title: Apolipoprotein B-containing lipoprotein particle assembly: lipid capacity of the nascent lipoprotein particle.
abstract: We previously proposed that the N-terminal 1000-residue betaalpha(1) domain of apolipoprotein B (apoB) forms a bulk lipid pocket homologous to that of lamprey lipovitellin. In support of this "lipid pocket" hypothesis, we demonstrated that apoB:1000 (residues 1-1000) is secreted by a stable transformant of McA-RH7777 cells as a monodisperse particle with high density lipoprotein 3 (HDL(3)) density. In contrast, apoB:931 (residues 1-931), missing only 69 residues of the sequence homologous to lipovitellin, was secreted as a particle considerably more dense than HDL(3). In the present study we have determined the stoichiometry of the lipid component of the apoB:931 and apoB:1000 particles. The secreted [(3)H]glycerol-labeled apoB:1000 particles, isolated by nondenaturing gradient gel electrophoresis, contained 50 phospholipid (PL) and 11 triacylglycerol (TAG) molecules/particle. In contrast, apoB:931 particles contained only a few molecules of PL and were devoid of TAG. The unlabeled apoB:1000 particles, isolated by immunoaffinity chromatography, contained 56 PL, 8 TAG, and 7 cholesteryl ester molecules/particle. The surface to core lipid ratio of apoB:1000-containing particles was approximately 4:1 and was not affected by oleate supplementation. Although very small amounts of microsomal triglyceride transfer protein (MTP) were associated with apoB:1000 particles, it never approached a 1:1 molar ratio of MTP to apoB. These results support a model in which (i) the first 1000 amino acid residues of apoB are competent to complete the lipid pocket without a structural requirement for MTP; (ii) a portion, or perhaps all, of the amino acid residues between 931 and 1000 of apoB-100 are critical for the formation of a stable, bulk lipid-containing nascent lipoprotein particle, and (iii) the lipid pocket created by the first 1000 residues of apoB-100 is PL-rich, suggesting a small bilayer type organization and has a maximum capacity on the order of 50 molecules of phospholipid.
authors.affiliation: Department of Medicine, Atherosclerosis Research Unit, University of Alabama at Birmingham Medical Center, Birmingham, Alabama 35294, USA.
authors.forename: Medha
authors.initials: M
authors.lastname: Manchekar
authors.forename: Paul E
authors.initials: PE
authors.lastname: Richardson
authors.forename: Trudy M
authors.initials: TM
authors.lastname: Forte
authors.forename: Geeta
authors.initials: G
authors.lastname: Datta
authors.forename: Jere P
authors.initials: JP
authors.lastname: Segrest
authors.forename: Nassrin
authors.initials: N
authors.lastname: Dashti
mesh_terms: D000818:Animals; D001055:Apolipoproteins B / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D002250:Carbon Radioisotopes; D006528:Carcinoma, Hepatocellular; D045744:Cell Line, Tumor; D005990:Glycerol / Q000378:metabolism / Q000494:pharmacology; D006801:Humans; D008854:Microscopy, Electron; D008970:Molecular Weight; D016296:Mutagenesis; D019301:Oleic Acid / Q000378:metabolism / Q000494:pharmacology; D010316:Particle Size; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D051381:Rats; D014316:Tritium
chemical_list: D001055:Apolipoproteins B; D002250:Carbon Radioisotopes; D014316:Tritium; D019301:Oleic Acid; D005990:Glycerol