title: Influence of ApoA-I structure on the ABCA1-mediated efflux of cellular lipids.
abstract: The influence of apolipoprotein (apo) A-I structure on ABCA1-mediated efflux of cellular unesterified (free) cholesterol (FC) and phospholipid (PL) is not well understood. To address this issue, we used a series of apoA-I mutants to examine the contributions of various domains in the molecule to ABCA1-mediated FC and PL efflux from mouse J774 macrophages and human skin fibroblasts. Irrespective of the cell type, deletion or disruption of the C-terminal lipid-binding domain of apoA-I drastically reduced the FC and PL efflux ( approximately 90%), indicating that the C-terminal amphipathic alpha-helix is required for high affinity microsolubilization of FC and PL. Deletion in the N-terminal region of apoA-I also reduced the lipid efflux ( approximately 30%) and increased the K(m) about 2-fold compared with wild type apoA-I, whereas deletion of the central domain (Delta123-166) had no effect on either K(m) or V(max). These results indicate that ABCA1-mediated lipid efflux is relatively insensitive to the organization of the apoA-I N-terminal helix-bundle domain. Alterations in apoA-I structure caused parallel changes in its ability to bind to a PL bilayer and to induce efflux of FC and PL. Overall, these results are consistent with a two-step model for ABCA1-mediated lipid efflux. In the first step, apoA-I binds to ABCA1 and hydrophobic alpha-helices in the C-terminal domain of apoA-I insert into the region of the perturbed PL bilayer created by the PL transport activity of ABCA1, thereby allowing the second step of lipidation of apoA-I and formation of nascent high density lipoprotein particles to occur.
authors.affiliation: Division of GI/Nutrition, The Children's Hospital of Philadelphia, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 19104-4318, USA.
authors.forename: Charulatha
authors.initials: C
authors.lastname: Vedhachalam
authors.forename: Lijuan
authors.initials: L
authors.lastname: Liu
authors.forename: Margaret
authors.initials: M
authors.lastname: Nickel
authors.forename: Padmaja
authors.initials: P
authors.lastname: Dhanasekaran
authors.forename: G M
authors.initials: GM
authors.lastname: Anantharamaiah
authors.forename: Sissel
authors.initials: S
authors.lastname: Lund-Katz
authors.forename: George H
authors.initials: GH
authors.lastname: Rothblat
authors.forename: Michael C
authors.initials: MC
authors.lastname: Phillips
mesh_terms: D064286:ATP Binding Cassette Transporter 1; D018528:ATP-Binding Cassette Transporters / Q000378:metabolism*; D016632:Apolipoprotein A-I / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D002784:Cholesterol / Q000378:metabolism*; D005347:Fibroblasts / Q000378:metabolism; D006801:Humans; D008264:Macrophages / Q000378:metabolism; D009154:Mutation; D010743:Phospholipids / Q000378:metabolism*; D017384:Sequence Deletion
chemical_list: C577836:ABCA1 protein, human; D064286:ATP Binding Cassette Transporter 1; D018528:ATP-Binding Cassette Transporters; D016632:Apolipoprotein A-I; D010743:Phospholipids; D002784:Cholesterol