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Phosphorylation-dependent activity of the deubiquitinase DUBA

Nature Structural & Molecular Biology volume 19, pages 171175 (2012) | Download Citation


Addition and removal of ubiquitin or ubiquitin chains to and from proteins is a tightly regulated process that contributes to cellular signaling and protein stability. Here we show that phosphorylation of the human deubiquitinase DUBA (OTUD5) at a single residue, Ser177, is both necessary and sufficient to activate the enzyme. The crystal structure of the ubiquitin aldehyde adduct of active DUBA reveals a marked cooperation between phosphorylation and substrate binding. An intricate web of interactions involving the phosphate and the C-terminal tail of ubiquitin cause DUBA to fold around its substrate, revealing why phosphorylation is essential for deubiquitinase activity. Phosphoactivation of DUBA represents an unprecedented mode of protease regulation and a clear link between two major cellular signal transduction systems: phosphorylation and ubiquitin modification.

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We thank K.C. Dong (Genentech) for the gift of Ub-BEA, M.P. Kamps (University of California, San Diego) for the gift of ER-Hoxb8 retrovirus, T. Kitamura (University of Tokyo) for the gift of pMXs-puro, the Genentech baculovirus cloning and expression group for production of DUBA in insect cells, and the Genentech oligonucleotide synthesis and sequencing groups. Portions of this research were carried out at the Advanced Light Source, supported by the Director, Office of Science, Office of Basic Energy Sciences of the US Department of Energy, under Contract No. DE-AC02-05CH11231.

Author information


  1. Department of Early Discovery Biochemistry, Genentech, South San Francisco, California, USA.

    • Oscar W Huang
    •  & Andrea G Cochran
  2. Department of Structural Biology, Genentech, South San Francisco, California, USA.

    • Xiaolei Ma
    • , JianPing Yin
    • , Jeremy Flinders
    • , Till Maurer
    • , Ivan Bosanac
    • , Sarah G Hymowitz
    •  & Melissa A Starovasnik
  3. Department of Physiological Chemistry, Genentech, South San Francisco, California, USA.

    • Nobuhiko Kayagaki
    •  & Vishva M Dixit
  4. Department of Protein Chemistry, Genentech, South San Francisco, California, USA.

    • Qui Phung
    •  & David Arnott


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Proteins and complexes were purified by O.W.H. and J.Y.; O.W.H. carried out all biochemical studies, with advice from A.G.C., and J.Y. crystallized proteins under advice from M.A.S. J.F. and T.M. conducted the NMR analysis, with advice from M.A.S. I.B. and X.M. solved the crystal structures of apo DUBA and the ubiquitin complex, respectively, with advice from S.G.H. Q.P. and D.A. provided mass spectral data. Evaluation of DUBA phosphorylation in macrophages was done by N.K. and V.M.D. The manuscript was written by A.G.C., with contributions from O.W.H., X.M., J.Y., T.M., N.K., Q.P., S.G.H. and M.A.S.

Competing interests

All authors are employees of Genentech, a member of the Roche Group.

Corresponding author

Correspondence to Andrea G Cochran.

Supplementary information

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  1. 1.

    Supplementary Text and Figures

    Supplementary Figures 1–5 and Supplementary Methods


  1. 1.

    Supplementary Video 1

    DUBA apo structure “morphing” into the pS177 DUBA Ub-al complex structure.

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  1. 1.

    Supplementary Data

    Backbone 1H, 15N, and 13Cα assignments for apo DUBA OTU domain, pSer177 OTU domain, and the BEA-ubiquitin adduct of the pSer177 OTU domain.

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