Disruption of the C-terminal helix by single amino acid deletion is directly responsible for impaired cholesterol effl ux ability of apolipoprotein A-I Nichinan

Abstract

Apolipoprotein A-I (apoA-I) Nichinan, a naturally occurring variant with δ E235 in the C terminus, is associated with low plasma HDL levels. Here, we investigated the tertiary structure, lipid-binding properties, and ability to induce cellular cholesterol effl ux of apoA-I Nichinan and its C-terminal peptide. Thermal and chemical denaturation experiments demonstrated that the δ E235 mutation decreased the protein stability compared with wild type (WT). ApoA-I Nichinan exhibited capabilities to bind to or solubilize lipid vesicles that are intermediate to that of WT and a L230P/ L233P/Y236P variant in which the C-terminal α-helix folding is completely disrupted and forms relatively larger and unstable discoidal complexes, indicating that perturbation of the C-terminal α-helical structure by the δ E235 mutation leads to reduced lipid binding. Supporting this, apoA-I 209-241/ δ E235 peptide showed signifi cantly decreased ability to form α-helix both in the lipid-free and lipid-bound states, and reduced effi ciency to solubilize vesicles. In addition, both apoA-I Nichinan and its C-terminal peptide exhibited reduced activity in ABCA1-mediated cellular cholesterol ef-fl ux. Thus, the disruption of the ability of the C-terminal region to form α-helix caused by the E235 deletion appears to be the important determinant of impaired lipid binding and cholesterol effl ux ability and, consequently, the low plasma HDL levels of apoA-I Nichinan probands. Copyright © 2010 by the American Society for Biochemistry and Molecular Biology, Inc.