Dataset Information

Structural basis for ubiquitin recognition and autoubiquitination by Rabex-5.

ABSTRACT: Rabex-5 is an exchange factor for Rab5, a master regulator of endosomal trafficking. Rabex-5 binds monoubiquitin, undergoes covalent ubiquitination and contains an intrinsic ubiquitin ligase activity, all of which require an N-terminal A20 zinc finger followed immediately by a helix. The structure of the N-terminal portion of Rabex-5 bound to ubiquitin at 2.5-A resolution shows that Rabex-5-ubiquitin interactions occur at two sites. The first site is a new type of ubiquitin-binding domain, an inverted ubiquitin-interacting motif, which binds with approximately 29-microM affinity to the canonical Ile44 hydrophobic patch on ubiquitin. The second is a diaromatic patch on the A20 zinc finger, which binds with approximately 22-microM affinity to a polar region centered on Asp58 of ubiquitin. The A20 zinc-finger diaromatic patch mediates ubiquitin-ligase activity by directly recruiting a ubiquitin-loaded ubiquitin-conjugating enzyme.

SUBMITTER: Lee S

PROVIDER: S-EPMC1578505 | biostudies-literature | 2006 Mar

REPOSITORIES: biostudies-literature

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Structural basis for ubiquitin recognition and autoubiquitination by Rabex-5.

Lee Sangho S Tsai Yien Che YC Mattera Rafael R Smith William J WJ Kostelansky Michael S MS Weissman Allan M AM Bonifacino Juan S JS Hurley James H JH

Nature structural & molecular biology 20060205 3

Rabex-5 is an exchange factor for Rab5, a master regulator of endosomal trafficking. Rabex-5 binds monoubiquitin, undergoes covalent ubiquitination and contains an intrinsic ubiquitin ligase activity, all of which require an N-terminal A20 zinc finger followed immediately by a helix. The structure of the N-terminal portion of Rabex-5 bound to ubiquitin at 2.5-A resolution shows that Rabex-5-ubiquitin interactions occur at two sites. The first site is a new type of ubiquitin-binding domain, an in ...[more]

PMID: 16462746

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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 for ubiquitin recognition and autoubiquitination by Rabex-5.

Publications

Structural basis for ubiquitin recognition and autoubiquitination by Rabex-5.

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