1. Structural Biology and Molecular Biophysics
Download icon

Structure of Escherichia coli respiratory complex I reconstituted into lipid nanodiscs reveals an uncoupled conformation

  1. Rouslan G Efremov  Is a corresponding author
  2. Piotr Kolata
  1. VIB-VUB, Belgium
Research Article
  • Cited 3
  • Views 1,878
  • Annotations
Cite this article as: eLife 2021;10:e68710 doi: 10.7554/eLife.68710

Abstract

Respiratory complex I is a multi-subunit membrane protein complex that reversibly couples NADH oxidation and ubiquinone reduction with proton translocation against trans-membrane potential. Complex I from Escherichia coli is among the best functionally characterized complexes, but its structure remains unknown, hindering further mechanistic studies to understand the enzyme coupling mechanism. Here we describe the single particle cryo-electron microscopy (cryo-EM) structure of the entire catalytically active E. coli complex I reconstituted into lipid nanodiscs. The structure of this mesophilic bacterial complex I displays highly dynamic connection between the peripheral and membrane domains. The peripheral domain assembly is stabilized by unique terminal extensions and an insertion loop. The membrane domain structure reveals novel dynamic features. Unusual conformation of the conserved interface between the peripheral and membrane domains suggests an uncoupled conformation of the complex. Considering constraints imposed by the structural data we suggest a new simple hypothetical coupling mechanism for the molecular machine.

Data availability

Cryo-EM density maps and atomic models are deposited into the PDB and EMDB databases with the following accession codes: cytoplasmic domain (PDB ID: 7NZ1, EMD-12661), membrane domain (PDB ID: 7NYH, EMD-12652), entire complex conformation 1 (PDB ID: 7NYR, EMD-12653), conformation 2 (PDB ID: 7NYU,EMD-12654), conformation 3 (PDB ID: 7NYV, EMD-12655).

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

Article and author information

Author details

  1. Rouslan G Efremov

    VIB Center for Structural Biology, VUB Structural Biology Brussels, VIB-VUB, Brussels, Belgium
    For correspondence
    rouslan.efremov@vub.vib.be
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-7516-8658
  2. Piotr Kolata

    VIB Center for Structural Biology, VUB Structural Biology Brussels, VIB-VUB, Brussels, Belgium
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-9484-5025

Funding

Fonds Wetenschappelijk Onderzoek (G0H5916N)

  • Rouslan G Efremov

Fonds Wetenschappelijk Onderzoek (G.0266.15N)

  • Rouslan G Efremov

H2020 European Research Council (726436)

  • Rouslan G Efremov

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

Reviewing Editor

  1. Andrew P Carter, MRC Laboratory of Molecular Biology, United Kingdom

Publication history

  1. Received: March 23, 2021
  2. Preprint posted: April 10, 2021 (view preprint)
  3. Accepted: July 23, 2021
  4. Accepted Manuscript published: July 26, 2021 (version 1)
  5. Accepted Manuscript updated: July 27, 2021 (version 2)
  6. Version of Record published: August 11, 2021 (version 3)

Copyright

© 2021, Efremov & Kolata

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

  • 1,878
    Page views
  • 268
    Downloads
  • 3
    Citations

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

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
    2. Structural Biology and Molecular Biophysics
    Nazia Ahmad et al.
    Research Article Updated

    L,D-transpeptidase function predominates in atypical 3 → 3 transpeptide networking of peptidoglycan (PG) layer in Mycobacterium tuberculosis. Prior studies of L,D-transpeptidases have identified only the catalytic site that binds to peptide moiety of the PG substrate or β-lactam antibiotics. This insight was leveraged to develop mechanism of its activity and inhibition by β-lactams. Here, we report identification of an allosteric site at a distance of 21 Å from the catalytic site that binds the sugar moiety of PG substrates (hereafter referred to as the S-pocket). This site also binds a second β-lactam molecule and influences binding at the catalytic site. We provide evidence that two β-lactam molecules bind co-operatively to this enzyme, one non-covalently at the S-pocket and one covalently at the catalytic site. This dual β-lactam-binding phenomenon is previously unknown and is an observation that may offer novel approaches for the structure-based design of new drugs against M. tuberculosis.

    1. Structural Biology and Molecular Biophysics
    Stephanie A Wankowicz et al.
    Research Article Updated

    While protein conformational heterogeneity plays an important role in many aspects of biological function, including ligand binding, its impact has been difficult to quantify. Macromolecular X-ray diffraction is commonly interpreted with a static structure, but it can provide information on both the anharmonic and harmonic contributions to conformational heterogeneity. Here, through multiconformer modeling of time- and space-averaged electron density, we measure conformational heterogeneity of 743 stringently matched pairs of crystallographic datasets that reflect unbound/apo and ligand-bound/holo states. When comparing the conformational heterogeneity of side chains, we observe that when binding site residues become more rigid upon ligand binding, distant residues tend to become more flexible, especially in non-solvent-exposed regions. Among ligand properties, we observe increased protein flexibility as the number of hydrogen bonds decreases and relative hydrophobicity increases. Across a series of 13 inhibitor-bound structures of CDK2, we find that conformational heterogeneity is correlated with inhibitor features and identify how conformational changes propagate differences in conformational heterogeneity away from the binding site. Collectively, our findings agree with models emerging from nuclear magnetic resonance studies suggesting that residual side-chain entropy can modulate affinity and point to the need to integrate both static conformational changes and conformational heterogeneity in models of ligand binding.