Identification of antigenic epitopes of 1D antigen recognized by antibodies in the serum of patients with thyroid-associated ophthalmopathy.
Random fragments of full-length 1D cDNA encoding the 64-kDa antigen were cloned and expressed in Escherichia coli as fusion proteins. Antigenic peptides were detected by colony blotting with sera from patients with thyroid-associated ophthalmopathy (TAO) and corresponding cDNA inserts were sequenced. Four antigenic peptides were characterized and positioned with respect to the 1D amino acid sequence. One antigenic peptide (G-8) was found to be close to the C-terminal end of 1D, and three others (G-2, G-4, and G-5) were clustered at sites between amino acids 76 and 205. The immunoreactivities of a panel of sera were tested against full-length 1D and the four peptides. A panel consisting of sera from (1) 15 patients with TAO, (2) 9 patients with Graves' hyperthyroidism without ophthalmopathy, (3) 6 patients with Hashimoto's thyroiditis without ophthalmopathy, and (4) 12 normal subjects were tested against the four peptides and full-length 1D protein by Western blotting. Sera from patients with TAO also reacted more often with G-4 and/or G-5 peptides than did those from normal subjects [7 of 15 (47%) sera from TAO patients versus 1 of 12 (8%) sera from normal subjects; P < 0.05]. Based on positioning of peptides onto the full-length 1D sequence and the reaction pattern of the serum panel, six immunogenic epitopes were identified on the 1D molecule.
Zhang
Z G
ZG
Thyroid Studies Center, Montreal General Hospital Research Institute, Quebec, Canada.
Wall
J R
JR
Bernard
N F
NF
D000328:Adult; D000941:Antigens / Q000276:immunology*; D001323:Autoantibodies / Q000097:blood / Q000276:immunology*; D015551:Autoimmunity / Q000276:immunology*; D001483:Base Sequence; D018076:DNA, Complementary / Q000235:genetics / Q000302:isolation & purification; D000939:Epitopes / Q000235:genetics / Q000276:immunology; D005128:Eye Diseases / Q000276:immunology*; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D008969:Molecular Sequence Data; D013959:Thyroid Diseases / Q000276:immunology*
D000941:Antigens; D001323:Autoantibodies; D018076:DNA, Complementary; D000939:Epitopes