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, C-tether, 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 C-tether, we hypothesize that both leaflets of the bilayer are morphologically decoupled at high C-tether 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.