Complex of an active μ-opioid receptor with a cyclic peptide agonist modeled from experimental constraints

Carol B. Fowler, Irina D. Pogozheva, Andrei L. Lomize, Harry LeVine, Henry I. Mosberg

Research output: Contribution to journalArticlepeer-review

55 Scopus citations


Site-directed mutagenesis and design of Zn2+-binding centers have been used to determine a set of specific tertiary interactions between the μ-opioid receptor, a rhodopsin-like G protein-coupled receptor (GPCR), and its cyclic peptide agonist ligand, Tyr1-c(S-Et-S)[D-Cys 2-Phe3-D-Pen4]NH2 (JOM6). The binding affinity of the tetrapeptide is strongly dependent on the nature of its first and third residues and on substitutions at positions 213, 216, 237, 300, 315, and 318 of the μ-opioid receptor. His1 and His3 analogues of the ligand were able to form metal-binding complexes with the V300C and G213C/T315C receptor mutants, respectively. Direct contact of the Phe 3 residue of JOM6 with Gly213, Asp216, Thr 315, and Trp318 of the receptor was suggested by the binding affinities of His3-, Nle3-, Leu3-, Aci3-, ΔEPhe3-, and Δ ZPhe3-substituted peptides with the G213C/T315C, D216V, T315C, and W318L mutants. The improved binding affinity of the free carboxylate analogue of JOM6 for binding to the E229D mutant revealed an interaction between the C-terminal group of the peptide and Glu229 of the receptor. The experimental constraints that were obtained were applied for distance geometry modeling of the μ-receptor in complex with the tetrapeptide agonist ligand, JOM6. The active conformation of the opioid receptor. was calculated using the crystal structure of "inactive" rhodopsin and published engineered and intrinsic metal-binding sites and disulfide bonds that allow or facilitate activation of GPCRs. Interhelical H-bonds existing in the μ-receptor were applied as additional distance constraints. The calculated model of the receptor-ligand complex can serve as a prototype of the active state for all rhodopsin-like GPCRs. It displays a strongly shifted transmembrane helix 6 (TM6) and reorientation of the conserved Trp293 residue in TM6 upon its interaction with the agonist. Importantly, the binding pockets of the active and inactive states are not identical, which implies distinct interaction modes of agonists and antagonists. In the active state, the binding pocket of the μ-receptor is complementary to the previously proposed receptor-bound conformation of JOM6.

Original languageEnglish
Pages (from-to)15796-15810
Number of pages15
Issue number50
StatePublished - Dec 21 2004

ASJC Scopus subject areas

  • Biochemistry


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