Significance of the hydrophobic residues 225–230 of apoA-I for the biogenesis of HDL

We studied the significance of four hydrophobic residues within the 225-230 region of apoA-I on its structure and functions and their contribution to the biogenesis of HDL. Adenovirus-mediated gene transfer of an apoA-I[F225A/V227A/F229A/L230A] mutant in apoA-I-/- mice decreased plasma cholesterol, HDL cholesterol, and apoA-I levels. When expressed in apoA-I-/- x apoE(-/-) mice, approximately 40% of the mutant apoA-I as well as mouse apoA-IV and apoB-48 appeared in the VLDL/IDL/LDL. In both mouse models, the apoA-I mutant generated small spherical particles of pre-beta- and alpha 4-HDL mobility. Coexpression of the apoA-I mutant and LCAT increased and shifted the-HDL cholesterol peak toward lower densities, created normal alpha HDL subpopulations, and generated spherical-HDL particles. Biophysical analyses suggested that the apoA-I[225-230] mutations led to a more compact folding that may limit the conformational flexibility of the protein. The mutations also reduced the ability of apoA-I to promote ABCA1-mediated cholesterol efflux and to activate LCAT to 31% and 66%, respectively, of the WT control. Overall, the apoA-I[225-230] mutations inhibited the biogenesis of-HDL and led to the accumulation of immature pre-beta- and alpha 4-HDL particles, a phenotype that could be corrected by administration of LCAT.