Definition of a nonlinear conformational epitope for the apolipoprotein B-100-specific monoclonal antibody, MB47.
The apolipoprotein (apo) B-100-specific monoclonal antibody MB47 has been widely used in lipoprotein metabolism and atherosclerosis research. When bound to apoB-100 on low density lipoproteins (LDL), antibody MB47 completely blocks the binding of LDL to the LDL receptor. The epitope for antibody MB47 has previously been mapped to the vicinity of apoB-100 amino acid (aa) residue 3500. To map the epitope for antibody MB47 more precisely, we used recombinant bacterial fusion proteins. Antibody MB47 bound strongly to a fusion protein containing apoB-100 aa 3214-3728, but no specific binding was observed to fusion proteins containing aa 3214-3351, 3214-3506, 3351-3506, or a fusion protein containing aa 3214-3351 and 3506-3728. Although antibody MB47 did not bind to aa 3214-3506, it did bind to aa 3214-3510. Further fusion protein studies revealed that antibody MB47 bound to aa 3429-3510, but bound only very weakly to aa 3453-3510, indicating that aa 3429-3453 constitute an important part of the MB47 epitope. Subsequent fusion protein studies revealed that MB47 bound much more strongly to aa 3429-3523, 3429-3544, 3429-3565, and 3429-3590 than to aa 3429-3510. Thus, aa 3507-3523 also constitute an important part of the MB47 epitope. In summary, the fusion protein data indicated that two nonlinear domains of apoB-100 separated by approximately 53 aa (the 25 residues from aa 3429 to 3453 and the 17 residues from aa 3507 to 3523) form key parts of the MB47 epitope.(ABSTRACT TRUNCATED AT 250 WORDS)
Young
S G
SG
Gladstone Institute for Cardiovascular Disease, Department of Medicine, University of California, San Francisco 94141-9100.
Koduri
R K
RK
Austin
R K
RK
Bonnet
D J
DJ
Smith
R S
RS
Curtiss
L K
LK
D000911:Antibodies, Monoclonal / Q000276:immunology*; D016632:Apolipoprotein A-I / Q000235:genetics / Q000276:immunology; D053299:Apolipoprotein B-100; D001055:Apolipoproteins B / Q000235:genetics / Q000276:immunology*; D015153:Blotting, Western; D004262:DNA Restriction Enzymes; D000939:Epitopes / Q000737:chemistry / Q000276:immunology*; D008077:Lipoproteins, LDL / Q000276:immunology / Q000378:metabolism; D010446:Peptide Fragments / Q000276:immunology / Q000378:metabolism; D010957:Plasmids; D011487:Protein Conformation; D011973:Receptors, LDL / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000276:immunology / Q000378:metabolism; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
D000911:Antibodies, Monoclonal; D016632:Apolipoprotein A-I; D053299:Apolipoprotein B-100; D001055:Apolipoproteins B; D000939:Epitopes; D008077:Lipoproteins, LDL; D010446:Peptide Fragments; D011973:Receptors, LDL; D011993:Recombinant Fusion Proteins; D004262:DNA Restriction Enzymes; D001616:beta-Galactosidase