Mapping of epitopes on the 86 kDa subunit of the Ku autoantigen.
Patients with rheumatic disorders develop autoantibodies to a nuclear protein antigen termed Ku. The Ku antigen has been identified as a non-histone DNA-binding protein complex composed of two polypeptides: 86 kDa and 70 kDa. Initial competition experiments with four monoclonal antibodies specific for the 86 kDa subunit of Ku protein suggested there was more than one epitope on this polypeptide. To determine the region for these epitopes, cDNA deletions were made from both 5' and 3' ends. The immunoreactivity of the expressed proteins was assayed by immunoblotting. A 40 amino acids region located near the C-terminus of the polypeptide between amino acids 667-708 was essential for binding the monoclonal and autoimmune antibodies. Within this region, we differentiated three epitopes for the monoclonal antibodies. The sequence critical for binding of the autoimmune antibodies was in the same region, between amino acids 689-708. None of the four monoclonal antibodies blocked the binding of the autoantibodies. The implications of these results for the etiology of the anti-Ku autoantibody response are discussed.
Wen
J
J
Department of Pharmacology, Baylor College of Medicine, Houston, Texas 77030.
Yaneva
M
M
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal; D034961:Antigens, Nuclear; D000954:Antigens, Surface / Q000737:chemistry / Q000276:immunology*; D001323:Autoantibodies; D001324:Autoantigens / Q000737:chemistry / Q000276:immunology*; D001667:Binding, Competitive; D002872:Chromosome Deletion; D004265:DNA Helicases; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000276:immunology*; D000939:Epitopes / Q000737:chemistry / Q000276:immunology; D004926:Escherichia coli / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D007457:Iodine Radioisotopes; D000072200:Ku Autoantigen; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000276:immunology
D000911:Antibodies, Monoclonal; D034961:Antigens, Nuclear; D000954:Antigens, Surface; D001323:Autoantibodies; D001324:Autoantigens; D004268:DNA-Binding Proteins; D000939:Epitopes; D007457:Iodine Radioisotopes; D011993:Recombinant Fusion Proteins; D004265:DNA Helicases; C106893:XRCC5 protein, human; C000606509:Xrcc6 protein, human; D000072200:Ku Autoantigen