Sequential binding of agonists to the β2 adrenoceptor. Kinetic evidence for intermediate conformational states

Gayathri Swaminath, Yang Kevin Xiang, Tae Weon Lee, Jacqueline Steenhuis, Charles Parnot, Brian K. Kobilka

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Abstract

The β2 adrenoreceptor (β2AR) is a prototypical G protein-coupled receptor (GPCR) activated by catecholamines. Agonist activation of GPCRs leads to sequential interactions with heterotrimeric G proteins, which activate cellular signaling cascades, and with GPCR kinases and arrestins, which attenuate GPCR-mediated signaling. We used fluorescence spectroscopy to monitor catecholamine-induced conformational changes in purified β 2AR. Here we show that upon catecholamine binding, β 2ARs undergo transitions to two kinetically distinguishable conformational states. Using a panel of chemically related catechol derivatives, we identified the specific chemical groups on the agonist responsible for the rapid and slow conformational changes in the receptor. The conformational changes observed in our biophysical assay were correlated with biologic responses in cellular assays. Dopamine, which induces only a rapid conformational change, is efficient at activating Gs but not receptor internalization. In contrast, norepinephrine and epinephrine, which induce both rapid and slow conformational changes, are efficient at activating Gs and receptor internalization. These results support a mechanistic model for GPCR activation where contacts between the receptor and structural determinants of the agonist stabilize a succession of conformational states with distinct cellular functions.

Original languageEnglish (US)
Pages (from-to)686-691
Number of pages6
JournalJournal of Biological Chemistry
Volume279
Issue number1
DOIs
StatePublished - Jan 2 2004
Externally publishedYes

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ASJC Scopus subject areas

  • Biochemistry

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