Lipid-protein interactions in high density lipoproteins

Area of Science:

• Lipid and lipoprotein metabolism
• Protein biochemistry
• Structural biology

Background:

• High density lipoprotein (HDL) plays a crucial role in reverse cholesterol transport.
• Apolipoproteins, particularly apoA-I and apoA-II, are key structural and functional components of HDL.
• Understanding the in vitro recombination of apolipoproteins with lipids is essential for elucidating HDL assembly and function.

Purpose of the Study:

• To investigate the in vitro recombination capabilities of various apolipoproteins and their fragments with specific phospholipids.
• To determine the structural and physical properties of reconstituted lipid-protein complexes.
• To explore the role of protein-protein interactions in the assembly of reconstituted HDL particles.

Main Methods:

• In vitro recombination of delipidated apoHDL, isolated apolipoproteins (apoA-I, apoA-II, apoC-III), modified apoA-II, and apolipoprotein fragments with phosphatidylcholine (PC) and sphingomyelin (SPM).
• Analysis of reconstituted complexes using ultracentrifugal flotation, agarose gel chromatography, and circular dichroism.
• Characterization of particle size, lipid-protein binding, and helical structure.

Main Results:

• Delipidated apoHDL, apoA-II, and S-carboxymethylated apoA-II efficiently recombined with PC or SPM, forming particles similar in size to native HDL.
• The COOH-terminal CNBr peptide of apoA-II recombined with lipids, but the NH(2)-terminal peptide did not.
• ApoA-I and its COOH-terminal CNBr peptide showed limited recombination with lipids, suggesting the importance of apoA-I/apoA-II interactions for apoA-I integration.
• Circular dichroism analysis revealed an increase in helical structure upon lipid-protein binding in reconstituted complexes.
• Reconstituted particles exhibited properties similar to native lipoproteins.

Conclusions:

• ApoA-II and its COOH-terminal fragment are readily incorporated into lipid structures, contributing to the formation of HDL-like particles.
• ApoA-I integration into reconstituted HDL is dependent on interactions with other apolipoproteins, likely apoA-II.
• Reconstitution with lipids induces conformational changes, increasing the helical content of apolipoproteins.
• These findings provide insights into the molecular mechanisms underlying HDL assembly and the functional roles of its constituent apolipoproteins.