Structure of the bacterial ribosome at 2 Å resolution
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).
-
70S ribosome composite mapElectron Microsopy Databank, EMD-22586.
-
30S subunit headElectron Microsopy Databank, EMD-22635.
-
30S subunit platformElectron Microsopy Databank, EMD-22636.
-
50S subunit CP mapsElectron Microsopy Databank, EMD-22637.
-
E. coli Proteoform Families and G-PTM-D Database ExpansionMassIVE, accession MSV000081144.
Article and author information
Author details
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.
Reviewing Editor
- Sjors HW Scheres, MRC Laboratory of Molecular Biology, United Kingdom
Version history
- Received: June 27, 2020
- Accepted: September 11, 2020
- Accepted Manuscript published: September 14, 2020 (version 1)
- Version of Record published: October 12, 2020 (version 2)
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
-
- 12,998
- views
-
- 1,375
- downloads
-
- 158
- citations
Views, downloads and citations are aggregated across all versions of this paper published by eLife.
Download links
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)
Further reading
-
- Biochemistry and Chemical Biology
- Structural Biology and Molecular Biophysics
The type II class of RAF inhibitors currently in clinical trials paradoxically activate BRAF at subsaturating concentrations. Activation is mediated by induction of BRAF dimers, but why activation rather than inhibition occurs remains unclear. Using biophysical methods tracking BRAF dimerization and conformation, we built an allosteric model of inhibitor-induced dimerization that resolves the allosteric contributions of inhibitor binding to the two active sites of the dimer, revealing key differences between type I and type II RAF inhibitors. For type II inhibitors the allosteric coupling between inhibitor binding and BRAF dimerization is distributed asymmetrically across the two dimer binding sites, with binding to the first site dominating the allostery. This asymmetry results in efficient and selective induction of dimers with one inhibited and one catalytically active subunit. Our allosteric models quantitatively account for paradoxical activation data measured for 11 RAF inhibitors. Unlike type II inhibitors, type I inhibitors lack allosteric asymmetry and do not activate BRAF homodimers. Finally, NMR data reveal that BRAF homodimers are dynamically asymmetric with only one of the subunits locked in the active αC-in state. This provides a structural mechanism for how binding of only a single αC-in inhibitor molecule can induce potent BRAF dimerization and activation.
-
- Structural Biology and Molecular Biophysics
We integrate evolutionary predictions based on the neutral theory of molecular evolution with protein dynamics to generate mechanistic insight into the molecular adaptations of the SARS-COV-2 spike (S) protein. With this approach, we first identified candidate adaptive polymorphisms (CAPs) of the SARS-CoV-2 S protein and assessed the impact of these CAPs through dynamics analysis. Not only have we found that CAPs frequently overlap with well-known functional sites, but also, using several different dynamics-based metrics, we reveal the critical allosteric interplay between SARS-CoV-2 CAPs and the S protein binding sites with the human ACE2 (hACE2) protein. CAPs interact far differently with the hACE2 binding site residues in the open conformation of the S protein compared to the closed form. In particular, the CAP sites control the dynamics of binding residues in the open state, suggesting an allosteric control of hACE2 binding. We also explored the characteristic mutations of different SARS-CoV-2 strains to find dynamic hallmarks and potential effects of future mutations. Our analyses reveal that Delta strain-specific variants have non-additive (i.e., epistatic) interactions with CAP sites, whereas the less pathogenic Omicron strains have mostly additive mutations. Finally, our dynamics-based analysis suggests that the novel mutations observed in the Omicron strain epistatically interact with the CAP sites to help escape antibody binding.