Membrane Lateral Mobility Obstructed by Polymer-Tethered Lipids Studied at the Single Molecule Level

Abstract

Obstructed long-range lateral diffusion of phospholipids (TRITC-DHPE) and membrane proteins (bacteriorhodopsin) in a planar polymer-tethered 1-stearoyl-2-oleoyl-sn-glycero-3-phosphocholine bilayer is studied using wide-field single molecule fluorescence microscopy. The obstacles are well-controlled concentrations of hydrophobic lipid-mimicking dioctadecylamine moieties in the polymer-exposed monolayer of the model membrane. Diffusion of both types of tracer molecules is well described by a percolating system with different percolation thresholds for lipids and proteins. Data analysis using a free area model of obstructed lipid diffusion indicates that phospholipids and tethered lipids interact via hard-core repulsion. A comparison to Monte Carlo lattice calculations reveals that tethered lipids act as immobile obstacles, are randomly distributed, and do not self-assemble into large-scale aggregates for low to moderate tethering concentrations. A procedure is presented to identify anomalous subdiffusion from tracking data at a single time lag. From the analysis of the cumulative distribution function of the square displacements, it was found that TRITC-DHPE and W80i show normal diffusion at lower concentrations of tethered lipids and anomalous diffusion at higher ones. This study may help improve our understanding of how lipids and proteins in biomembranes may be obstructed by very small obstacles comprising only one or very few molecules.

Structural characterization of an elevated lipid bilayer obtained by stepwise functionalization of a self-assembled alkenyl silane film

“…These data compare well with previous reports, where diffusion constants were even correlated with height and mole percentage of tethers. 40,41 Some have also measured diffusion constants of tethered bilayers and of bilayers supported directly on silicon and have shown that comparable high diffusion constants have been obtained in both cases. 17 The somewhat higher absolute values of the diffusion constants measured here, i.e., a minor difference of ϳ2 versus ϳ1 m 2 / s, could be inherent to the different analyzing techniques.…”

T cells like a firm molecular handshake

“…Equally critical will be the direct measurements of half-lives in adhesive interfaces. Two approaches that are currently tractable are the use of supported planar bilayers to visualize interactions (20) and erythrocyte-based interaction sensors (17), but moving these measurements into native cell-cell interfaces would also be an important challenge for the future.…”

Surface supported bilayer platform for studies of lateral association of proteins in membranes (Mini Review)

“…[15][16][17] The lipid diffusion coefficients calculated using the BPE method are very similar to previously reported values obtained using other methods, such as flash bleaching and single molecule measurements. 18 The diffusion coefficients measured for the peptides, however, were very low. We therefore explored whether the incorporation of a PEG cushion between the bilayer and the substrate increases protein mobility.…”