1. Biochemistry and Chemical Biology
  2. Structural Biology and Molecular Biophysics
Download icon

Allosteric modulation in monomers and oligomers of a G protein-coupled receptor

  1. Rabindra V Shivnaraine  Is a corresponding author
  2. Brendan Kelly
  3. Krishana S Sankar
  4. Dar'ya S Redka
  5. Yi Rang Han
  6. Fei Huang
  7. Gwendolynne Elmslie
  8. Daniel Pinto
  9. Yuchong Li
  10. Jonathan V Rocheleau
  11. Claudiu C Gradinaru
  12. John Ellis
  13. James W Wells
  1. Stanford University School of Medicine, United States
  2. Stanford University, United States
  3. University of Toronto, Canada
  4. Hershey Medical Center, United States
Research Article
  • Cited 18
  • Views 2,072
  • Annotations
Cite this article as: eLife 2016;5:e11685 doi: 10.7554/eLife.11685

Abstract

The M2 muscarinic receptor is the prototypic model of allostery in GPCRs, yet the molecular and the supramolecular determinants of such effects are unknown. Monomers and oligomers of the M2 muscarinic receptor therefore have been compared to identify those allosteric properties that are gained in oligomers. Allosteric interactions were monitored by means of a FRET-based sensor of conformation at the allosteric site and in pharmacological assays involving mutants engineered to preclude intramolecular effects. Electrostatic, steric, and conformational determinants of allostery at the atomic level were examined in molecular dynamics simulations. Allosteric effects in monomers were exclusively negative and derived primarily from intramolecular electrostatic repulsion between the allosteric and orthosteric ligands. Allosteric effects in oligomers could be positive or negative, depending upon the allosteric-orthosteric pair, and they arose from interactions within and between the constituent protomers. The complex behavior of oligomers is characteristic of muscarinic receptors in myocardial preparations.

Article and author information

Author details

  1. Rabindra V Shivnaraine

    Department of Molecular and Cellular Physiology, Stanford University School of Medicine, Stanford, United States
    For correspondence
    rvshiv@stanford.edu
    Competing interests
    The authors declare that no competing interests exist.
  2. Brendan Kelly

    Department of Computer Science, Stanford University, Stanford, United States
    Competing interests
    The authors declare that no competing interests exist.
  3. Krishana S Sankar

    Department of Physiology, University of Toronto, Toronto, Canada
    Competing interests
    The authors declare that no competing interests exist.
  4. Dar'ya S Redka

    Department of Pharmaceutical Sciences, Leslie Dan Faculty of Pharmacy, University of Toronto, Toronto, Canada
    Competing interests
    The authors declare that no competing interests exist.
  5. Yi Rang Han

    Department of Pharmaceutical Sciences, Leslie Dan Faculty of Pharmacy, University of Toronto, Toronto, Canada
    Competing interests
    The authors declare that no competing interests exist.
  6. Fei Huang

    Department of Pharmaceutical Sciences, Leslie Dan Faculty of Pharmacy, University of Toronto, Toronto, Canada
    Competing interests
    The authors declare that no competing interests exist.
  7. Gwendolynne Elmslie

    Departments of Psychiatry and Pharmacology, Hershey Medical Center, Hershey, United States
    Competing interests
    The authors declare that no competing interests exist.
  8. Daniel Pinto

    Department of Pharmaceutical Sciences, Leslie Dan Faculty of Pharmacy, University of Toronto, Toronto, Canada
    Competing interests
    The authors declare that no competing interests exist.
  9. Yuchong Li

    Department of Physics, University of Toronto, Toronto, Canada
    Competing interests
    The authors declare that no competing interests exist.
  10. Jonathan V Rocheleau

    Department of Physiology, University of Toronto, Toronto, Canada
    Competing interests
    The authors declare that no competing interests exist.
  11. Claudiu C Gradinaru

    Department of Physics, University of Toronto, Toronto, Canada
    Competing interests
    The authors declare that no competing interests exist.
  12. John Ellis

    Departments of Psychiatry and Pharmacology, Hershey Medical Center, Hershey, United States
    Competing interests
    The authors declare that no competing interests exist.
  13. James W Wells

    Department of Pharmaceutical Sciences, Leslie Dan Faculty of Pharmacy, University of Toronto, Toronto, Canada
    Competing interests
    The authors declare that no competing interests exist.

Reviewing Editor

  1. Werner Kühlbrandt, Max Planck Institute of Biophysics, Germany

Publication history

  1. Received: September 17, 2015
  2. Accepted: April 30, 2016
  3. Accepted Manuscript published: May 6, 2016 (version 1)
  4. Version of Record published: June 9, 2016 (version 2)

Copyright

© 2016, Shivnaraine et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

Metrics

  • 2,072
    Page views
  • 570
    Downloads
  • 18
    Citations

Article citation count generated by polling the highest count across the following sources: Scopus, Crossref, PubMed Central.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Download citations (links to download the citations from this article in formats compatible with various reference manager tools)

Open citations (links to open the citations from this article in various online reference manager services)

Further reading

    1. Biochemistry and Chemical Biology
    Erica N Thomas et al.
    Research Article

    Similar to DNA replication, translation of the genetic code by the ribosome is hypothesized to be exceptionally sensitive to small chemical changes to its template mRNA. Here we show that addition of common alkylating agents to growing cultures of E. coli leads to accumulation of several adducts within RNA, including N(1)-methyladenosine (m1A). As expected, the introduction of m1A to model mRNAs was found to reduce the rate of peptide-bond formation by three orders of magnitude in a well-defined in vitro system. These observations suggest that alkylative stress is likely to stall translation in vivo and necessitates activation of ribosome-rescue pathways. Indeed, the addition of alkylation agents was found to robustly activate the transfer-messenger RNA system, even when transcription was inhibited. Our findings suggest that bacteria carefully monitor the chemical integrity of their mRNA and they evolved rescue pathways to cope with its effect on translation.

    1. Biochemistry and Chemical Biology
    2. Cell Biology
    Mikel Garcia-Marcos et al.
    Tools and Resources

    Heterotrimeric G-proteins are signal transducers involved in mediating the action of many natural extracellular stimuli as well as of many therapeutic agents. Non-invasive approaches to manipulate the activity of G-proteins with high precision are crucial to understand their regulation in space and time. Here, we developed LOV2GIVe, an engineered modular protein that allows the activation of heterotrimeric G-proteins with blue light. This optogenetic construct relies on a versatile design that differs from tools previously developed for similar purposes, i.e. metazoan opsins, which are light-activated GPCRs. Instead, LOV2GIVe consists of the fusion of a G-protein activating peptide derived from a non-GPCR regulator of G-proteins to a small plant protein domain, such that light uncages the G-protein activating module. Targeting LOV2GIVe to cell membranes allowed for light-dependent activation of Gi proteins in different experimental systems. In summary, LOV2GIVe expands the armamentarium and versatility of tools available to manipulate heterotrimeric G-protein activity.