Limited proteolysis selectively destroys epitopes on apolipoprotein B in low density lipoproteins.
Apolipoprotein B (apoB), the major apoprotein in human low density lipoprotein (LDL), is a large protein and complex immunogen, against which seven monoclonal IgG antibodies have been produced in our laboratory. These antibodies were used to define at least five individual epitopes on holo-LDL. The aim of the present experiments was to ascertain whether limited proteolysis of LDL would selectively affect the expression of the epitopes. LDL was digested with staphylococcal protease (SP), trypsin (T) or pronase; approximately 20% of LDL protein was lost by treatment with SP and T, and approximately 60% by treatment with pronase. Structurally stable SP- and T-LDL cores and soluble peptides were separated by gel permeation chromatography and zonal ultracentrifugation. The cores resembled holo-LDL in composition and flotation rates but apoB was fragmented into peptides less than 100,000 in molecular weight. On analysis by competitive radioimmunoassays some epitopes of the LDL cores were destroyed partially, some completely, and some were spared. The enzymes each produced individual patterns of epitope modulation. SP- and T-LDL cores retained most of their abilities to be bound and degraded by normal human cultured fibroblasts. Some soluble peptides also manifested both antigenicity and cell reactivity. The retention of activities by the cores while active fragments are lost is compatible with, but does not constitute unequivocal evidence for, the hypothesis that apoB may consist of repeating structures.
D000911:Antibodies, Monoclonal; D001053:Apolipoproteins / Q000276:immunology*; D001055:Apolipoproteins B; D001666:Binding Sites, Antibody; D002478:Cells, Cultured; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes / Q000032:analysis*; D005347:Fibroblasts / Q000276:immunology; D006801:Humans; D007118:Immunoassay; D008077:Lipoproteins, LDL / Q000276:immunology*; D010446:Peptide Fragments / Q000276:immunology*; D013329:Structure-Activity Relationship
D000911:Antibodies, Monoclonal; D001053:Apolipoproteins; D001055:Apolipoproteins B; D000939:Epitopes; D008077:Lipoproteins, LDL; D010446:Peptide Fragments