Nanodiscs are phospholipid-protein complexes which are relevant to nascent high-density lipoprotein and are applicable as a drug carrier and a tool to immobilize membrane proteins. We evaluated the structure and dynamics of the nanoparticles consisting of dimyristoylphosphatidylcholine (DMPC) and apolipoprotein A-I (apoA-I) with small-angle neutron scattering (SANS) and fluorescence methods and compared them with static/dynamic properties for large unilamellar vesicles. SANS revealed that the nanodisc includes a lipid bilayer with a thickness of 44 A and a radius of 37 A, in which each lipid occupies a smaller area than the reported molecular area of DMPC in vesicles. Fluorescence measurements suggested that DMPC possesses a lower entropy in nanodiscs than in vesicles, because apoA-I molecules, which surround the bilayer, force closer lipid packing, but allow water penetration to the acyl chain ends. Time-resolved SANS experiments revealed that nanodiscs represent a 20-fold higher lipid transfer via an entropically favorable process. The results put forward a conjunction of static/dynamic properties of nanodiscs, where the entropic constraints are responsible for the accelerated desorption of lipids.

We applied a time-resolved small-angle neutron scattering technique to vesicle systems to determine interparticle transfer and flip-flop of phospholipids. Measurements were performed for large unilamellar vesicles, consisting of dimyristoylphosphatidylcholine (DMPC), 1-palmitoyl-2-oleoylphosphatidylcholine (POPC), or 1-palmitoyl-2-oleoylphosphatidic acid (POPA), which differ either in their acyl chains or headgroup. POPC, which is analogous to naturally occurring phosphatidylcholines, exhibited no transbilayer transfer and very slow interbilayer migration. POPC on the inner leaflet of vesicles did not flop even when phospholipase D converted all POPC molecules on the outer leaflet into POPA, which was shown to exhibit fast flip-flop. From these results, together with the observation that the flip-flop of DMPC was entirely inhibited in the presence of cholesterol, it is deduced that the flip-flop of phosphatidylcholines does not take place spontaneously in cellular plasma membranes rich in cholesterol and that it requires enzymatic activities of energy-dependent and/or -independent flippases/floppases.

We investigated interparticle transfer of cholesterol (Chol) between large unilamellar vesicles (LUVs) and phospholipid bilayer nanodiscs. The Chol transfer rate from LUVs to nanodiscs was decreased by an increase in the Chol content or incorporation of sphingomyelin in donor phosphatidylcholine/Chol LUVs but was not influenced by the lipid composition of acceptor particles. These results suggest that Chol dissociation from the lipid bilayer into aqueous phase is the rate-limiting step of the transfer and that the process depends on the fluidity of the donor membranes. The Chol dissociation rate from nanodiscs was faster than that from LUVs with similar lipid composition. Chol preferably partitioned to LUVs rather than nanodiscs, which is consistent with the faster dissociation rate from nanodiscs. The activation energy of Chol dissociation from nanodiscs was 1.7 kJ/mol lower than that from LUV, which was brought by increased (less negative) activation entropy and enthalpy. In addition, fluorescence lifetime and anisotropy data revealed that the lipid bilayer of nanodiscs is more tightly packed than that of LUVs. These results suggest that the tighter lipid packing in nanodiscs destabilizes the Chol-containing bilayer by reducing the entropy, which facilitates Chol dissociation.