title: Evidence for lipid-dependent structural changes in specific domains of apolipoprotein B100.
abstract: The structural organization and stability of apoB100 in complexes containing triglyceride (TG) and phospholipid have been examined. LDL was delipidated to form aqueous soluble apoB100-TG complexes that retain approximately 70% of LDL TG, but contain no other lipids. The apoB100-TG complexes exhibited reduced amphipathic alpha-helical content (17%) and net negative charge (-2.9 mV) as compared to native LDL-apoB100 (49% and -6 mV, respectively). Of 28 anti-apoB monoclonal antibodies tested, 15 showed partial or full reactivity with apoB100-TG. The immunoreactive epitopes of apoB100-TG were restricted to those situated in either the amino terminal globular domain (4 of 6) or in regions of apoB100 that are predicted to be composed of amphipathic beta-strands (11 of 13). Incubation of the apoB100-TG complex with palmitoyloleoylphosphatidylcholine (POPC) spontaneously (< 10 min) formed homogeneous lipoproteins (20 nm) that contained approximately 300 molecules of POPC per particle (apoB100-PC). Phospholipidation of apoB100-TG complexes partially recovered the alpha-helical content (34%) and net negative charge (-4.9 mV) of the native LDL and restored resistance of apoB100 to denaturation by guanidine HCl (5.8 M). Addition of phospholipids to apoB100-TG also increased the immunoreactivity of specific epitopes that are located primarily in regions of apoB100 that are thought to be constituted of amphipathic beta-strands. The effects of TG and phospholipid on apoB100 conformation appear to be highly domain-specific. On the basis of these results, we propose that the beta-strands of apoB100 may represent a nonflexible lipid-associating backbone, while the amphipathic alpha-helical domains may represent flexible lipid-binding regions that allow the particle to accommodate varying amounts of lipid.
authors.affiliation: Lipoprotein and Atherosclerosis Group, University of Ottawa Heart Institute, Ontario, Canada.
authors.forename: V
authors.initials: V
authors.lastname: Chauhan
authors.forename: X
authors.initials: X
authors.lastname: Wang
authors.forename: T
authors.initials: T
authors.lastname: Ramsamy
authors.forename: R W
authors.initials: RW
authors.lastname: Milne
authors.forename: D L
authors.initials: DL
authors.lastname: Sparks
mesh_terms: D000911:Antibodies, Monoclonal / Q000378:metabolism; D053299:Apolipoprotein B-100; D001055:Apolipoproteins B / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism; D002478:Cells, Cultured; D003840:Deoxycholic Acid; D004134:Dimyristoylphosphatidylcholine / Q000378:metabolism; D005347:Fibroblasts; D006801:Humans; D050356:Lipid Metabolism; D008055:Lipids / Q000502:physiology*; D008077:Lipoproteins, LDL / Q000378:metabolism; D010713:Phosphatidylcholines / Q000378:metabolism; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011973:Receptors, LDL / Q000378:metabolism; D014280:Triglycerides / Q000378:metabolism
chemical_list: D000911:Antibodies, Monoclonal; D053299:Apolipoprotein B-100; D001055:Apolipoproteins B; D008055:Lipids; D008077:Lipoproteins, LDL; D010713:Phosphatidylcholines; D011973:Receptors, LDL; D014280:Triglycerides; D003840:Deoxycholic Acid; C028694:1-palmitoyl-2-oleoylphosphatidylcholine; D004134:Dimyristoylphosphatidylcholine