|Title||Propagation of conformational changes during μ-opioid receptor activation.|
|Publication Type||Journal Article|
|Year of Publication||2015|
|Authors||Sounier, R., Mas C., Steyaert J., Laeremans T., Manglik A., Huang W., Kobilka B. K., Déméné H., and Granier S.|
|Date Published||2015 Aug 20|
|Keywords||Allosteric Regulation, Animals, Binding Sites, Heterotrimeric GTP-Binding Proteins, Lysine, Mice, Models, Molecular, Morphinans, Nuclear Magnetic Resonance, Biomolecular, Protein Binding, Protein Conformation, Pyrroles, Receptors, Adrenergic, beta-2, Receptors, Opioid, mu, Single-Chain Antibodies, Structure-Activity Relationship, Substrate Specificity|
µ-Opioid receptors (µORs) are G-protein-coupled receptors that are activated by a structurally diverse spectrum of natural and synthetic agonists including endogenous endorphin peptides, morphine and methadone. The recent structures of the μOR in inactive and agonist-induced active states (Huang et al., ref. 2) provide snapshots of the receptor at the beginning and end of a signalling event, but little is known about the dynamic sequence of events that span these two states. Here we use solution-state NMR to examine the process of μOR activation using a purified receptor (mouse sequence) preparation in an amphiphile membrane-like environment. We obtain spectra of the μOR in the absence of ligand, and in the presence of the high-affinity agonist BU72 alone, or with BU72 and a G protein mimetic nanobody. Our results show that conformational changes in transmembrane segments 5 and 6 (TM5 and TM6), which are required for the full engagement of a G protein, are almost completely dependent on the presence of both the agonist and the G protein mimetic nanobody, revealing a weak allosteric coupling between the agonist-binding pocket and the G-protein-coupling interface (TM5 and TM6), similar to that observed for the β2-adrenergic receptor. Unexpectedly, in the presence of agonist alone, we find larger spectral changes involving intracellular loop 1 and helix 8 compared to changes in TM5 and TM6. These results suggest that one or both of these domains may play a role in the initial interaction with the G protein, and that TM5 and TM6 are only engaged later in the process of complex formation. The initial interactions between the G protein and intracellular loop 1 and/or helix 8 may be involved in G-protein coupling specificity, as has been suggested for other family A G-protein-coupled receptors.
|PubMed Central ID||PMC4820006|
|Grant List||R37 DA036246 / DA / NIDA NIH HHS / United States |
T32 GM008294 / GM / NIGMS NIH HHS / United States
DA036246 / DA / NIDA NIH HHS / United States
Propagation of conformational changes during μ-opioid receptor activation.