Molecular Mechanism of Apolipoprotein E Binding to Lipoprotein Particles

Abstract

The exchangeability of apolipoprotein (apo) E between lipoprotein particles such as very low density lipoprotein (VLDL) and high density lipoprotein (HDL) is critical for lipoprotein metabolism but, despite its importance, the kinetics and mechanism of apoE-lipoprotein interaction are not known. We have used surface plasmon resonance (SPR) to monitor in real time the reversible binding of apoE to human VLDL and HDL3; biotinylated lipoproteins were immobilized on a streptavidin-coated SPR sensor chip and solutions containing various human apoE molecules at different concentrations were flowed across the surface. Analysis of the resultant sensorgrams indicated that the apoE3/lipoprotein interaction is a two-step process. After an initial interaction, the second slower step involves opening of the N-terminal helix bundle domain of the apoE molecule. Destabilization of this domain leads to more rapid interfacial rearrangement as is seen when comparing the lipoprotein binding of apoE4 relative to that of apoE3. The resultant differences in interfacial packing seem to underlie the differing abilities of apoE4 and apoE3 to bind to VLDL and HDL3. The measured dissociation constants for apoE binding to these lipoprotein particles are in the micromolar range. C-terminal truncations of apoE to remove the lipid binding region spanning residues 250–299 reduces binding to both types of lipoprotein but the effect is less with HDL3; this suggests that protein-protein interactions are important for apoE binding to this lipoprotein while apoE-lipid interactions are more significant for VLDL binding. The two-step mechanism of lipoprotein binding exhibited by apoE is likely to apply to other members of the exchangeable apolipoprotein family.

Surface Plasmon Resonance Assay of Binding Properties of Antisense Oligonucleotides to Serum Albumins and Lipoproteins

“…32 In the case, the response of analytes did not increase with increases in analyte concentration, indicating that the system had reached saturation. As the immobilization amount of LDL was increased to approximately 11000 RU, the responses of Cho-ASO in the LDL immobilized sensor chip were investigated.…”

Helical structure, stability, and dynamics in human apolipoprotein E3 and E4 by hydrogen exchange and mass spectrometry

“…Human apoE3 and apoE4 were expressed in Escherichia coli as thioredoxin fusion proteins and isolated and purified as described previously (19,20). Cleavage with thrombin leaves the target apoE with two extra amino acids, glycine and serine (designated residues -2, -1), at the N terminus that do not significantly alter the properties of the protein.…”

“…The C112R substitution in apoE4 leads to unfolding of certain helical segments that reduces selfassociation and is expected to enhance the binding of apoE4 to triglyceride-rich lipoprotein particles in plasma and to amyloid-β deposits in the brain. hydrophobic surface that mediates the self-association of apoE in a monomer−tetramer equilibrium (18), its binding to phospholipid in lipoprotein particles (19)(20)(21), and its interaction with amyloid-β (13).…”

“…Interaction of apoE with lipid surfaces is initiated by the C domain (24). Whereas the lipid-free apoE3 and E4 tetramers do not interact with the LDLR because the binding site in N-domain helix 4 is masked, lipid-bound apoE in which the N-terminal helix bundle is opened (19,20) does interact (1). Importantly, the apoE4 C112R substitution is located in helix 3 (residues 89-125) of the N-terminal helix bundle, but the substitution also alters lipid-binding activity mediated by the C-terminal domain.…”

Cholesterol-lowering phytosterols: factors affecting their use and efficacy

“…56,57 In contrast, SR-BI and CD36 facilitate intestinal uptake of sterols to the membrane but did not affect cholesterol absorption in mice. 58 These transporters are also present in the liver and perform a similar role of transporting sterols across the biliary canalicular membrane. To date, little work has been done to determine the transport of phytosterols involving these proteins or the direct competition between phytosterols and cholesterol for these transporters.…”