Ligand discrimination and gating in cyclic nucleotide-gated ion channels from apo and partial agonist-bound cryo-EM structures
- Weill Cornell Medical College, United States
Abstract
Cyclic nucleotide-modulated channels have important roles in visual signal transduction and pacemaking. Binding of cyclic nucleotides (cAMP/cGMP) elicits diverse functional responses in different channels within the family despite their high sequence and structure homology. The molecular mechanisms responsible for ligand discrimination and gating are unknown due to lack of correspondence between structural information and functional states. Using single particle cryo-electron microscopy and single-channel recording, we assigned functional states to high-resolution structures of SthK, a prokaryotic cyclic nucleotide-gated channel. The structures for apo, cAMP-bound, and cGMP-bound SthK in lipid nanodiscs, correspond to no, moderate, and low single-channel activity, respectively, consistent with the observation that all structures are in resting, closed states. The similarity between apo and ligand-bound structures indicates that ligand-binding domains are strongly coupled to pore and SthK gates in an allosteric, concerted fashion. The different orientations of cAMP and cGMP in the 'resting' and 'activated' structures suggest a mechanism for ligand discrimination.
Data availability
The 3 density maps and 3 atomic models have been deposited in PDB under the following accession codes: 6CJQ, 6CJU and 6CJT (coordinates of atomic models), EMD-7482, EMD-7484 and EMD-7483 (density maps).
-
apo-bound structure of the prokaryotic SthK channel (coordinates of atomic model)Publicly available at the Electron Microscopy Data Bank (accession no: 6CJQ).
-
cAMP-bound structure of the prokaryotic SthK channel (coordinates of atomic model)Publicly available at the Electron Microscopy Data Bank (accession no: 6CJU).
-
cGMP-bound structure of the prokaryotic SthK channel (coordinates of atomic model)Publicly available at the Electron Microscopy Data Bank (accession no: 6CJT).
-
apo-bound structure of the prokaryotic SthK channel (density map)Publicly available at the Electron Microscopy Data Bank (accession no: EMD-7482).
-
cAMP-bound structure of the prokaryotic SthK channel (density map)Publicly available at the Electron Microscopy Data Bank (accession no: EMD-7484).
-
cGMP-bound structure of the prokaryotic SthK channel (density map)Publicly available at the Electron Microscopy Data Bank (accession no: EMD-7483).
Article and author information
Author details
Philipp AM Schmidpeter
Funding
National Institute of General Medical Sciences (R01GM124451 and R01GM088352)
- Crina M Nimigean
Deutsche Forschungsgemeinschaft (SCHM 3198/1-1)
- Philipp AM Schmidpeter
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2018, Rheinberger 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
-
- 3,545
- views
-
- 658
- downloads
-
- 44
- citations
Views, downloads and citations are aggregated across all versions of this paper published by eLife.
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)
Open citations (links to open the citations from this article in various online reference manager services)
Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)
Ligand discrimination and gating in cyclic nucleotide-gated ion channels from apo and partial agonist-bound cryo-EM structures
eLife 7:e39775.
https://doi.org/10.7554/eLife.39775
Further reading
-
- Computational and Systems Biology
- Structural Biology and Molecular Biophysics
Viral adhesion to host cells is a critical step in infection for many viruses, including monkeypox virus (MPXV). In MPXV, the H3 protein mediates viral adhesion through its interaction with heparan sulfate (HS), yet the structural details of this interaction have remained elusive. Using AI-based structural prediction tools and molecular dynamics (MD) simulations, we identified a novel, positively charged α-helical domain in H3 that is essential for HS binding. This conserved domain, found across orthopoxviruses, was experimentally validated and shown to be critical for viral adhesion, making it an ideal target for antiviral drug development. Targeting this domain, we designed a protein inhibitor, which disrupted the H3-HS interaction, inhibited viral infection in vitro and viral replication in vivo, offering a promising antiviral candidate. Our findings reveal a novel therapeutic target of MPXV, demonstrating the potential of combination of AI-driven methods and MD simulations to accelerate antiviral drug discovery.
-
- Chromosomes and Gene Expression
- Structural Biology and Molecular Biophysics
Type II nuclear receptors (T2NRs) require heterodimerization with a common partner, the retinoid X receptor (RXR), to bind cognate DNA recognition sites in chromatin. Based on previous biochemical and overexpression studies, binding of T2NRs to chromatin is proposed to be regulated by competition for a limiting pool of the core RXR subunit. However, this mechanism has not yet been tested for endogenous proteins in live cells. Using single-molecule tracking (SMT) and proximity-assisted photoactivation (PAPA), we monitored interactions between endogenously tagged RXR and retinoic acid receptor (RAR) in live cells. Unexpectedly, we find that higher expression of RAR, but not RXR, increases heterodimerization and chromatin binding in U2OS cells. This surprising finding indicates the limiting factor is not RXR but likely its cadre of obligate dimer binding partners. SMT and PAPA thus provide a direct way to probe which components are functionally limiting within a complex TF interaction network providing new insights into mechanisms of gene regulation in vivo with implications for drug development targeting nuclear receptors.
