Evidence for lipid-dependent structural changes in specific domains of apolipoprotein B100.

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.