title: Helix-constrained nociceptin peptides are potent agonists and antagonists of ORL-1 and nociception.
abstract: Nociceptin (orphanin FQ) is a 17-residue neuropeptide hormone with roles in both nociception and analgesia. It is an opioid-like peptide that binds to and activates the G-protein-coupled receptor opioid receptor-like-1 (ORL-1, NOP, orphanin FQ receptor, kappa-type 3 opioid receptor) on central and peripheral nervous tissue, without activating classic delta-, kappa-, or mu-opioid receptors or being inhibited by the classic opioid antagonist naloxone. The three-dimensional structure of ORL-1 was recently published, and the activation mechanism is believed to involve capture by ORL-1 of the high-affinity binding, prohelical C-terminus. This likely anchors the receptor-activating N-terminus of nociception nearby for insertion in the membrane-spanning helices of ORL-1. In search of higher agonist potency, two lysine and two aspartate residues were strategically incorporated into the receptor-binding C-terminus of the nociceptin sequence and two Lys(i)→Asp(i+4) side chain-side chain condensations were used to generate lactam cross-links that constrained nociceptin into a highly stable α-helix in water. A cell-based assay was developed using natively expressed ORL-1 receptors on mouse neuroblastoma cells to measure phosphorylated ERK as a reporter of agonist-induced receptor activation and intracellular signaling. Agonist activity was increased up to 20-fold over native nociceptin using a combination of this helix-inducing strategy and other amino acid modifications. An NMR-derived three-dimensional solution structure is described for a potent ORL-1 agonist derived from nociceptin, along with structure-activity relationships leading to the most potent known α-helical ORL-1 agonist (EC₅₀ 40 pM, pERK, Neuro-2a cells) and antagonist (IC₅₀ 7 nM, pERK, Neuro-2a cells). These α-helix-constrained mimetics of nociceptin(1-17) had enhanced serum stability relative to unconstrained peptide analogues and nociceptin itself, were not cytotoxic, and displayed potent thermal analgesic and antianalgesic properties in rats (ED₅₀ 70 pmol, IC₅₀ 10 nmol, s.c.), suggesting promising uses in vivo for the treatment of pain and other ORL-1-mediated responses.
authors: Institute for Molecular Bioscience, University of Queensland, Brisbane, Queensland, Australia.
authors: Rink-Jan
authors: RJ
authors: Lohman
authors: Institute for Molecular Bioscience, University of Queensland, Brisbane, Queensland, Australia.
authors: Rosemary S
authors: RS
authors: Harrison
authors: Institute for Molecular Bioscience, University of Queensland, Brisbane, Queensland, Australia.
authors: Gloria
authors: G
authors: Ruiz-Gómez
authors: Institute for Molecular Bioscience, University of Queensland, Brisbane, Queensland, Australia.
authors: Huy N
authors: HN
authors: Hoang
authors: Institute for Molecular Bioscience, University of Queensland, Brisbane, Queensland, Australia.
authors: Nicholas E
authors: NE
authors: Shepherd
authors: Institute for Molecular Bioscience, University of Queensland, Brisbane, Queensland, Australia.
authors: Shiao
authors: S
authors: Chow
authors: Institute for Molecular Bioscience, University of Queensland, Brisbane, Queensland, Australia.
authors: Timothy A
authors: TA
authors: Hill
authors: Institute for Molecular Bioscience, University of Queensland, Brisbane, Queensland, Australia.
authors: Praveen K
authors: PK
authors: Madala
authors: Institute for Molecular Bioscience, University of Queensland, Brisbane, Queensland, Australia. Electronic address: d.fairlie@imb.uq.edu.au.
authors: David P
authors: DP
authors: Fairlie
mesh_terms: D000701:Analgesics, Opioid / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*; D000818:Animals; D015195:Drug Design; D015507:Drugs, Investigational / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D006801:Humans; D009292:Narcotic Antagonists / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*; D009419:Nerve Tissue Proteins / Q000819:agonists / Q000037:antagonists & inhibitors / Q000378:metabolism*; D009474:Neurons / Q000187:drug effects* / Q000378:metabolism; D059225:Nociception / Q000187:drug effects*; D009842:Oligopeptides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D018847:Opioid Peptides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D010455:Peptides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D011487:Protein Conformation; D015202:Protein Engineering; D011957:Receptors, Opioid / Q000819:agonists / Q000737:chemistry / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D000094942:Nociceptin Receptor; D000097629:Nociceptin
chemical_list: D000701:Analgesics, Opioid; D015507:Drugs, Investigational; D009292:Narcotic Antagonists; D009419:Nerve Tissue Proteins; D009842:Oligopeptides; D018847:Opioid Peptides; D010455:Peptides; D011957:Receptors, Opioid; D011994:Recombinant Proteins; D000094942:Nociceptin Receptor; D000097629:Nociceptin; C000722283:OPRL1 protein, human
keywords: Agonist; Analgesia; Helix; Nociceptin; Nociception; ORL