Affinity Labelling of the Thyrotropin Receptor

  • Paul R. Auckland
  • Fionuala Creagh
  • Bernard Rees Smith
Part of the Methodological Surveys in Biochemistry and Analysis book series (MSBA, volume 13)


Thyrotropin (TSH) receptors from different species and tissues have been covalently cros s-linked to 125I-labelled TSH, and the mol. wts. of the TSH-TSH receptor complexes elucidated. Two successful methods of cross-linking entail use of the reagents disuccinimidyl suberate (DSS) and N-hydroxysuccin-imidyl-4-azidobenzoate (HSAB). Highly purified bovine TSH (70 u./mg) was coupled to HSAB (which is photoreactive), labelled wi th 125I, then incubated with several different TSH receptor preparations and coupled to the receptor by photolysis. Alternatively, the TSH was labelled with 125I and incubated with solubilized receptors; DSS effected cross-linking.

The TSH-TSH receptor complexes were heated in sodium do-decyl sulphate (SDS) solution and analyzed by SDS-polyacrylamide gel electrophoresis (PAGE) with and without reduction by mercaptoethanol; the dried gel was autoradiographed. Porcine, human and guinea pig thyroid TSH receptors and guinea pig fat-cell TSH receptors were all found to contain two subunits linked by one or more disulphide bridges, of mol. wt. x 10–3: por-cine, 45 & 27; human, 50 & 32; guinea pig thyroid, 60 & 31, and fat, 57 & 32. Further information was gained by examining the effects on the cross-linking of chemical reagents and trypsin.


Receptor Subunit Disulphide Bridge Porcine Thyroid Disuccinimidyl Suberate Decyl Sulphate 
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  1. 1.
    Rees Smith, B. (1981) Receptors and Recognition, Series B, 13, 217–244.Google Scholar
  2. 2.
    Rees Smith, B. & Buckland, P.R. (1982) in Receptors, Antibodies and Disease [Ciba Found. Symp. 90] (Evered, D. & Whelan, J., eds.), Pitman, London, pp. 114–132.Google Scholar
  3. 3.
    Brennan, A., Petersen, V.B., Petersen, M.M., Rees Smith, B. & Hall, R. (1980) FEBS Lett. 111, 35–38.CrossRefGoogle Scholar
  4. 4.
    Rickards, C.R., Buckland, P.R., Rees Smith, B. & Hall, R. (1981) FEBS Lett. 127, 17–21.CrossRefGoogle Scholar
  5. 5.
    Rees Smith, B., McLachlan, S.M., Ginsberg, J., Rickards, C. & Hall, R. (1981) in Hormones in Normal and Abnormal Human Tissues, Vol. 11 (Fotherby, K. & Pal, S.B., eds.), Walter de Gruyter, Berlin, pp. 519–546.Google Scholar
  6. 6.
    Ginsberg, J., Rees Smith, B. & Hall, R. (1982) Mol. Cell. Endocrin.26, 95–102.CrossRefGoogle Scholar
  7. 7.
    Buckland, P.R., Rickards, C.R., Howells, R.D., Jones, E.D. & Rees Smith, B. (1982) FEBS Lett. 145, 245–249.CrossRefGoogle Scholar
  8. 8.
    Buckland, P.R., Rickards, C.R., Howells, R.D., Jones, E.D. & Rees Smith, B. (1983) Biochem. Soc. Trans.11, 189–190.Google Scholar
  9. 9.
    Ji, T.H. (1979) Biochim. Biophys. Acta559, 39–69.CrossRefGoogle Scholar
  10. 10.
    Bolton, A.E. & Hunter, W.M. (1973) Biochem. J.133, 529–539.Google Scholar
  11. 11.
    Pierce, J.G., Liao, T.H., Howard, S.M., Shome, B. & Cornell, J.S. (1971) Rec. Prog. Horm. Res.27, 165–212.Google Scholar
  12. 12.
    Rees Smith, B. & Hall, R. (1981) Meths. Enzymol. 74Pt.C, 405–420.CrossRefGoogle Scholar
  13. 13.
    Davies, T.F., Teng, C.S., McLachlan, S.M., Rees Smith, B. & Hall, R. (1978) Mol. Cell Endocrin.9, 303–310.CrossRefGoogle Scholar
  14. 14.
    Fayet, G., Pacheco, M. & Tixier, R. (1970) Bull. Soc. Chim. Biol.52, 299–306.Google Scholar
  15. 15.
    Swanstrom, R. & Shank, P.R. (1978) Anal. Biochem.86, 184–192.CrossRefGoogle Scholar
  16. 16.
    Griffith, I.P. (1972) Biochem. J.126, 553–560.Google Scholar
  17. 17.
    Kohn, L.D., Valente, W.A., Laccetti, P., Cohen, J.L., Aloj, S.M. & Grollman, E.F. (1983) Life Sci. 32, 15–30.CrossRefGoogle Scholar

Copyright information

© Plenum Press, New York 1984

Authors and Affiliations

  • Paul R. Auckland
    • 1
  • Fionuala Creagh
    • 1
  • Bernard Rees Smith
    • 1
  1. 1.Endocrine Immunology Unit, Department of MedicineWelsh National School of MedicineCardiffUK

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