In vivo studies of HDL assembly and metabolism using adenovirus-mediated transfer of apoA-I mutants in apoA-I-deficient mice

We have used adenovirus-mediated gene transfer in apoA-I-deficient (A-I-/-) mice to probe the in vivo assembly and metabolism of HDL using apoA-I vari ants, focusing primarily on the role of the C-terminal 32 amino acids (heli ces 9-10). Lipid, lipoprotein, and apoA-I analyses showed that plasma level s of apoA-I and HDL of the mutants were 40-88% lower than that of wild type (WT) human apoA-I despite comparable levels of expression in the liver. WT apoA-I and mutant 1 (P165A, E172A) formed spherical particles with the siz e and density of HDL2 and HDL3. Mutant 2 (E234A, E235A, K238A, K239A) gener ated spherical particles with density between HDL2 and HDL3. Mutant 3 (L211 V, L214V, L218V, L219V) and mutant 4 (L222K, F225K, F229K), which have subs titutions of hydrophobic residues in the C-terminus, generated discoidal HD L particles indicating a defect in their conversion to mature spherical HDL . Significant amounts of mutant 4 and mutant 5 (truncated at residue 219) w ere found in the lipid poor fractions after ultracentrifugation of the plas ma (18 and 35%, respectively, of total apoA-I). These findings suggest that hydrophobic residues in and/or between helices 9 and 10 are important for the maturation of HDL in vivo.