting authority can be held responsible for them.