Conformation of apolipoprotein B-100 in the low density lipoproteins of tangier disease. Identification of localized conformational response to triglyceride content.
The low density lipoproteins (LDL) from patients with Tangier disease are enriched in triglycerides, 27% of LDL mass versus 7% for normal LDL. To study whether this unique LDL core lipid composition affects the surface disposition of apolipoprotein (apo) B-100, we analyzed the LDL by protease digestion and in competitive radioimmunoassays. Limited proteolytic digestion of Tangier LDL by Staphylococcus aureus V8 protease generated a prominent fragment of 120 kDa (cleavage site at residue 1076), which was not visible in similarly digested normal LDL. In competitive radioimmunoassay, Tangier LDL bound weakly to the apoB-specific monoclonal antibody MB20, compared with control LDL. We localized the MB20 epitope between residues 1031 and 1084 of apoB-100, probably very near residue 1076. DNA sequencing of exon 21 of apoB genomic clones (coding for residues 1014-1084) from a Tangier patient revealed no difference from the normal DNA sequence, thus eliminating a protein polymorphism as a basis for the altered protease sensitivity and antibody binding. When the triglyceride contents of Tangier LDL were reduced to 10% of mass by incubation with normal high density lipoproteins, production of the 120-kDa fragment by proteolysis decreased and MB20 binding increased in affinity, implying a change toward normal conformation of apoB-100. Thus, using two independent techniques, proteolytic digestion and binding of monoclonal antibodies, we have demonstrated an alternative conformation of apoB-100 in the vicinity of residue 1076, which reflects the content of triglycerides in the LDL particle.
Kunitake
S T
ST
Cardiovascular Research Institute, University of California, San Francisco 94143.
Young
S G
SG
Chen
G C
GC
Pullinger
C R
CR
Zhu
S
S
Pease
R J
RJ
Scott
J
J
Hass
P
P
Schilling
J
J
Kane
J P
JP
D000595:Amino Acid Sequence; D053299:Apolipoprotein B-100; D001055:Apolipoproteins B / Q000097:blood / Q000737:chemistry / Q000235:genetics*; D001483:Base Sequence; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics; D000939:Epitopes / Q000032:analysis; D006801:Humans; D007438:Introns; D008077:Lipoproteins, LDL / Q000097:blood*; D008214:Lymphocytes / Q000378:metabolism; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes; D011487:Protein Conformation; D011863:Radioimmunoassay; D012016:Reference Values; D013631:Tangier Disease / Q000097:blood* / Q000235:genetics; D014280:Triglycerides / Q000097:blood*
D053299:Apolipoprotein B-100; D001055:Apolipoproteins B; D000939:Epitopes; D008077:Lipoproteins, LDL; D015345:Oligonucleotide Probes; D014280:Triglycerides; D004247:DNA