Role of apolipoprotein A-I in cholesterol transfer between lipoproteins. Evidence for involvement of specific apoA-I domains.

A series of monoclonal antibodies against epitopes spanning different domains of apoA-I have been tested for their effects on unesterified cholesterol transfer between low density lipoprotein (LDL) and well-defined homogenous lipoproteins reconstituted with phosphatidylcholine, cholesterol, and apoA-I (LpA-I). Antibodies 2G11 (reacting between residues 25 and 110), A05 (residues 25-82), A03 (residues 135-140), A44 and r5G9 (residues 149-186), and 4A12 (residues 173-205) significantly inhibit cholesterol transfer from LDL to Lp2A-I while they enhance transfer in the opposite direction, thus causing an increased net transfer to LDL. Most of these monoclonal antibodies (mAbs) also enhance phospholipid transfer to LDL but in a lesser and variable proportion relative to cholesterol. Their epitopes are mainly contained within domains that are predicted to be amphipathic alpha-helices. In contrast, mAbs 4H1 (residues 2-8), 3G10 (residues 96-121), and 5F6 (residues 116-141) have little or no effect on either cholesterol or phospholipid transfer, and the epitopes for these three mAbs have been shown in earlier studies to be structurally and functionally related. Their immunoreactivity responds similarly to variation in lipoprotein cholesterol content, and the antibodies binding to these sites compete with one another and have similar effects on the cholesterol esterification reaction. Thus, the current results are compatible with the hypothesis that they form an integrated domain with a common function in cholesterol metabolism, possibly as part of a hinge domain. Most mAbs were found to increase significantly the alpha-helicity of apoA-I in the Lp2A-I immunecomplexes, suggesting that they may increase the stability of the lipid-bound apoA-I. However, not unexpectedly, there is no correlation between the effects of mAbs on alpha-helicity and their effects on cholesterol or phospholipid transfer since each mAb has a discrete effect on these transfers. These studies demonstrate the specificity of LpA-I particles in cholesterol transport and document the existence of apoA-I domains with different functions in cholesterol transport.