Kinetic and Thermodynamic Analyses of Spontaneous Exchange between High-Density Lipoprotein-Bound and Lipid-Free Apolipoprotein A-I

Abstract

It is thought that apolipoprotein A-I (apoA-I) spontaneously exchanges between high-density lipoprotein (HDL)-bound and lipid-free states, which is relevant to the occurrence of preβ-HDL particles in plasma. To improve our understanding of the mechanistic basis for this phenomenon, we performed kinetic and thermodynamic analyses for apoA-I exchange between discoidal HDL-bound and lipid-free forms using fluorescence-labeled apoA-I variants. Gel filtration experiments demonstrated that addition of excess lipid-free apoA-I to discoidal HDL particles promotes exchange of apoA-I between HDL-associated and lipid-free pools without alteration of the steady-state HDL particle size. Kinetic analysis of time-dependent changes in NBD fluorescence upon the transition of NBD-labeled apoA-I from HDL-bound to lipid-free state indicates that the exchange kinetics are independent of the collision frequency between HDL-bound and lipid-free apoA-I, in which the lipid binding ability of apoA-I affects the rate of association of lipid-free apoA-I with the HDL particles and not the rate of dissociation of HDL-bound apoA-I. Thus, C-terminal truncations or mutations that reduce the lipid binding affinity of apoA-I strongly impair the transition of lipid-free apoA-I to the HDL-bound state. Thermodynamic analysis of the exchange kinetics demonstrated that the apoA-I exchange process is enthalpically unfavorable but entropically favorable. These results explain the thermodynamic basis of the spontaneous exchange reaction of apoA-I associated with HDL particles. The altered exchangeability of dysfunctional apoA-I would affect HDL particle rearrangement, leading to perturbed HDL metabolism.

Effects of aspirin in combination with EPA and DHA on HDL-C cholesterol and ApoA1 exchange in individuals with type 2 diabetes mellitus

“…A key atheroprotective function of apoA-I is its ability to exchange on and off of HDL particles. 30–32 Lipid-poor/lipid-free apoA-I is the preferred substrate of ATP binding cassette transporter A1 (ABCA1). 30,33,34 Since apoA-I is not expressed in the intima but arrives there associated with HDL, apoA-I exchange is a critical step to ABCA1-mediated cholesterol efflux and de novo HDL biogenesis.…”

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“…ApoA-I is conformationally dynamic and lipid-free ApoA-I exchanges with ApoA-I bound to HDL particles, a process critical to HDL formation, maturation, and reverse cholesterol transport. [34][35][36][37] Lipid-free ApoA-I undergoes significant conformational changes when exchanging onto HDL particles. The addition of a nitroxide spin label permits this change to be monitored by EPR.…”

“…ApoA-I is the major protein component of HDL particles, and lipid-free ApoA-I can exchange spontaneously with HDL-bound ApoA-I, permitting in vitro quantitation of HDL-ApoA-I exchange as a measure of HDL particle functionality in human plasma. [34][35][36][37] HDL-ApoA-I exchange was reduced in sickle plasma, indicating dysfunctionality of the sickle HDL and this decrease was more pronounced during vaso-occlusive episodes (VOE) of sickle patients even after they received RBC concentrate transfusion. Our data indicate that the highly oxidative environment of sickle blood induced sulfhydryl alterations of the essential components in both plasma and RBC to maintain CE formation.…”

High-density lipoproteins for therapeutic delivery systems

“…10 Apolipoprotein association with HDL is a dynamic process, as apolipoprotein spontaneously exchanges between its HDL-bound and lipid-free state. 11 HDLs have also been found to carry microRNA, and the microRNA complement of HDL differs in patients with varying disease states. 6 This rich structural and signaling diversity of high-density lipoproteins raises the possibility that there exists within HDL functionally specialized subpopulations with tailored combinations of biological macromolecules.…”

“…HDL particles are highly stable structures 30, 31 whose stability is both of thermodynamic as well as kinetic origin. 11, 30-33 Further, their built-in receptor-mediated interactions 29 allow for the potential for specific targeting. 3 The concentration of HDL particles in the blood is quite high, approximately 30 μM, suggesting that HDL biomimetics could be tolerated at similar concentrations as well.…”