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Structural basis of substrate recognition by a bacterial deubiquitinase important for dynamics of phagosome ubiquitination.

ABSTRACT: Manipulation of the host's ubiquitin network is emerging as an important strategy for counteracting and repurposing the posttranslational modification machineries of the host by pathogens. Ubiquitin E3 ligases encoded by infectious agents are well known, as are a variety of viral deubiquitinases (DUBs). Bacterial DUBs have been discovered, but little is known about the structure and mechanism underlying their ubiquitin recognition. In this report, we found that members of the Legionella pneumophila SidE effector family harbor a DUB module important for ubiquitin dynamics on the bacterial phagosome. Structural analysis of this domain alone and in complex with ubiquitin vinyl methyl ester (Ub-VME) reveals unique molecular contacts used in ubiquitin recognition. Instead of relying on the Ile44 patch of ubiquitin, as commonly used in eukaryotic counterparts, the SdeADub module engages Gln40 of ubiquitin. The architecture of the active-site cleft presents an open arrangement with conformational plasticity, permitting deubiquitination of three of the most abundant polyubiquitin chains, with a distinct preference for Lys63 linkages. We have shown that this preference enables efficient removal of Lys63 linkages from the phagosomal surface. Remarkably, the structure reveals by far the most parsimonious use of molecular contacts to achieve deubiquitination, with less than 1,000 Å(2) of accessible surface area buried upon complex formation with ubiquitin. This type of molecular recognition appears to enable dual specificity toward ubiquitin and the ubiquitin-like modifier NEDD8.

SUBMITTER: Sheedlo MJ

PROVIDER: S-EPMC4679006 | biostudies-literature | 2015 Dec

REPOSITORIES: biostudies-literature

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Structural basis of substrate recognition by a bacterial deubiquitinase important for dynamics of phagosome ubiquitination.

Sheedlo Michael J MJ Qiu Jiazhang J Tan Yunhao Y Paul Lake N LN Luo Zhao-Qing ZQ Das Chittaranjan C

Proceedings of the National Academy of Sciences of the United States of America 20151123 49

Manipulation of the host's ubiquitin network is emerging as an important strategy for counteracting and repurposing the posttranslational modification machineries of the host by pathogens. Ubiquitin E3 ligases encoded by infectious agents are well known, as are a variety of viral deubiquitinases (DUBs). Bacterial DUBs have been discovered, but little is known about the structure and mechanism underlying their ubiquitin recognition. In this report, we found that members of the Legionella pneumoph ...[more]

PMID: 26598703

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Project description:Pathogenic bacteria have acquired a vast array of eukaryotic-protein-like proteins via intimate interaction with host cells. Bacterial effector proteins that function as ubiquitin ligases and deubiquitinases (DUBs) are remarkable examples of such molecular mimicry. LotA, a Legionella pneumophila effector, belongs to the ovarian tumor (OTU) superfamily, which regulates diverse ubiquitin signals by their DUB activities. LotA harbors two OTU domains that have distinct reactivities; the first one is responsible for the cleavage of the K6-linked ubiquitin chain, and the second one shows an uncommon preference for long chains of ubiquitin. Here, we report the crystal structure of a middle domain of LotA (LotAM), which contains the second OTU domain. LotAM consists of two distinct subdomains, a catalytic domain having high structural similarity with human OTU DUBs and an extended helical lobe (EHL) domain, which is characteristically conserved only in Legionella OTU DUBs. The docking simulation of LotAM with ubiquitin suggested that hydrophobic and electrostatic interactions between the EHL of LotAM and the C-terminal region of ubiquitin are crucial for the binding of ubiquitin to LotAM. The structure-based mutagenesis demonstrated that the acidic residue in the characteristic short helical segment termed the "helical arm" is essential for the enzymatic activity of LotAM. The EHL domain of the three Legionella OTU DUBs, LotA, LotB, and LotC, share the "helical arm" structure, suggesting that the EHL domain defines the Lot-OTUs as a unique class of DUBs. IMPORTANCE To successfully colonize, some pathogenic bacteria hijack the host ubiquitin system. Legionella OTU-like-DUBs (Lot-DUBs) are novel bacterial deubiquitinases found in effector proteins of L. pneumophila. LotA is a member of Lot-DUBs and has two OTU domains (OTU1 and OTU2). We determined the structure of a middle fragment of LotA (LotAM), which includes OTU2. LotAM consists of the conserved catalytic domain and the Legionella OTUs-specific EHL domain. The docking simulation with ubiquitin and the mutational analysis suggested that the acidic surface in the EHL is essential for enzymatic activity. The structure of the EHL differs from those of other Lot-DUBs, suggesting that the variation of the EHL is related to the variable cleaving specificity of each DUB.

Dataset Information

Structural basis of substrate recognition by a bacterial deubiquitinase important for dynamics of phagosome ubiquitination.

Publications

Structural basis of substrate recognition by a bacterial deubiquitinase important for dynamics of phagosome ubiquitination.

Similar Datasets

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