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Structural basis for specific cleavage of Lys 63-linked polyubiquitin chains

A Corrigendum to this article was published on 13 November 2008


Deubiquitinating enzymes (DUBs) remove ubiquitin from conjugated substrates to regulate various cellular processes. The Zn2+-dependent DUBs AMSH and AMSH-LP regulate receptor trafficking by specifically cleaving Lys 63-linked polyubiquitin chains from internalized receptors. Here we report the crystal structures of the human AMSH-LP DUB domain alone and in complex with a Lys 63-linked di-ubiquitin at 1.2 Å and 1.6 Å resolutions, respectively. The AMSH-LP DUB domain consists of a Zn2+-coordinating catalytic core and two characteristic insertions, Ins-1 and Ins-2. The distal ubiquitin interacts with Ins-1 and the core, whereas the proximal ubiquitin interacts with Ins-2 and the core. The core and Ins-1 form a catalytic groove that accommodates the Lys 63 side chain of the proximal ubiquitin and the isopeptide-linked carboxy-terminal tail of the distal ubiquitin. This is the first reported structure of a DUB in complex with an isopeptide-linked ubiquitin chain, which reveals the mechanism for Lys 63-linkage-specific deubiquitination by AMSH family members.

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Figure 1: Overall structure of the AMSH-LP DUB domain alone and in complex with K63–Ub2.
Figure 2: Proximal ubiquitin recognition.
Figure 3: Distal ubiquitin recognition.
Figure 4: Catalytic mechanism.
Figure 5: Conservation among the DUB domains of AMSH, AMSH-LP and Rpn11.

Accession codes

Primary accessions

Protein Data Bank

Data deposits

The coordinates and structure factors of the AMSH-LP DUB domain and its complex with K63–Ub2 have been deposited in the Protein Data Bank with the accession codes 2ZNR and 2ZNV, respectively.


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We thank C. Toyoshima for support of this research. We thank M. Shirakawa and T. Tenno for advice about the K63–Ub2 preparation. We thank S. Kaiser for critical reading and improvement of this manuscript. We thank the beam-line staffs at NW12A of PF-AR (Tsukuba, Japan) for technical help during data collection. This work was supported by grants from MEXT to S.F. and O.N. Y.S. is supported by JSPS research fellowships for young scientists.

Author Contributions Y.S. carried out the crystallization and structure determination of the AMSH-LP DUB domain and its complex with K63–Ub2. A.Y. prepared K63–Ub2 and polyubiquitin chains with support from K.O., M.Y., O.N., K.I. and M.K. Y.S. and A.Y. carried out the in vitro DUB assays. K.I. and M.K. provided resources for K63–Ub2 preparation. A.Y., H.M. and S.F. assisted with the crystallization and structure determination. Y.S. and S.F. wrote the paper, with editing from A.Y., H.M., O.N., K.I. and M.K. S.F. and M.K. designed the research. All authors discussed the results and commented on the manuscript. S.F. supervised the work.

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Correspondence to Shuya Fukai.

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Sato, Y., Yoshikawa, A., Yamagata, A. et al. Structural basis for specific cleavage of Lys 63-linked polyubiquitin chains. Nature 455, 358–362 (2008).

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