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Principal Investigator: Brian K Kobilka
Organization: STANFORD UNIVERSITY
Fiscal Year: 2024
Award: $549,378
Funding agency: National Institute on Drug Abuse
Project Summary/Abstract
The structural basis for pathway-selective signaling by the µ-opioid receptor.
For many patients experiencing acute and chronic pain, opioids like codeine, morphine, and fentanyl have
improved their quality of life. Unfortunately, these benefits can be offset by dose-limiting liabilities, like addiction
and the respiratory depression responsible for opioid overdose deaths. The µ-opioid receptor (µOR) is
responsible for mediating the beneficial and adverse effects of most opioid analgesics. The µOR has complex
signaling behavior, activating six different G proteins subtypes (Gi1, Gi2, Gi3, GoA, GoB, Gz) and two arrestin
subtypes. Most of the opioid agonists currently used to treat pain activate all of these signaling pathways. Yet
there is a growing body of evidence that only a subset of these signaling pathways mediate analgesia, while a
different subset may be responsible for the adverse effects. Moreover, it is possible that the Gi/o/z subtypes
responsible for alleviating acute pain are different from the subtypes responsible for alleviating chronic pain.
We have identified several agonists that preferentially activate subsets of Gi/o/z proteins that retain analgesic
efficacy but have fewer adverse effects. The overall goal of the proposal is to determine the structural basis for
this pathway-selective G protein signaling. This information will facilitate the development of more pathway-
selective agonists. These agonists will provide useful tools for understanding the complex signaling behavior of
the µOR and the role of specific pathways in mediating the therapeutic and adverse effects of opioid agonists.
The Specific Aims of the overall proposal are:
Aim 1A. Determine the structural basis for subtype-selective signaling to Gi/o/z proteins. We will use cryo-
electron microscopy to determine structures of the µOR bound to different Gi/o/z subtypes and different
pathway selective agonists.
Aim1B. Structure-guided synthesis of novel pathway-selective µOR agonists.
Aim 2. Characterize the steady-state conformational changes in the cytoplasmic surface of the µOR stabilized
by agonists with distinct signaling profiles. We will use double electron-electron resonance (DEER)
spectroscopy to map conformational changes in the cytoplasmic surface of the µOR bound to pathway
selective and non-selective agonists.
Aim 3. Characterize the dynamics of TM6, ICL2, TM7, and the C-terminus in response to the binding of
agonists with distinct signaling profiles in the presence and absence of specific G proteins and arrestins. We
will use single-molecule Förster Resonance Energy Transfer to monitor conformational changes of these
cytoplasmic domains in real-time.
Terms: <Absence of pain sensation><Absence of sensibility to pain><Actiq><Acute Pain><Acute pain management><Adverse effects><Agonist><Analgesic Agents><Analgesic Drugs><Analgesic Preparation><Analgesics><Anodynes><Antinociceptive Agents><Antinociceptive Drugs><Arrestins><Behavior><Binding><Cause of Death><Cell Communication and Signaling><Cell Signaling><Clinical><Codeine><Complex><Coupled><Coupling><Cryo-electron Microscopy><Cryoelectron Microscopy><Cytoplasm><Cytoplasmic Domain><Cytoplasmic Tail><Development><Dose Limiting><Drugs><Duragesic><Electron Cryomicroscopy><Electrons><Europe><Exhibits><FRET><Feels no pain><Fentanest><Fentanyl><Fentyl><Fluorescence Resonance Energy Transfer><Förster Resonance Energy Transfer><G-Proteins><GTP-Binding Proteins><GTP-Regulatory Proteins><Goals><Guanine Nucleotide Coupling Protein><Guanine Nucleotide Regulatory Proteins><Infumorph><Intracellular Communication and Signaling><Isoforms><Kadian><MS Contin><MSir><Maps><Mediating><Medication><Medicine><Mice><Mice Mammals><Molecular Configuration><Molecular Conformation><Molecular Interaction><Molecular Stereochemistry><Monitor><Morphia><Morphine><Murine><Mus><N-Methylmorphine><Negative Beta Particle><Negatrons><No sensitivity to pain><North America><Opiate Receptors><Opiate agonist><Opiate receptor agonist><Opiates><Opioid><Opioid Analgesics><Opioid Receptor><Opioid Receptor Binding><Opioid agonist><Opioid receptor agonist><Oramorph><Oramorph SR><Pain><Painful><Pathway interactions><Patients><Pharmaceutical Preparations><Phentanyl><Property><Protein Isoforms><QOL><Quality of life><Receptor Protein><Respiratory Depression><Retinal S-Antigen><Role><Roxanol><Signal Pathway><Signal Transduction><Signal Transduction Systems><Signaling><Signaling Factor Proto-Oncogene><Signaling Pathway Gene><Signaling Protein><Spectroscopy><Spectrum Analyses><Spectrum Analysis><Statex SR><Structure><Structure-Activity Relationship><Surface><Therapeutic><Therapeutic Effect><Time><Ventilatory Depression><addiction><addictive disorder><analgesia><analog><arr3><arrestin 2><arrestin 3><arrestin3><biological signal transduction><biophysical approaches><biophysical methodology><biophysical methods><biophysical techniques><chemical structure function><chronic pain><conformation><conformational><conformational state><conformationally><conformations><cryo-EM><cryoEM><cryogenic electron microscopy><depressed breathing><depression of breathing><developmental><drug action><drug/agent><experience><improved><insight><mitragynine pseudoindoxyl><novel><opiate analgesia><opiate analgesic><opiate consumption><opiate crisis><opiate deaths><opiate drug use><opiate intake><opiate mortality><opiate pain medication><opiate pain reliever><opiate use><opioid analgesia><opioid anesthetic><opioid consumption><opioid crisis><opioid deaths><opioid drug use><opioid epidemic><opioid intake><opioid mortality><opioid overdose death><opioid pain medication><opioid pain reliever><opioid painkiller><opioid related death><opioid use><overdose death><overdose fatalities><pain killer><pain medication><pain reliever><painkiller><particle><pathway><plasma protein Z><protein Z><receptor><respiratory><response><single molecule><social role><structure function relationship><therapeutic target><therapeutically effective><tool>