Transbilayer coupling of obstructed lipid diffusion in polymer-tethered phospholipid bilayers

The current study reports on the inter-leaflet coupling of obstructed lipid diffusion in a polymer-tethered phospholipid bilayer, where the obstruction of diffusion is caused by lipopolymers which form non-bilayer-spanning membrane pinning sites in the bottom leaflet of the bilayer. Monolayer-specific wide-field single molecule fluorescence microscopy experiments of fluorescence-tagged phospholipids (TRITC-DHPE) over a wide range of lipopolymer concentrations, ctether, reveal a strong, polymer-induced inter-leaflet coupling of obstructed lipid diffusion for different types of lipopolymers, including those based on poly(ethylene glycol), poly(2-methyl-2-oxazoline), and poly(2-ethyl-2-oxazoline) as the hydrophilic polymer moiety. Remarkably, the degree of inter-leaflet diffusional coupling can be regulated by the cholesterol (CHOL) content which affects membrane bending elasticity. This latter finding suggests that the inter-leaflet coupling of obstructed diffusion is caused by polymer-induced bilayer deformations around membrane pinning sites, thus creating membrane-spanning regions of high membrane tension. Because the inter-leaflet coupling of obstructed diffusion at an elevated CHOL molar concentration also decreases with increasing ctether, we hypothesize that both leaflets of the bilayer are morphologically decoupled at high ctether with the outer (lipopolymer-free) monolayer being flatter than the inner one. Our findings could be of biological relevance because a similar mechanism of transbilayer coupling of obstructed diffusion may occur in some regions of cellular membranes.