Abstract
Eight different types of ubiquitin linkages are present in eukaryotic cells that regulate diverse biological processes. Proteins that mediate specific assembly and disassembly of atypical Lys6, Lys27, Lys29 and Lys33 linkages are mainly unknown. We here reveal how the human ovarian tumor (OTU) domain deubiquitinase (DUB) TRABID specifically hydrolyzes both Lys29- and Lys33-linked diubiquitin. A crystal structure of the extended catalytic domain reveals an unpredicted ankyrin repeat domain that precedes an A20-like catalytic core. NMR analysis identifies the ankyrin domain as a new ubiquitin-binding fold, which we have termed AnkUBD, and DUB assays in vitro and in vivo show that this domain is crucial for TRABID efficiency and linkage specificity. Our data are consistent with AnkUBD functioning as an enzymatic S1′ ubiquitin-binding site, which orients a ubiquitin chain so that Lys29 and Lys33 linkages are cleaved preferentially.
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Change history
29 February 2012
In the version of Supplementary Figure 5b originally posted online, three images were incorrect: those for DMSO K27only Ub, MG132 K27only Ub and MG132 K29only Ub. The correct figure is now shown. The conclusions from this figure remain unchanged. These errors have been corrected in this file as of 29 February 2012.
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Acknowledgements
We would like to thank R. Williams, M. Allen, M. Fiedler, N. Soler, S. Freund, C. Johnson, S. McLaughlin, Y. Kulathu, Y. Ye, A. Bremm, M. Busch (Medical Research Council Laboratory of Molecular Biology), D.S. Hameed (Netherlands Cancer Institute) and K. Hofmann (Miltenyi Biotec) for reagents, help with experiments and discussions. Crystallographic data were collected at the European Synchrotron Radiation Facility beamline ID23-2. J.D.F.L. was supported by an Association for International Cancer Research grant (no. 07-0040 to M.B.). D.K. is an European Molecular Biology Organization (EMBO) Young Investigator. This work was supported by the Medical Research Council (MC_US_A024_0059 to M.B., MC_US_A024_0056 to D.K., and MC_US_A024_0051 to J.W.C.).
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D.K., M.B. and J.D.F.L. designed the research. J.D.F.L., D.K., J.M., T.E.T.M., T.J.R. and M.A. conducted the experiments. F.E., H.O., S.V. and J.W.C. contributed reagents. D.K. wrote the manuscript, with help from all authors.
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H.O. and F.E. are cofounders of UbiQ Bio BV.
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Licchesi, J., Mieszczanek, J., Mevissen, T. et al. An ankyrin-repeat ubiquitin-binding domain determines TRABID's specificity for atypical ubiquitin chains. Nat Struct Mol Biol 19, 62–71 (2012). https://doi.org/10.1038/nsmb.2169
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DOI: https://doi.org/10.1038/nsmb.2169
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