Insights into the ubiquitin transfer cascade catalyzed by the Legionella effector SidC
Abstract
The causative agent of Legionnaires' disease, Legionella pneumophila, delivers more than 330 virulent effectors to its host to establish an intracellular membrane-bound organelle called the Legionella containing vacuole. Among the army of Legionella effectors, SidC and its paralog SdcA have been identified as novel bacterial ubiquitin (Ub) E3 ligases. To gain insight into the molecular mechanism of SidC/SdcA as Ub ligases, we determined the crystal structures of a binary complex of the N-terminal catalytic SNL domain of SdcA with its cognate E2 UbcH5C and a ternary complex consisting of the SNL domain of SidC with the Ub-linked E2 UbcH7. These two structures reveal the molecular determinants governing the Ub transfer cascade catalyzed by SidC. Together, our data support a common mechanism in the Ub transfer cascade in which the donor Ub is immobilized with its C-terminal tail locked in an extended conformation, priming the donor Ub for catalysis.
Data availability
Atomic coordinates and structure factors for the reported structures have been deposited into the Protein Data Bank under the accession codes 6CP0 (SdcA-UbcH5C), 6CP2 (SidC-UbcH7~Ub)
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structure for SdcA-UbcH5CPublicly available at the RCSB Protein Data Bank (accession no. 6CP0).
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structure for SidC-UbcH7~UbPublicly available at the RCSB Protein Data Bank (accession no. 6CP2).
Article and author information
Author details
Funding
National Institute of General Medical Sciences (5R01GM116964)
- Yuxin Mao
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Reviewing Editor
- Cynthia Wolberger, Johns Hopkins University, United States
Version history
- Received: February 23, 2018
- Accepted: July 16, 2018
- Accepted Manuscript published: July 17, 2018 (version 1)
- Version of Record published: July 27, 2018 (version 2)
Copyright
© 2018, Wasilko 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.
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Further reading
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Previously we showed that 2D template matching (2DTM) can be used to localize macromolecular complexes in images recorded by cryogenic electron microscopy (cryo-EM) with high precision, even in the presence of noise and cellular background (Lucas et al., 2021; Lucas et al., 2022). Here, we show that once localized, these particles may be averaged together to generate high-resolution 3D reconstructions. However, regions included in the template may suffer from template bias, leading to inflated resolution estimates and making the interpretation of high-resolution features unreliable. We evaluate conditions that minimize template bias while retaining the benefits of high-precision localization, and we show that molecular features not present in the template can be reconstructed at high resolution from targets found by 2DTM, extending prior work at low-resolution. Moreover, we present a quantitative metric for template bias to aid the interpretation of 3D reconstructions calculated with particles localized using high-resolution templates and fine angular sampling.
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