1. Structural Biology and Molecular Biophysics

Structure of the bacterial ribosome at 2 Å resolution

  1. Zoe L Watson
  2. Fred R Ward
  3. Raphaël Méheust
  4. Omer Ad
  5. Alanna Schepartz
  6. Jillian F Banfield
  7. Jamie HD Cate  Is a corresponding author
  1. University of California, Berkeley, United States
https://doi.org/10.7554/eLife.60482
Share this article
Cite this article
  1. Zoe L Watson
  2. Fred R Ward
  3. Raphaël Méheust
  4. Omer Ad
  5. Alanna Schepartz
  6. Jillian F Banfield
  7. Jamie HD Cate
(2020)
Structure of the bacterial ribosome at 2 Å resolution
eLife 9:e60482.
https://doi.org/10.7554/eLife.60482
  2. Download BibTeX
  3. Download .RIS

Abstract

Using cryo-electron microscopy (cryo-EM), we determined the structure of the Escherichia coli 70S ribosome with a global resolution of 2.0 Å. The maps reveal unambiguous positioning of protein and RNA residues, their detailed chemical interactions, and chemical modifications. Notable features include the first examples of isopeptide and thioamide backbone substitutions in ribosomal proteins, the former likely conserved in all domains of life. The maps also reveal extensive solvation of the small (30S) ribosomal subunit, and interactions with A-site and P-site tRNAs, mRNA, and the antibiotic paromomycin. The maps and models of the bacterial ribosome presented here now allow a deeper phylogenetic analysis of ribosomal components including structural conservation to the level of solvation. The high quality of the maps should enable future structural analysis of the chemical basis for translation and aid the development of robust tools for cryo-EM structure modeling and refinement.

Data availability

Ribosome coordinates have been deposited in the Protein Data Bank (entry 7K00), maps in the EM Database (entries EMD-22586, EMD-22607, EMD-22614, EMD-22632, EMD-22635, EMD-22636, and EMD-22637 for the 70S ribosome composite map, 70S ribosome, 50S subunit, 30S subunit, 30S subunit head, 30S subunit platform, and 50S subunit CP maps, respectively), and raw movies in EMPIAR (entry EMPIAR-10509).

The following data sets were generated
The following previously published data sets were used

Article and author information

Author details

  1. Zoe L Watson

    Chemistry, University of California, Berkeley, Berkeley, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-4877-7914

  2. Fred R Ward

    Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-3825-5095

  3. Raphaël Méheust

    Earth and Planetary Science, University of California, Berkeley, Berkeley, United States
    Competing interests
    The authors declare that no competing interests exist.

  4. Omer Ad

    Chemistry, University of California, Berkeley, Berkeley, United States
    Competing interests
    The authors declare that no competing interests exist.

  5. Alanna Schepartz

    Chemistry, University of California, Berkeley, Berkeley, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-2127-3932

  6. Jillian F Banfield

    Department of Earth and Planetary Science, University of California, Berkeley, Berkeley, United States
    Competing interests
    The authors declare that no competing interests exist.

  7. Jamie HD Cate

    Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA, United States
    For correspondence
    j-h-doudna-cate@berkeley.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-5965-7902

Funding

National Science Foundation

  • Zoe L Watson
  • Omer Ad
  • Alanna Schepartz
  • Jamie HD Cate

National Institutes of Health

  • Fred R Ward

Innovative Genomics Institute

  • Raphaël Méheust
  • Jillian F Banfield

Chan Zuckerberg Biohub

  • Raphaël Méheust
  • Jillian F Banfield

Agilent Technologies

  • Omer Ad

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Copyright

© 2020, Watson 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

  • 13,915
    views
  • 1,471
    downloads
  • 183
    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)

  1. Zoe L Watson
  2. Fred R Ward
  3. Raphaël Méheust
  4. Omer Ad
  5. Alanna Schepartz
  6. Jillian F Banfield
  7. Jamie HD Cate
(2020)
Structure of the bacterial ribosome at 2 Å resolution
eLife 9:e60482.
https://doi.org/10.7554/eLife.60482

Share this article

https://doi.org/10.7554/eLife.60482

Categories and tags

Research organism

Further reading

    1. Computational and Systems Biology
    2. Structural Biology and Molecular Biophysics

    Discovery of a heparan sulfate binding domain in monkeypox virus H3 as an anti-poxviral drug target combining AI and MD simulations

    Bin Zheng, Meimei Duan ... Peng Zheng
    Research Article

    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.

    1. Chromosomes and Gene Expression
    2. Structural Biology and Molecular Biophysics

    Surprising features of nuclear receptor interaction networks revealed by live-cell single-molecule imaging

    Liza Dahal, Thomas GW Graham ... Xavier Darzacq
    Research Article

    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.

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

    Recognition and cleavage of human tRNA methyltransferase TRMT1 by the SARS-CoV-2 main protease

    Angel D'Oliviera, Xuhang Dai ... Jeffrey S Mugridge
    Research Article

    The SARS-CoV-2 main protease (Mpro or Nsp5) is critical for production of viral proteins during infection and, like many viral proteases, also targets host proteins to subvert their cellular functions. Here, we show that the human tRNA methyltransferase TRMT1 is recognized and cleaved by SARS-CoV-2 Mpro. TRMT1 installs the N2,N2-dimethylguanosine (m2,2G) modification on mammalian tRNAs, which promotes cellular protein synthesis and redox homeostasis. We find that Mpro can cleave endogenous TRMT1 in human cell lysate, resulting in removal of the TRMT1 zinc finger domain. Evolutionary analysis shows the TRMT1 cleavage site is highly conserved in mammals, except in Muroidea, where TRMT1 is likely resistant to cleavage. TRMT1 proteolysis results in reduced tRNA binding and elimination of tRNA methyltransferase activity. We also determined the structure of an Mpro-TRMT1 peptide complex that shows how TRMT1 engages the Mpro active site in an uncommon substrate binding conformation. Finally, enzymology and molecular dynamics simulations indicate that kinetic discrimination occurs during a later step of Mpro-mediated proteolysis following substrate binding. Together, these data provide new insights into substrate recognition by SARS-CoV-2 Mpro that could help guide future antiviral therapeutic development and show how proteolysis of TRMT1 during SARS-CoV-2 infection impairs both TRMT1 tRNA binding and tRNA modification activity to disrupt host translation and potentially impact COVID-19 pathogenesis or phenotypes.