Competing scaffolding proteins determine capsid size during mobilization of Staphylococcus aureus pathogenicity islands
- National Institutes of Health, United States
- Virginia Commonwealth University, United States
- University of Alabama at Birmingham, United States
- Direct Electron, LP, United States
- Florida State University, United States
Abstract
Staphylococcus aureus pathogenicity islands (SaPIs), such as SaPI1, exploit specific helper bacteriophages, like 80a, for their high frequency mobilization, a process termed 'molecular piracy'. SaPI1 redirects the helper's assembly pathway to form small capsids that can only accommodate the smaller SaPI1 genome, but not a complete phage genome. SaPI1 encodes two proteins, CpmA and CpmB, that are responsible for this size redirection. We have determined the structures of the 80a and SaPI1 procapsids to near-atomic resolution by cryo-electron microscopy, and show that CpmB competes with the 80a scaffolding protein (SP) for a binding site on the capsid protein (CP), and works by altering the angle between capsomers. We probed these interactions genetically and identified second-site suppressors of lethal mutations in SP. Our structures show, for the first time, the detailed interactions between SP and CP in a bacteriophage, providing unique insights into macromolecular assembly processes.
Data availability
-
Capsid protein and C-terminal part of scaffolding protein in the Staphylococcus aureus phage 80alpha procapsidPublicly available at the EMBL-EBI EM Data Bank with accession number -EMDB-7030.
-
Capsid protein and C-terminal part of scaffolding protein in the Staphylococcus aureus phage 80alpha procapsidPublicly available at the RCSB Protein Data Bank with accession number 6B0X.
-
Capsid protein and C-terminal part of CpmB protein in the Staphylococcus aureus pathogenicity island 1 80alpha-derived procapsidPublicly available at the EMBL-EBI EM Data Bank with accession number -EMDB-7035.
-
Capsid protein and C-terminal part of CpmB protein in the Staphylococcus aureus pathogenicity island 1 80alpha-derived procapsidPublicly available at the RCSB Protein Data Bank with accession number 6B23.
Article and author information
Author details
Funding
National Institutes of Health (R01 AI083255)
- Terje Dokland
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2017, Dearborn 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
-
- 1,811
- views
-
- 328
- downloads
-
- 47
- 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)
Competing scaffolding proteins determine capsid size during mobilization of Staphylococcus aureus pathogenicity islands
eLife 6:e30822.
https://doi.org/10.7554/eLife.30822
Further reading
-
- Biochemistry and Chemical Biology
- Structural Biology and Molecular Biophysics
Dynamic conformational and structural changes in proteins and protein complexes play a central and ubiquitous role in the regulation of protein function, yet it is very challenging to study these changes, especially for large protein complexes, under physiological conditions. Here, we introduce a novel isobaric crosslinker, Qlinker, for studying conformational and structural changes in proteins and protein complexes using quantitative crosslinking mass spectrometry. Qlinkers are small and simple, amine-reactive molecules with an optimal extended distance of ~10 Å, which use MS2 reporter ions for relative quantification of Qlinker-modified peptides derived from different samples. We synthesized the 2-plex Q2linker and showed that the Q2linker can provide quantitative crosslinking data that pinpoints key conformational and structural changes in biosensors, binary and ternary complexes composed of the general transcription factors TBP, TFIIA, and TFIIB, and RNA polymerase II complexes.
-
- Structural Biology and Molecular Biophysics
Many voltage-gated potassium (Kv) channels display a time-dependent phenomenon called C-type inactivation, whereby prolonged activation by voltage leads to the inhibition of ionic conduction, a process that involves a conformational change at the selectivity filter toward a non-conductive state. Recently, a high-resolution structure of a strongly inactivated triple-mutant channel kv1.2-kv2.1-3m revealed a novel conformation of the selectivity filter that is dilated at its outer end, distinct from the well-characterized conductive state. While the experimental structure was interpreted as the elusive non-conductive state, our molecular dynamics simulations and electrophysiological measurements show that the dilated filter of kv1.2-kv2.1-3m is conductive and, as such, cannot completely account for the inactivation of the channel observed in the structural experiments. The simulation shows that an additional conformational change, implicating isoleucine residues at position 398 along the pore lining segment S6, is required to effectively block ion conduction. The I398 residues from the four subunits act as a state-dependent hydrophobic gate located immediately beneath the selectivity filter. By mutating I398 to Asparagine, ion permeation can be resumed in the kv1.2-kv2.1-3m channel, which was not a reversion C-type inactivation, since AgTxII fails to block the ionic permeation of kv1.2-kv2.1-3m_I398N. As a critical piece of the C-type inactivation machinery, this structural feature is the potential target of a broad class of QA blockers and negatively charged activators thus opening new research directions towards the development of drugs that specifically modulate gating-states of Kv channels.
