Apolipoprotein B-38.9 does not associate with apo[a] and forms two distinct HDL density particle populations that are larger than HDL.
We have identified a new truncated apolipoprotein B (apoB) that provides insights into the interaction of apoB with apo[a] and with lipids. Both total and LDL-cholesterol were below the 5th percentile in the proband; Lp[a] was 28 mg/dl. Four other affected individuals were identified in this kindred. Immunoblotting of plasma apoB-containing lipoproteins with an anti-apoB monoclonal antibody revealed a major band for apoB-100 and a minor band with apparent M(r) 217 kDa. The apoB truncation is due to a -1 frameshift mutation, consisting of a cytosine deletion at cDNA position 5444, that results in the translation of 22 novel amino acids terminating at residue 1767. The mutation was confirmed in the affected subjects by allele-specific oligonucleotide (ASO) analysis. Gel filtration of whole plasma revealed that the minority of apoB-38.9 eluted with IDL- and LDL-sized particles, while the majority (approximately 60%) eluted between LDL and HDL. Lp[a] eluted between VLDL and LDL. Upon preparative density gradient ultracentrifugation (DGUC), the majority of the plasma apoB-38.9 (approximately 65%) floated at a density of 1.12 g/ml coincident with the major peak of HDL cholesterol. Lp[a] floated at a peak density of 1.08 g/ml between LDL and HDL. Immunoblots of the apoB-38.9-containing HDL density DGUC fractions subjected to nondenaturing gradient gel electrophoresis (GGE) demonstrated two apoB-38.9-containing particle populations with diameters of approximately 15 nm and approximately 18 nm, respectively. Lipoproteins of these sizes were also detected when whole plasma was subjected to GGE and immunoblotting. The 15-18 nm lipoproteins correspond to the gel filtration populations eluting between LDL and HDL. Lysine-Sepharose chromatography of plasma yielded retained products that contained apo[a] and apoB-100 but not apoB-38.9. Immunoprecipitation of whole plasma with monospecific polyclonal anti-human apo[a] showed apo[a] and apoB-100, but no apoB-38.9 to be present in precipitates. ApoB-100 and apoB-38.9 were present in supernates. In in vitro incubations, recombinant apo[a] formed complexes with apoB-100 but not with apoB-38.9-containing particles. Our results show that the apoB-38.9 protein can be found in a variety of lipoproteins; however, the majority of apoB-38.9-containing lipoproteins float at a density equivalent to HDL but are larger than HDL, being intermediate in size between apoB-100 LDL and HDL. The heterogeneity of apoB-38.9 lipoproteins may reflect their dual tissue source, i.e., liver and intestine, and the discordance between size and density indicates a disproportionately reduced association of lipids with apoB-38.9. Finally, our data suggest that apoB-38.9 is incapable of forming complexes with apo[a] in plasma.
Groenewegen
W A
WA
Department of Medicine, Washington University School of Medicine, St. Louis, MO 63110.
Averna
M R
MR
Pulai
J
J
Krul
E S
ES
Schonfeld
G
G
D000368:Aged; D000595:Amino Acid Sequence; D001054:Apolipoproteins A / Q000378:metabolism*; D001055:Apolipoproteins B / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D002499:Centrifugation, Density Gradient; D002850:Chromatography, Gel; D017353:Gene Deletion; D006801:Humans; D008075:Lipoproteins, HDL / Q000737:chemistry / Q000378:metabolism*; D008297:Male; D008969:Molecular Sequence Data; D009154:Mutation; D010316:Particle Size; D010375:Pedigree; D011994:Recombinant Proteins / Q000378:metabolism
D001054:Apolipoproteins A; D001055:Apolipoproteins B; D008075:Lipoproteins, HDL; D011994:Recombinant Proteins