Structure of a type IV pilus machinery in the open and closed state

  1. Vicki A M Gold  Is a corresponding author
  2. Ralf Salzer
  3. Beate Averhoff
  4. Werner Kühlbrandt
  1. Max Planck Institute of Biophysics, Germany
  2. Goethe University Frankfurt, Germany

Abstract

Proteins of the secretin family form large macromolecular complexes, which assemble in the outer membrane of Gram-negative bacteria. Secretins are major components of type II and III secretion systems and are linked to extrusion of type IV pili (T4P) and to DNA uptake. By electron cryo-tomography of whole T. thermophilus cells, we determined the in situ structure of a T4P molecular machine in the open and the closed state. Comparison reveals a major conformational change whereby the N-terminal domains of the central secretin PilQ shift by ~30 Å, and two periplasmic gates open to make way for pilus extrusion. Furthermore, we determine the structure of the assembled pilus.

Article and author information

Author details

  1. Vicki A M Gold

    Department of Structural Biology, Max Planck Institute of Biophysics, Frankfurt am Main, Germany
    For correspondence
    vicki.gold@biophys.mpg.de
    Competing interests
    No competing interests declared.
  2. Ralf Salzer

    Molecular Microbiology and Bioenergetics, Institute of Molecular Biosciences, Goethe University Frankfurt, Frankfurt am Main, Germany
    Competing interests
    No competing interests declared.
  3. Beate Averhoff

    Molecular Microbiology and Bioenergetics, Institute of Molecular Biosciences, Goethe University Frankfurt, Frankfurt am Main, Germany
    Competing interests
    No competing interests declared.
  4. Werner Kühlbrandt

    Department of Structural Biology, Max Planck Institute of Biophysics, Frankfurt am Main, Germany
    Competing interests
    Werner Kühlbrandt, Reviewing editor, eLife.

Copyright

© 2015, Gold 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

  • 5,617
    views
  • 1,201
    downloads
  • 106
    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. Vicki A M Gold
  2. Ralf Salzer
  3. Beate Averhoff
  4. Werner Kühlbrandt
(2015)
Structure of a type IV pilus machinery in the open and closed state
eLife 4:e07380.
https://doi.org/10.7554/eLife.07380

Share this article

https://doi.org/10.7554/eLife.07380

Further reading

    1. Biochemistry and Chemical Biology
    2. Structural Biology and Molecular Biophysics
    Yamato Niitani, Kohei Matsuzaki ... Michio Tomishige
    Research Article

    The two identical motor domains (heads) of dimeric kinesin-1 move in a hand-over-hand process along a microtubule, coordinating their ATPase cycles such that each ATP hydrolysis is tightly coupled to a step and enabling the motor to take many steps without dissociating. The neck linker, a structural element that connects the two heads, has been shown to be essential for head–head coordination; however, which kinetic step(s) in the chemomechanical cycle is ‘gated’ by the neck linker remains unresolved. Here, we employed pre-steady-state kinetics and single-molecule assays to investigate how the neck-linker conformation affects kinesin’s motility cycle. We show that the backward-pointing configuration of the neck linker in the front kinesin head confers higher affinity for microtubule, but does not change ATP binding and dissociation rates. In contrast, the forward-pointing configuration of the neck linker in the rear kinesin head decreases the ATP dissociation rate but has little effect on microtubule dissociation. In combination, these conformation-specific effects of the neck linker favor ATP hydrolysis and dissociation of the rear head prior to microtubule detachment of the front head, thereby providing a kinetic explanation for the coordinated walking mechanism of dimeric kinesin.

    1. Structural Biology and Molecular Biophysics
    Christopher T Schafer, Raymond F Pauszek III ... David P Millar
    Research Article

    The canonical chemokine receptor CXCR4 and atypical receptor ACKR3 both respond to CXCL12 but induce different effector responses to regulate cell migration. While CXCR4 couples to G proteins and directly promotes cell migration, ACKR3 is G-protein-independent and scavenges CXCL12 to regulate extracellular chemokine levels and maintain CXCR4 responsiveness, thereby indirectly influencing migration. The receptors also have distinct activation requirements. CXCR4 only responds to wild-type CXCL12 and is sensitive to mutation of the chemokine. By contrast, ACKR3 recruits GPCR kinases (GRKs) and β-arrestins and promiscuously responds to CXCL12, CXCL12 variants, other peptides and proteins, and is relatively insensitive to mutation. To investigate the role of conformational dynamics in the distinct pharmacological behaviors of CXCR4 and ACKR3, we employed single-molecule FRET to track discrete conformational states of the receptors in real-time. The data revealed that apo-CXCR4 preferentially populates a high-FRET inactive state, while apo-ACKR3 shows little conformational preference and high transition probabilities among multiple inactive, intermediate and active conformations, consistent with its propensity for activation. Multiple active-like ACKR3 conformations are populated in response to agonists, compared to the single CXCR4 active-state. This and the markedly different conformational landscapes of the receptors suggest that activation of ACKR3 may be achieved by a broader distribution of conformational states than CXCR4. Much of the conformational heterogeneity of ACKR3 is linked to a single residue that differs between ACKR3 and CXCR4. The dynamic properties of ACKR3 may underly its inability to form productive interactions with G proteins that would drive canonical GPCR signaling.