The Mechanism of Vesicle Solubilization by the Detergent Sodium Dodecyl Sulfate

Abstract

Membrane solubilization by sodium dodecyl sulfate (SDS) is indispensable for many established biotechnological applications, including viral inactivation and protein extraction. Although the ensemble thermodynamics have been thoroughly explored, the underlying molecular dynamics have remained inaccessible, owing to major limitations of traditional measurement tools. Here, we integrate multiple advanced biophysical approaches to gain multi-angle insight into the time-dependence and fundamental kinetic steps associated with the solubilization of single sub-micron sized vesicles in response to SDS. We find that the accumulation of SDS molecules on in-tact vesicles triggers biphasic solubilization kinetics comprising an initial vesicle expansion event followed by rapid lipid loss and micellization. Our findings support a general mechanism of detergent-induced membrane solubilization and we expect the framework of correlative biophysical technologies presented here will form a general platform for elucidating the complex kinetics of membrane perturbation induced by a wide variety of surfactants and disrupting agents.

Crowding induced morphological changes in synthetic lipid vesicles determined using smFRET

“…To further visualize the observed compaction event, we next evaluated the mean FRET response from single surface-tethered vesicles under crowding conditions by simultaneously capturing DiI and DiD fluorescence intensities using a custom built wide-field objective-type total internal reflection fluorescence microscope 45 . Prior to the extrusion process, the lipid suspension contained 1 mol% of Biotin-PE, allowing formed vesicles to be tethered to a glass substrate via BSA-Biotin and NeutrAvidin as previously described 9 . As shown in Figure 3A , the application of picomolar vesicle concentrations to a NeutrAvidin coated surface led to the detection of 195 ± 14 FRET-active vesicles per field of view (25 × 50 μm), with a mean nearest-neighbour vesicle separation distance of ∼1 μm.…”

“…1,1'-Dioctadecyl-3,3,3',3' Tetramethylindocarbocyanine Perchlorate (DiI) and 1,1-Dioctadecyl-3,3,3,3-tetramethylindodicarbocyanine (DiD) membrane stains were obtained from ThermoFisher Scientific. Synthetic lipid vesicles were prepared via the extrusion method as previously described [8][9] . Briefly, mixtures of lipids and membrane stains as described in the main text were mixed in chloroform at typical final lipid concentrations of 10 mg lipid/ml.…”

“…Furthermore, inducing porosity into the membrane using bacterial toxins such as alpha haemolysin, enables buffer exchange without washing out the encapsulated biomolecules, rendering the vesicle an effective nanocontainer for allowing the interrogation of constrained biomolecules under various solution conditions 4 . Vesicles can also be biochemically programmed to fuse, enabling lipid mixing and content exchange [5][6][7] , and the use of perturbative detergent molecules, which dramatically alter vesicle morphology [8][9] , are important in the context of lysis and for triggering the release of membrane-bound or -encapsulated biomolecules 10 .…”

“…LUVs have found particular utility as nanoscale containers for constraining biomolecules for single-molecule analysis, where inducing porosity into their membrane facilitates buffer exchange without removing the biomolecule under interrogation 4 . Additionally, vesicles can be biochemically programmed to interact, enabling lipid mixing and content exchange [5][6][7] , and the use of perturbative detergents, which alter their morphology [8][9] , are important in the context of lysis and for triggering release of encapsulated molecules 10 . It is also now clear that the structure and dynamics of synthetic vesicles are influenced by factors such as molecular crowding but new experimental approaches are required to explore these interactions further.…”

Single-molecule FRET dynamics of molecular motors in an ABEL Trap

“…A major hallmark of the FRET approach is the ability to unveil population distributions of inter-dye distances, and reveal transient conformations that may be otherwise hidden by conventional ensemble-averaging. Coupled with the ability to probe individual molecules for long periods of time (seconds to minutes) with millisecond time resolution, the FRET approach has been successfully applied to reveal conformational dynamics across a wide range of proteins [8], nucleic acids [9][10][11] and biomolecular complexes [12][13][14]. However, one has to make a choice between investigating freely diffusing molecules for inherently short observation times or extend this observation time at the price of potential impairment in biological function by attaching molecules to a surface.…”