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Article

  • The EMBO Journal (1997) 16, 6737 - 6747
  • doi:10.1093/emboj/16.22.6737

Agonists induce conformational changes in transmembrane domains III and VI of the bold beta2 adrenoceptor

Ulrik Gether1,4, Sansan Lin1, Pejman Ghanouni1, Juan A. Ballesteros3, Harel Weinstein3 and Brian K. Kobilka1,2

  1. Howard Hughes Medical Institute, Stanford University Medical School, Stanford, CA 94305 USA
  2. Division of Cardiovascular Medicine, Stanford University Medical School, Stanford, CA 94305 USA
  3. Department of Physiology and Biophysics, Mount Sinai School of Medicine, New York, NY 10029, USA
  4. Present address: Department of Medical Physiology, The Panum Institute, University of Copenhagen, DK-2200 Copenhagen, Denmark

Correspondence to:

Ulrik Gether, E-mail: gether@mfi.ku.dk

Brian K. Kobilka, E-mail: kobilka@cmgm.stanford.edu

Received 18 April 1997; Revised 28 July 1997

Agonist binding to G protein-coupled receptors is believed to promote a conformational change that leads to the formation of the active receptor state. However, the character of this conformational change which provides the important link between agonist binding and G protein coupling is not known. Here we report evidence that agonist binding to the beta2 adrenoceptor induces a conformational change around 125Cys in transmembrane domain (TM) III and around 285Cys in TM VI. A series of mutant beta2 adrenoceptors with a limited number of cysteines available for chemical derivatization were purified, site-selectively labeled with the conformationally sensitive, cysteine-reactive fluorophore IANBD and analyzed by fluorescence spectroscopy. Like the wild-type receptor, mutant receptors containing 125Cys and/or 285Cys showed an agonist-induced decrease in fluorescence, while no agonist-induced response was observed in a receptor where these two cysteines were mutated. These data suggest that IANBD bound to 125Cys and 285Cys are exposed to a more polar environment upon agonist binding, and indicate that movements of transmembrane segments III and VI are involved in activation of G protein-coupled receptors.

  • Keywords:

    • fluorescence spectroscopy,
    • G protein-coupled receptors,
    • molecular modeling,
    • signal transduction