Conformational dynamics of the nucleotide binding domains and the power stroke of a heterodimeric ABC transporter

  1. Smriti Mishra
  2. Brandy Verhalen
  3. Richard A Stein
  4. Po-Chao Wen
  5. Emad Tajkhorshid
  6. Hassane S Mchaourab  Is a corresponding author
  1. Vanderbilt University, United States
  2. University of Illinois, United States

Abstract

Multidrug ATP binding cassette (ABC) exporters are ubiquitous ABC transporters that extrude cytotoxic molecules across cell membranes. Despite recent progress in structure determination of these transporters, the conformational motion that transduces the energy of ATP hydrolysis to the work of substrate translocation remains undefined. Here, we have investigated the conformational cycle of BmrCD, a representative of the heterodimer family of ABC exporters that have an intrinsically impaired nucleotide binding site. We measured distances between pairs of spin labels monitoring the movement of the nucleotide binding (NBD) and transmembrane domains (TMD). The results expose previously unobserved structural intermediates of the NBDs arising from asymmetric configuration of catalytically inequivalent nucleotide binding sites. The two-state transition of the TMD, from an inward- to an outward-facing conformation, is driven exclusively by ATP hydrolysis. These findings provide direct evidence of divergence in the mechanism of ABC exporters.

Article and author information

Author details

  1. Smriti Mishra

    Vanderbilt University, Nashville, United States
    Competing interests
    The authors declare that no competing interests exist.
  2. Brandy Verhalen

    Vanderbilt University, Nashville, United States
    Competing interests
    The authors declare that no competing interests exist.
  3. Richard A Stein

    Vanderbilt University, Nashville, United States
    Competing interests
    The authors declare that no competing interests exist.
  4. Po-Chao Wen

    University of Illinois, Urbana, United States
    Competing interests
    The authors declare that no competing interests exist.
  5. Emad Tajkhorshid

    University of Illinois, Urbana, United States
    Competing interests
    The authors declare that no competing interests exist.
  6. Hassane S Mchaourab

    Vanderbilt University, Nashville, United States
    For correspondence
    hassane.mchaourab@vanderbilt.edu
    Competing interests
    The authors declare that no competing interests exist.

Reviewing Editor

  1. Volker Dötsch, Goethe University, Germany

Version history

  1. Received: March 8, 2014
  2. Accepted: May 13, 2014
  3. Accepted Manuscript published: May 16, 2014 (version 1)
  4. Version of Record published: June 10, 2014 (version 2)

Copyright

© 2014, Mishra 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

  • 3,426
    Page views
  • 502
    Downloads
  • 100
    Citations

Article citation count generated by polling the highest count across the following sources: Crossref, Scopus, 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)

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. Smriti Mishra
  2. Brandy Verhalen
  3. Richard A Stein
  4. Po-Chao Wen
  5. Emad Tajkhorshid
  6. Hassane S Mchaourab
(2014)
Conformational dynamics of the nucleotide binding domains and the power stroke of a heterodimeric ABC transporter
eLife 3:e02740.
https://doi.org/10.7554/eLife.02740

Share this article

https://doi.org/10.7554/eLife.02740

Further reading

    1. Biochemistry and Chemical Biology
    Valentin Bohl, Nele Merret Hollmann ... Axel Mogk
    Research Article

    Heat stress can cause cell death by triggering the aggregation of essential proteins. In bacteria, aggregated proteins are rescued by the canonical Hsp70/AAA+ (ClpB) bi-chaperone disaggregase. Man-made, severe stress conditions applied during, e.g., food processing represent a novel threat for bacteria by exceeding the capacity of the Hsp70/ClpB system. Here, we report on the potent autonomous AAA+ disaggregase ClpL from Listeria monocytogenes that provides enhanced heat resistance to the food-borne pathogen enabling persistence in adverse environments. ClpL shows increased thermal stability and enhanced disaggregation power compared to Hsp70/ClpB, enabling it to withstand severe heat stress and to solubilize tight aggregates. ClpL binds to protein aggregates via aromatic residues present in its N-terminal domain (NTD) that adopts a partially folded and dynamic conformation. Target specificity is achieved by simultaneous interactions of multiple NTDs with the aggregate surface. ClpL shows remarkable structural plasticity by forming diverse higher assembly states through interacting ClpL rings. NTDs become largely sequestered upon ClpL ring interactions. Stabilizing ring assemblies by engineered disulfide bonds strongly reduces disaggregation activity, suggesting that they represent storage states.

    1. Biochemistry and Chemical Biology
    2. Structural Biology and Molecular Biophysics
    Tien M Phan, Young C Kim ... Jeetain Mittal
    Research Article

    The heterochromatin protein 1 (HP1) family is a crucial component of heterochromatin with diverse functions in gene regulation, cell cycle control, and cell differentiation. In humans, there are three paralogs, HP1α, HP1β, and HP1γ, which exhibit remarkable similarities in their domain architecture and sequence properties. Nevertheless, these paralogs display distinct behaviors in liquid-liquid phase separation (LLPS), a process linked to heterochromatin formation. Here, we employ a coarse-grained simulation framework to uncover the sequence features responsible for the observed differences in LLPS. We highlight the significance of the net charge and charge patterning along the sequence in governing paralog LLPS propensities. We also show that both highly conserved folded and less-conserved disordered domains contribute to the observed differences. Furthermore, we explore the potential co-localization of different HP1 paralogs in multicomponent assemblies and the impact of DNA on this process. Importantly, our study reveals that DNA can significantly reshape the stability of a minimal condensate formed by HP1 paralogs due to competitive interactions of HP1α with HP1β and HP1γ versus DNA. In conclusion, our work highlights the physicochemical nature of interactions that govern the distinct phase-separation behaviors of HP1 paralogs and provides a molecular framework for understanding their role in chromatin organization.