STRUCTURAL DOMAIN OF APOLIPOPROTEIN-A-I INVOLVED IN ITS INTERACTION WITH CELLS

Apolipoprotein A-I (ape A-I) is the major protein constituent of high- density lipoprotein (HDL), the lipoprotein fraction which mediates the reverse cholesterol transport. This apolipoprotein plays an important role in the binding of HDL to cells and participates in the efflux of cellular cholesterol. We have recently compared six different genetic variants of apo A-I and found that the apo A-I (Pro 165 --> Arg) muta nt is defective in promoting cellular cholesterol efflux from murine a dipocytes and peritoneal macrophages and we have proposed that this re gion of apo A-I may be involved in their interaction with cells. To co nfirm this hypothesis, four monoclonal antibodies (mAbs) specific for apo A-I were used to study the inhibition of the interaction of palmit oyloleoylphosphatidylcholine(POPC):apoA-I complexes with HeLa cells an d adipocytes. Among these antibodies, the apo A-I epitope recognized b y the A44 mAb lies in the COOH terminal region (amino acid residues 14 9-186) including the proposed region. The antibodies A05, and A03 reac t with residues 25-82, 135-140, respectively and the A11 mAb correspon ds to a discontinuous epitope at residues 99-105 and 126-132. Our resu lts show clearly that the A44 and A05 mAbs reduce both the binding to HeLa cells and the cholesterol efflux from adipocytes. The inhibition of POPC:apoA-I complexes binding to both cell types is more strictly o bserved with the Fab fragments of monoclonal antibodies A44 and A05. P artial cotitration curves of these mAbs in a solid phase assay (RIA), indicated partial competition between these two antibodies. We propose a structural model for the POPC:apoA-I complexes where the N-terminal domain of one apo A-I molecule is in close spatial relationship with the C-terminal domain of the adjacent apo A-I molecule. We therefore s uggest that the domain around amino acid 165 of apo A-I and which is r ecognized by mAb A44 (149-186) farms or contains some specific regions which mediate selectively the interaction with the binding site of ce lls and is involved in the efflux of cellular cholesterol.