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

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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.