Colicin E1 opens its hinge to plug TolC
- University of Kansas, United States
- University of Michigan, United States
- Rutgers University, United States
- Rutgers, The State University of New Jersey, United States
Abstract
The double membrane architecture of Gram-negative bacteria forms a barrier that is impermeable to most extracellular threats. Bacteriocin proteins evolved to exploit the accessible, surface-exposed proteins embedded in the outer membrane to deliver cytotoxic cargo. Colicin E1 is a bacteriocin produced by, and lethal to, Escherichia coli that hijacks the outer membrane proteins TolC and BtuB to enter the cell. Here we capture the colicin E1 translocation domain inside its membrane receptor, TolC, by high-resolution cryoEM to obtain the first reported structure of a bacteriocin bound to TolC. Colicin E1 binds stably to TolC as an open hinge through the TolC pore-an architectural rearrangement from colicin E1's unbound conformation. This binding is stable in live E. coli cells as indicated by single-molecule fluorescence microscopy. Finally, colicin E1 fragments binding to TolC plug the channel, inhibiting its native efflux function as an antibiotic efflux pump and heightening susceptibility to three antibiotic classes. In addition to demonstrating that these protein fragments are useful starting points for developing novel antibiotic potentiators, this method could be expanded to other colicins to inhibit other outer membrane protein functions.
Data availability
The manuscript has been deposited in BioRXiv BIORXIV/2019/692251CryoEM maps and models have been deposited with accession codes EMD-21960, EMD-21959, PDB ID 6WXI, and PDB ID 6WXH. The following data are publically available for the two structures:6WXH TolC + colE1Structure: https://files.rcsb.org/download/6WXH.cifEM map: https://ftp.wwpdb.org/pub/emdb/structures/EMD-21959/map/emd_21959.map.gzValidation report: https://files.rcsb.org/pub/pdb/validation_reports/wx/6wxh/6wxh_full_validation.pdf6WXI TolC aloneStructure: https://files.rcsb.org/download/6WXI.cifEM map: https://ftp.wwpdb.org/pub/emdb/structures/EMD-21960/map/emd_21960.map.gzValidation report: https://files.rcsb.org/pub/pdb/validation_reports/wx/6wxi/6wxi_full_validation.pdf
Article and author information
Author details
Jacqueline J Stevens
Ayotunde P Ikujuni
Virangika K Wimalasena
Funding
National Institute of General Medical Sciences (DP2GM128201)
- Joanna SG Slusky
National Institute of General Medical Sciences (P20GM113117)
- Joanna SG Slusky
National Institute of General Medical Sciences (P20GM103638)
- Joanna SG Slusky
Gordon and Betty Moore Foundation (Moore Inventor Fellowship)
- Joanna SG Slusky
National Institute of General Medical Sciences (P20 GM103418)
- S Jimmy Budiardjo
National Institute of General Medical Sciences (2K12GM063651)
- S Jimmy Budiardjo
National Institute of General Medical Sciences (R21-GM128022)
- Julie S Biteen
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Reviewing Editor
- David Drew, Stockholm University, Sweden
Publication history
- Preprint posted: July 4, 2019 (view preprint)
- Received: August 24, 2021
- Accepted: February 21, 2022
- Accepted Manuscript published: February 24, 2022 (version 1)
- Version of Record published: April 20, 2022 (version 2)
Copyright
© 2022, Budiardjo 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
-
- 1,893
- Page views
-
- 338
- Downloads
-
- 2
- 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)
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)
Colicin E1 opens its hinge to plug TolC
eLife 11:e73297.
https://doi.org/10.7554/eLife.73297
Further reading
-
- Microbiology and Infectious Disease
CRISPR-Cas systems are a family of adaptive immune systems that use small CRISPR RNAs (crRNAs) and CRISPR-associated (Cas) nucleases to protect prokaryotes from invading plasmids and viruses (i.e. phages). Type III systems launch a multi-layered immune response that relies upon both Cas and non-Cas cellular nucleases, and although the functions of Cas components have been well described, the identities and roles of non-Cas participants remain poorly understood. Previously, we showed that the Type III-A CRISPR-Cas system in Staphylococcus epidermidis employs two degradosome-associated nucleases, PNPase and RNase J2, to promote crRNA maturation and eliminate invading nucleic acids (Chou-Zheng and Hatoum-Aslan, 2019). Here, we identify RNase R as a third 'housekeeping' nuclease critical for immunity. We show that RNase R works in concert with PNPase to complete crRNA maturation, and identify specific interactions with Csm5, a member of the Type III effector complex, which facilitate nuclease recruitment/stimulation. Further, we demonstrate that RNase R and PNPase are required to maintain robust anti-plasmid immunity, particularly when targeted transcripts are sparse. Altogether, our findings expand the known repertoire of accessory nucleases required for Type III immunity and highlight the remarkable capacity of these systems to interface with diverse cellular pathways to ensure successful defense.
-
- Microbiology and Infectious Disease
In their natural environment, most bacteria preferentially live as complex surface-attached multicellular colonies called biofilms. Biofilms begin with a few cells adhering to a surface, where they multiply to form a mature colony. When conditions deteriorate, cells can leave the biofilm. This dispersion is thought to be an important process that modifies the overall biofilm architecture and that promotes colonization of new environments. In Caulobacter crescentus biofilms, extracellular DNA (eDNA) is released upon cell death and prevents newborn cells from joining the established biofilm. Thus, eDNA promotes the dispersal of newborn cells and the subsequent colonization of new environments. These observations suggest that eDNA is a cue for sensing detrimental environmental conditions in the biofilm. Here we show that the toxin-antitoxin system (TAS) ParDE4 stimulates cell death in areas of a biofilm with decreased O2 availability. In conditions where O2 availability is low, eDNA concentration is correlated with cell death. Cell dispersal away from biofilms is decreased when parDE4 is deleted, probably due to the lower local eDNA concentration. Expression of parDE4 is positively regulated by O2 and the expression of this operon is decreased in biofilms where O2 availability is low. Thus, a programmed cell death mechanism using an O2-regulated TAS stimulates dispersal away from areas of a biofilm with decreased O2 availability and favors colonization of a new, more hospitable environment.
