Prokaryotic expression of the thyrotropin receptor and identification of an immunogenic region of the protein using synthetic peptides.
Graves' disease is characterized by hypersecretion of thyroid hormones due to binding of autoantibodies to the thyrotropin receptor (TSHR). In order to study immunological aspects of the TSHR we expressed the extracellular domain of the rat TSHR (ETSHR) as a fusion protein with beta-galactosidase in a prokaryotic system. The identity of this ETSHR-fusion protein was confirmed by Western blot, using antibodies to synthetic peptides derived from TSHR. Patients' sera reacted to a significantly greater extent with the affinity purified ETSHR relative to control sera. Similarly, sera from patients with Graves' disease displayed significant reactivity with only one of five peptides, RH2 (residues 352-366), when compared with normal sera. These data, together with the predicted hydrophilicity of the peptide RH2, suggest that amino acids 352-366 which lie within one of the unique regions of the extracellular domain of the TSHR may be important for antibody binding.
Takai
O
O
Laboratory of Oral Medicine, NIDR National Institutes of Health, Bethesda, MD 20892.
Desai
R K
RK
Seetharamaiah
G S
GS
Jones
C A
CA
Allaway
G P
GP
Akamizu
T
T
Kohn
L D
LD
Prabhakar
B S
BS
D000595:Amino Acid Sequence; D000818:Animals; D000936:Antigen-Antibody Complex; D001323:Autoantibodies / Q000378:metabolism*; D003001:Cloning, Molecular; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli / Q000235:genetics; D006111:Graves Disease / Q000276:immunology*; D006801:Humans; D008969:Molecular Sequence Data; D010455:Peptides / Q000138:chemical synthesis* / Q000276:immunology; D010957:Plasmids; D051381:Rats; D011989:Receptors, Thyrotropin / Q000235:genetics* / Q000276:immunology; D011993:Recombinant Fusion Proteins / Q000276:immunology; D012016:Reference Values; D015183:Restriction Mapping
D000936:Antigen-Antibody Complex; D001323:Autoantibodies; D010455:Peptides; D011989:Receptors, Thyrotropin; D011993:Recombinant Fusion Proteins