Journal of Clinical Endocrinology & Metabolism, Vol 81, 700-706, Copyright © 1996 by Endocrine Society

ARTICLES

Binding of antithyrotropin receptor autoantibodies in Graves' disease serum to nascent, in vitro translated thyrotropin receptor: ability to map epitopes recognized by antibodies

NG Morgenthaler, J Tremble, G Huang, WA Scherbaum, AM McGregor and JP Banga
Department of Medicine, King's College School of Medicine, London, United Kingdom.

The binding of Graves' disease autoantibodies and xenogeneic antibodies to the human TSH receptor (TSH-R) has been studied using receptor preparations generated in an in vitro transcription and translation reaction. The complementary DNAs encoding for the full-length (764 amino acids) and the extracellular region of TSH-R (amino acids 20-414, lacking the signal sequence) were used to generate the translated receptor antigen. Stable [35S]methionine-labeled nascent protein for full-length and extracellular regions of TSH-R of approximate size 87 and 50 kDa, respectively, together with other smaller proteins were generated. The 87- and 50-kDa translated receptor proteins react by immunoprecipitation analysis with monoclonal antibodies, polyclonal antisera, and unfractionated Graves' disease serum containing autoantibody to TSH-R. The translated products of the extracellular TSH- R were examined in detail. Using three well characterized murine monoclonal antibodies whose epitopes encompass the amino-, central, and carboxyl-terminals of the extracellular region of the receptor led to immunochemical identification of the smaller translated products to derive from internal methionine start sites of TSH-R. These smaller, N- terminal-truncated translated proteins were also recognized by polyclonal antisera generated against recombinant TSH-R, thus allowing epitope mapping of some antibodies. A large proportion of Graves' disease autoantibodies (> 70%) bind to the translated extracellular region of TSH-R. This indicates that the majority of pathogenic anti- TSH-R autoantibody binds the nascent translated extracellular region of TSH-R, which is not influenced by the lack of glycosylation of the receptor.

This article has been cited by other articles:

				J. Seissler, S. Wagner, M. Schott, M. Lettmann, J. Feldkamp, W. A. Scherbaum, and N. G. Morgenthaler Low Frequency of Autoantibodies to the Human Na+/I- Symporter in Patients with Autoimmune Thyroid Disease J. Clin. Endocrinol. Metab., December 1, 2000; 85(12): 4630 - 4634. [Abstract] [Full Text]

				R. A. Ajjan, E. H. Kemp, E. A. Waterman, P. F. Watson, T. Endo, T. Onaya, and A. P. Weetman Detection of Binding and Blocking Autoantibodies to the Human Sodium-Iodide Symporter in Patients with Autoimmune Thyroid Disease J. Clin. Endocrinol. Metab., May 1, 2000; 85(5): 2020 - 2027. [Abstract] [Full Text]

				Y. Oda, J. Sanders, S. Roberts, M. Maruyama, A. Kiddie, J. Furmaniak, and B. Rees Smith Analysis of Carbohydrate Residues on Recombinant Human Thyrotropin Receptor J. Clin. Endocrinol. Metab., June 1, 1999; 84(6): 2119 - 2125. [Abstract] [Full Text]

				S. Costagliola, N. G. Morgenthaler, R. Hoermann, K. Badenhoop, J. Struck, D. Freitag, S. Poertl, W. Weglöhner, J. M. Hollidt, B. Quadbeck, et al. Second Generation Assay for Thyrotropin Receptor Antibodies Has Superior Diagnostic Sensitivity for Graves' Disease J. Clin. Endocrinol. Metab., January 1, 1999; 84(1): 90 - 97. [Abstract] [Full Text]

				B. Rapoport, G. D. Chazenbalk, J. C. Jaume, and S. M. McLachlan The Thyrotropin (TSH)-Releasing Hormone Receptor: Interaction with TSH and Autoantibodies Endocr. Rev., December 1, 1998; 19(6): 673 - 716. [Abstract] [Full Text]

				G. D. Chazenbalk, J. C. Jaume, S. M. McLachlan, and B. Rapoport Engineering the Human Thyrotropin Receptor Ectodomain from a Non-secreted Form to a Secreted, Highly Immunoreactive Glycoprotein That Neutralizes Autoantibodies in Graves' Patients' Sera J. Biol. Chem., July 25, 1997; 272(30): 18959 - 18965. [Abstract] [Full Text] [PDF]

				A. Kakinuma, G. D. Chazenbalk, J. C. Jaume, B. Rapoport, and S. M. McLachlan The Human Thyrotropin (TSH) Receptor in a TSH Binding Inhibition Assay for TSH Receptor Autoantibodies J. Clin. Endocrinol. Metab., July 1, 1997; 82(7): 2129 - 2134. [Abstract] [Full Text] [PDF]

				L. Prentice, J. F. Sanders, M. Perez, R. Kato, J. Sawicka, Y. Oda, D. Jaskolski, J. Furmaniak, and B. R. Smith Thyrotropin (TSH) Receptor Autoantibodies Do Not Appear to Bind to the TSH Receptor Produced in an in Vitro Transcription/Translation System J. Clin. Endocrinol. Metab., April 1, 1997; 82(4): 1288 - 1292. [Abstract] [Full Text] [PDF]

				J. C. Jaume, A. Kakinuma, G. D. Chazenbalk, B. Rapoport, and S. M. McLachlan Thyrotropin Receptor Autoantibodies in Serum Are Present at Much Lower Levels Than Thyroid Peroxidase Autoantibodies: Analysis by Flow Cytometry J. Clin. Endocrinol. Metab., February 1, 1997; 82(2): 500 - 507. [Abstract] [Full Text] [PDF]