Multiple Intermediates in the Detergent-Induced Fusion of Lipid Vesicles

The structure, dynamics and function of lipid vesicles are heavily influenced by a range of physical forces, local microenvironmental effects and interactions with perturbative molecules, including detergents. Detergent-induced membrane interactions – critical for a wide range of applications including protein extraction and virus inactivation – varies in magnitude according to the detergent type and membrane composition, but the underlying mechanistic details remain largely under explored. Open questions relate to the precise molecular-level pathway of detergent-induced vesicle fusion, the nature of the fusion products, the influence of modulatory factors, and whether fusion states can be controllably harnessed for bionanotechnology. By using a lipid mixing assay based on Förster resonance energy transfer (FRET), and single-vesicle characterization approaches to assess vesicle heterogeneity, we identify that both freely-diffusing and surface-tethered sub-micron sized vesicles are induced to fuse by the widely-used non-ionic detergent Triton-X 100. We demonstrate that the fusion process is a multi-step mechanism, characterized by discrete values of FRET efficiency between membrane-embedded donor and acceptor fluorophores, and involves vesicle docking, hemi-fusion and full lipid mixing, even at sub-solubilizing detergent concentrations. We present evidence that the fusion process is regulated by environmental factors including membrane composition and phase, and we dissect the kinetics of vesicle fusion in contact with solid surfaces using a label free quartz-crystal microbalance with dissipation monitoring approach. The presented strategies are likely to be applicable beyond the vesicle sizes and compositions studied here, and not only provide mechanistic insight into the multifaceted dynamics of vesicle fusion but also have implications for a wide range of biotechnological applications including drug delivery, sensor development, surfactant sensing, biomimetic formation, and microfluidics, where transport and manipulation of encapsulated cargo is essential. ### Competing Interest Statement The authors have declared no competing interest.