Thermodynamic and Kinetic Stability of Discoidal High-Density Lipoprotein Formation from Phosphatidylcholine/Apolipoprotein A-I Mixture
ABSTRACT Nascent high-density lipoproteins (HDLs), which are also known as discoidal HDLs, are formed by the interaction of apolipoprotein A-I (apoA-I) with transmembrane ATP-binding cassette transporter A1 (ABCA1). However, the molecular mechanism governing disc formation is not fully understood. Here, we evaluated the thermodynamic and kinetic stability of disc formation from mixtures of 1-palmitoyl-2-oleoylphosphatidylcholine and apoA-I by quantifying the discs and vesicles produced. Sodium cholate dialysis experiments revealed that the discs are thermodynamically more stable than the vesicle/apoA-I mixture (Delta*G = -52 kJ/disc mol at 37.0 degrees C) because the decrease in enthalpy (Delta*H = -620 kJ/disc mol) exceeds the decrease in entropy (TDelta*S = -570 kJ/disc mol). Circular dichroism spectral measurements ascribed 68% of the decrease in enthalpy during disc formation to the formation of helices in apoA-I. Fluorescence measurements suggested that phospholipids enclosed in the discs are more closely packed than those in the vesicles so that they are entropically destabilized. To determine if the disc could be spontaneously produced from vesicles, we measured the decrease in the turbidity of vesicles in response to the addition of apoA-I. However, the rate of disc formation was very slow, suggesting that the large kinetic barrier against disc formation makes the vesicle/apoA-I mixtures metastable. These results raise the possibility that ABCA1 may act to lower the activation energy, thereby facilitating disc formation.
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ABSTRACT: One of biology's most pervasive nanostructures, the phospholipid membrane, represents an ideal scaffold for a host of nanotechnology applications. Whether engineering biomimetic technologies or designing therapies to interface with the cell, this adaptable membrane can provide the necessary molecular-level control of membrane-anchored proteins, glycopeptides, and glycolipids. If appropriately prepared, these components can replicate in vitro or influence in vivo essential living processes such as signal transduction, mass transport, and chemical or energy conversion. To satisfy these requirements, a lipid-based, synthetic nanoscale architecture with molecular-level tunability is needed. In this regard, discrete lipid particles, including reconstituted high density lipoprotein (HDL), have emerged as a versatile and elegant solution. Structurally diverse, native biological HDLs exist as discoidal lipid bilayers of 5-8 nm diameter and lipid monolayer-coated spheres 10-15 nm in diameter, all belted by a robust scaffolding protein. These supramolecular assemblies can be reconstituted using simple self-assembly methods to incorporate a broad range of amphipathic molecular constituents, natural or artificial, and provide a generic platform for stabilization and transport of amphipathic and hydrophobic elements capable of docking with targets at biological or inorganic surfaces. In conjunction with top-down or bottom-up engineering approaches, synthetic HDL can be designed, arrayed, and manipulated for a host of applications including biochemical analyses and fundamental studies of molecular structure. Also highly biocompatible, these assemblies are suitable for medical diagnostics and therapeutics. The collection of efforts reviewed here focuses on laboratory methods by which synthetic HDLs are produced, the advantages conferred by their nanoscopic dimension, and current and emerging applications.ACS Nano 01/2011; 5(1):42-57. DOI:10.1021/nn103098m · 12.03 Impact Factor
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ABSTRACT: Plasma triglyceride-rich lipoproteins vary in their lipid composition during metabolism. We investigated the effects of cholesterol (Chol) on the surface properties of lipid emulsions and on the interactions with two amphipathic peptides, acetyl-DWLKAFYDKVAEKLKEAF-amide (Ac-18A-NH(2)) and acetyl-KWLDAFYDEVAEKLKKAF-amide (Ac-18G*-NH(2)), which differ in charge distribution. The fluorescence lifetimes of N-dansyl phosphatidylethanolamine (dansyl-PE) and n-(9-anthroyloxy)stearic acid (n-AS, n = 2, 6, and 12) were used to assess the water penetration into the headgroup and acyl chain regions of phosphatidylcholine (PC), respectively. Steady-state fluorescence anisotropy of n-AS was also performed to evaluate the acyl chain fluidity in emulsion surface monolayers. Chol decreased the fluorescence lifetime of dansyl-PE and increased the lifetimes and anisotropy values of n-AS. These results demonstrated that Chol alters the surface properties of emulsions, i.e., induces PC headgroup separation and acyl chain condensation. The two peptides showed different responses to Chol in several experiments: Addition of Chol to emulsions decreased and increased the dissociation constants of Ac-18A-NH(2) and Ac-18G*-NH(2), respectively. Furthermore, the α-helical content of Ac-18A-NH(2) was decreased by Chol, whereas that of Ac-18G*-NH(2) was unchanged. The higher reduction in helicity for Ac-18A-NH(2) is probably due to its deeper penetration than Ac-18G*-NH(2) into the hydrocarbon region of surface monolayers in the absence of Chol, which was demonstrated by Trp quenching experiments with n-AS. From these results, the charge distribution of the amphipathic helices is suggested to be a determining factor in their response to Chol enrichment in emulsions.The Journal of Physical Chemistry B 12/2011; 116(1):476-82. DOI:10.1021/jp207062h · 3.30 Impact Factor
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ABSTRACT: Apolipoprotein A-I (apo A-I), the main protein component of high-density lipoprotein (HDL), reduces the risk for atherosclerosis by removing cholesterol from the membrane of foam cells. Experiments with model membrane systems have indicated, however, that membrane cholesterol reduces apo A-I binding to the membrane. Foam cells resolve this discrepancy electrostatically by co-inserting negatively charged phospholipids in their membrane. Here we present a statistical mechanical model to account for the effect of cholesterol. Our model is based on the Haugen and May model which takes into account the dipolar nature of the zwitterionic phospholipid head group in the membrane, in which the positive end of the zwitterionic dipole moment can move randomly on a hemispherical surface with a radius equal to the arm of the dipole moment and with the negative end fixed at the hydrocarbon layer. Adsorption of a positively charged apo A-I macroion to the surface of the membrane modifies the electric field within the head group region and induces lateral demixing of phospholipid molecules in the membrane. Results from numerical integration of model equations show that i) as a result of the strong charge-dipole electrostatic coupling, the positive end of the dipoles tilts away from the adsorbed macroion in a cooperative manner; and ii) cholesterol reduces macroion adsorption to the membrane by reducing the surface area of the membrane and restricting the dipoles range of rotation. Model predictions for the change in free energy of adsorption to zwitterionic membrane are in good agreement with previously reported experimental data with liposomes. The model can assist in designing new mimetic peptides.EPL (Europhysics Letters) 07/2012; 99(1). DOI:10.1209/0295-5075/99/18003 · 2.27 Impact Factor