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From Proteopedia
Structure of SdeA DUB Domain disulfide crosslinked with Ubiquitin
Structural highlights
FunctionSDEA_LEGPH Secreted effector that interferes with the host cell ubiquitin pathway and is required for intracellular bacterial replication. Catalyzes the ubiquitination of several mammalian Rab proteins (Rab33b, Rab1, Rab6a and Rab30) during L.pneumophila infection, without engaging the standard cellular enzyme cascade (E1 and E2). Transfers an ADP-ribose moiety from NAD to the 'Arg-42' residue of ubiquitin in a reaction that releases nicotinamide. The modified ubiquitin is subsequently transferred to the substrate protein through an unknown mechanism that results in the release of AMP. Cannot ubiquitinate the endosomal Rab5 or the cytoskeletal small GTPase Rac1 (PubMed:27049943). Also acts as a deubiquitinase (DUB), catalyzing the cleavage of three of the most abundant polyubiquitin chains ('Lys-11', 'Lys-48' and 'Lys-63') with a distinct preference for 'Lys-63' linkages; is thus able to efficiently remove 'Lys-63'-linked polyubiquitin chains from the phagosomal surface. Is also able to remove NEDD8 from neddylated proteins, but is unable to recognize SUMO. The DUB activity of SdeA is important for regulating the dynamics of ubiquitin association with the bacterial phagosome, but is dispensable for its role in intracellular bacterial replication (PubMed:26598703).[1] [2] Publication Abstract from PubMedStructural characterization of the recognition of ubiquitin (Ub) by deubiquitinases (DUBs) has largely relied on covalent complexation of the DUB through its catalytic cysteine with a Ub C-terminal electrophile. The Ub electrophiles are accessed through intein chemistry in conjunction with chemical synthesis. Here, it was asked whether DUB-Ub covalent complexes could instead be accessed by simpler disulfide chemistry using a Ub cysteine mutant in which the last glycine has been replaced with a cysteine. The Ub cysteine mutant displayed a wide variability in disulfide formation across a panel of eukaryotic and prokaryotic DUBs, with some showing no detectable reaction while others robustly produced a disulfide complex. Using this approach, two disulfide-linked ubiquitin-bound complexes were crystallized, one involving the Legionella pneumophila effector SdeA DUB and the other involving the Orientia effector OtDUB. These DUBs had previously been crystallized in Ub-bound forms using the C-terminal electrophile strategy and noncovalent complexation, respectively. While the disulfide-linked SdeA DUB-Ub complex crystallized as expected, in the OtDUB complex the disulfide bond to the Ub mutant involved a cysteine that differed from the catalytic cysteine. Disulfide formation with the SdeA DUB catalytic cysteine was accompanied by local distortion of the helix carrying the active-site cysteine, whereas OtDUB reacted with the Ub mutant using a surface-exposed cysteine. Cocrystallization of ubiquitin-deubiquitinase complexes through disulfide linkage.,Negron Teron KI, Das C Acta Crystallogr D Struct Biol. 2023 Nov 1;79(Pt 11):1044-1055. doi: , 10.1107/S2059798323008501. Epub 2023 Oct 25. PMID:37877948[3] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See AlsoReferences
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