Abstract 560: Protein and Lipid Composition of HDL Particles

Abstract

Objective: HDL cell-cholesterol efflux capacity is influenced by both the concentration and the functionality (efflux capacity per particle) of HDL particles. It is assumed that the lipid and protein composition of HDL particles significantly influence their functionality; however there is little data to support that. Our aim was to determine the protein and lipid composition of individual HDL particles to better understand the large variability observed in HDL functionality studies. Approach: ApoA-I-containing HDL particles (preβ-1, α-4, α-3, α-2, and α-1) were separated by non-denaturing 2d-PAGE, electro-eluted from the gel, and purified by immuno-affinity chromatography using anti-human apoA-I IgG. The protein and lipid composition of the particles were assessed by mass-spectrometry. The number of apoA-I molecules in each particle was determined by cross-linking experiments. Results: The number of protein species, associated with HDL particles, increased with particle size: the largest (α-1) particles carried more than 50, while the smallest (preβ-1) particles carried only a few different proteins. ApoE was present in trace amount, if any, in these HDL particles. The apoA-II/apoA-I ratio was very low in the α-1, α-4, and preβ-1 particles but substantially higher in the α-2 and α-3 particles. The maximum number of apoA-I molecules was 4 in α-1 and 3 in α-2, α-3, and α-4 particles. The largest (α-1) particles contained significantly more lipid molecules, especially triglycerides, for each apoA-I molecule compared to smaller particles. The lipid composition of the intermediate-size α-2 and α-3 particles was similar. In contrast to larger HDL particles, the small α-4 particles were enriched in sphingomyelin. Interestingly, the very-small preβ-1 particles contained about equal amounts of polar (phospholipids and free cholesterol) and non-polar (cholesteryl ester and triglycerides) lipids. Phospholipids were represented only by sphingomyelin. These findings contradict previous models that indicated that each preβ-1 particle contained one or two apoA-I and a few polar lipid molecules. Conclusions: Both the protein and the lipid composition of HDL particles are size dependent. Further studies are needed to determine how the protein and lipid composition of HDL particles influence HDL functions.