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

Mechanochemical coupling and bi-phasic force-velocity dependence in the ultra-fast ring ATPase SpoIIIE

  1. Ninning Liu
  2. Gheorghe Chistol
  3. Yuanbo Cui
  4. Carlos Bustamante  Is a corresponding author
  1. University of California, Berkeley, United States
Research Advance
  • Cited 3
  • Views 1,314
  • Annotations
Cite this article as: eLife 2018;7:e32354 doi: 10.7554/eLife.32354

Abstract

Multi-subunit ring-shaped ATPases are molecular motors that harness chemical free energy to perform vital mechanical tasks such as polypeptide translocation, DNA unwinding, and chromosome segregation. Previously we reported the intersubunit coordination and stepping behavior of the hexameric ring-shaped ATPase SpoIIIE (Liu et al., 2015). Here we use optical tweezers to characterize the motor's mechanochemistry. Analysis of the motor response to external force at various nucleotide concentrations identifies phosphate release as the likely force-generating step. Analysis of SpoIIIE pausing indicates that pauses are off-pathway events. Characterization of SpoIIIE slipping behavior reveals that individual motor subunits engage DNA upon ATP binding. Furthermore, we find that SpoIIIE's velocity exhibits an intriguing bi-phasic dependence on force. We hypothesize that this behavior is an adaptation of ultra-fast motors tasked with translocating DNA from which they must also remove DNA-bound protein roadblocks. Based on these results, we formulate a comprehensive mechanochemical model for SpoIIIE.

Article and author information

Author details

  1. Ninning Liu

    Jason L Choy Laboratory of Single Molecule Biophysics, University of California, Berkeley, Berkeley, United States
    Competing interests
    The authors declare that no competing interests exist.
  2. Gheorghe Chistol

    Jason L Choy Laboratory of Single Molecule Biophysics, University of California, Berkeley, Berkeley, United States
    Competing interests
    The authors declare that no competing interests exist.
  3. Yuanbo Cui

    Jason L Choy Laboratory of Single Molecule Biophysics, University of California, Berkeley, Berkeley, United States
    Competing interests
    The authors declare that no competing interests exist.
  4. Carlos Bustamante

    Jason L Choy Laboratory of Single Molecule Biophysics, University of California, Berkeley, Berkeley, United States
    For correspondence
    carlosb@berkeley.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-2970-0073

Funding

Howard Hughes Medical Institute

  • Carlos Bustamante

National Institutes of Health (R01GM071552)

  • Carlos Bustamante

National Institutes of Health (R01GM032543)

  • Carlos Bustamante

U.S. Department of Energy (DE-AC02-05CH11231)

  • Carlos Bustamante

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

Reviewing Editor

  1. Taekjip Ha, Johns Hopkins University School of Medicine, United States

Publication history

  1. Received: November 22, 2017
  2. Accepted: March 3, 2018
  3. Accepted Manuscript published: March 5, 2018 (version 1)
  4. Version of Record published: March 19, 2018 (version 2)

Copyright

© 2018, Liu 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

  • 1,314
    Page views
  • 213
    Downloads
  • 3
    Citations

Article citation count generated by polling the highest count across the following sources: PubMed Central, Crossref, 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. Cancer Biology
    Xiao-Tong Lin et al.
    Research Article Updated

    Dysregulation of tumor-relevant proteins may contribute to human hepatocellular carcinoma (HCC) tumorigenesis. FBXO45 is an E3 ubiquitin ligase that is frequently elevated expression in human HCC. However, it remains unknown whether FBXO45 is associated with hepatocarcinogenesis and how to treat HCC patients with high FBXO45 expression. Here, IHC and qPCR analysis revealed that FBXO45 protein and mRNA were highly expressed in 54.3% (57 of 105) and 52.2% (132 of 253) of the HCC tissue samples, respectively. Highly expressed FBXO45 promoted liver tumorigenesis in transgenic mice. Mechanistically, FBXO45 promoted IGF2BP1 ubiquitination at the Lys190 and Lys450 sites and subsequent activation, leading to the upregulation of PLK1 expression and the induction of cell proliferation and liver tumorigenesis in vitro and in vivo. PLK1 inhibition or IGF2BP1 knockdown significantly blocked FBXO45-driven liver tumorigenesis in FBXO45 transgenic mice, primary cells, and HCCs. Furthermore, IHC analysis on HCC tissue samples revealed a positive association between the hyperexpression of FBXO45 and PLK1/IGF2BP1, and both had positive relationship with poor survival in HCC patients. Thus, FBXO45 plays an important role in promoting liver tumorigenesis through IGF2BP1 ubiquitination and activation, and subsequent PLK1 upregulation, suggesting a new strategy for treating HCC by targeting FBXO45/IGF2BP1/PLK1 axis.

    1. Biochemistry and Chemical Biology
    2. Cell Biology
    Rania Elsabrouty et al.
    Research Article Updated

    UbiA prenyltransferase domain-containing protein-1 (UBIAD1) utilizes geranylgeranyl pyrophosphate (GGpp) to synthesize the vitamin K2 subtype menaquinone-4. The prenyltransferase has emerged as a key regulator of sterol-accelerated, endoplasmic reticulum (ER)-associated degradation (ERAD) of HMG CoA reductase, the rate-limiting enzyme in synthesis of cholesterol and nonsterol isoprenoids including GGpp. Sterols induce binding of UBIAD1 to reductase, inhibiting its ERAD. Geranylgeraniol (GGOH), the alcohol derivative of GGpp, disrupts this binding and thereby stimulates ERAD of reductase and translocation of UBIAD1 to Golgi. We now show that overexpression of Type 1 polyisoprenoid diphosphate phosphatase (PDP1), which dephosphorylates GGpp and other isoprenyl pyrophosphates to corresponding isoprenols, abolishes protein geranylgeranylation as well as GGOH-induced ERAD of reductase and Golgi transport of UBIAD1. Conversely, these reactions are enhanced in the absence of PDP1. Our findings indicate PDP1-mediated hydrolysis of GGpp significantly contributes to a feedback mechanism that maintains optimal intracellular levels of the nonsterol isoprenoid.