The Improved Ability of ApoA-I Amyloidogenic Variants at Mediating Cholesterol Efflux Relies on their Increased Structural Flexibility

Abstract

Apolipoprotein A-I (ApoA-I), the most abundant protein in high-density lipoprotein (HDL), plays a key role in the catabolism of cholesterol. However, specific mutations in the ApoA-I protein are responsible for late-onset systemic amyloidosis. Interestingly, carriers of such ApoA-I amyloidogenic mutations do not have higher cardiovascular disease risk despite reduced ApoA-I and HDL-cholesterol levels. Here we aimed at finding a correlation between ApoA-I variants’ structural properties in HDL and their ability to mediate cholesterol mobilization. Serum samples from L75P and L174S carriers, and from healthy donors, were analyzed by immunoblot and tested for their ability to stimulate cholesterol efflux from macrophages. Reconstituted 8.4 and 9.6 nm HDL particles (rHDL) containing ApoA-I variants were used for cholesterol efflux experiments and their structures were characterized by synchrotron radiation circular dichroism (SRCD) and hydrogen-deuterium exchange mass spectrometry (HDX-MS). Stability was measured by conventional CD. We found that serum from L75P and L174S carriers possess higher capacity to catalyze cholesterol efflux from macrophages compared to controls. Same conclusion was drawn when cholesterol efflux was mediated by rHDL. The HDL particle profile from carriers showed a higher relative abundance of the 8.4 nm vs 9.6 nm particles. Low-resolution structural analysis of the particles revealed that L75P- and L174S-ApoA-I proteins in 8.4 nm rHDL are characterized by altered secondary structure composition and a more relaxed binding to lipids compared to their native counterpart. However, the structural differences and increased flexibility did not affect particle stability. In conclusion, the unfavorable lipid profile of ApoA-I amyloidogenic variants’ carriers is balanced by a higher amount of smaller HDL particles that are particularly efficient at mediating cholesterol efflux. The improved functionality appears to be due to an increased plasticity of the protein structure.