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The structure of the deubiquitinase USP15 reveals a misaligned catalytic triad and an open ubiquitin-binding channel

Ward, Stephanie J.; Gratton, Hayley E.; Indrayudha, Peni; Michavila, Camille; Mukhopadhyay, Rishov; Maurer, Sigrun K.; Caulton, Simon G.; Emsley, Jonas; Dreveny, Ingrid

Authors

Stephanie J. Ward

Hayley E. Gratton

Peni Indrayudha

Camille Michavila

Rishov Mukhopadhyay

Sigrun K. Maurer

Simon G. Caulton

prof JONAS EMSLEY jonas.emsley@nottingham.ac.uk
Professor of Macromolecularcrystallography



Abstract

Ubiquitin specific protease 15 (USP15) regulates important cellular processes, including transforming growth factor β (TGF-β) signaling, mitophagy, mRNA processing, and innate immune responses; however, structural information on USP15’s catalytic domain is currently unavailable. Here, we determined crystal structures of the USP15 catalytic core domain, revealing a canonical USP fold, including a finger, palm, and thumb region. Unlike for the structure of paralog USP4, the catalytic triad is in an inactive configuration with the catalytic cysteine ~10Å apart from the catalytic histidine. This conformation is atypical, and a similar misaligned catalytic triad has so far been observed only for USP7, although USP15 and USP7 are differently regulated. Moreover, we found that the active site loops are flexible, resulting in a largely open ubiquitin tail binding channel. Comparison of the USP15 and USP4 structures points to a possible activation mechanism. Sequence differences between these two USPs mainly map to the S1’ region likely to confer specificity, whereas the S1 ubiquitin-binding pocket is highly conserved. Isothermal titration calorimetry monoubiquitin and linear diubiquitin binding experiments showed significant differences in their thermodynamic profiles, with USP15 displaying a lower affinity for monoubiquitin than USP4. Moreover, we report that USP15 is weakly inhibited by the antineoplastic agent mitoxantrone in vitro. A USP15-mitoxantrone complex structure disclosed that the anthracenedione interacts with the S1’ binding site. Our results reveal first insights into USP15’s catalytic domain structure, conformational changes, differences between paralogs, and small molecule interactions and establish a framework for cellular probe and inhibitor development.

Journal Article Type Article
Publication Date Nov 9, 2018
Journal Journal of Biological Chemistry
Print ISSN 0021-9258
Electronic ISSN 1083-351X
Publisher American Society for Biochemistry and Molecular Biology
Peer Reviewed Peer Reviewed
Volume 293
Issue 45
Pages 17362-17374
APA6 Citation Ward, S. J., Gratton, H. E., Indrayudha, P., Michavila, C., Mukhopadhyay, R., Maurer, S. K., …Dreveny, I. (2018). The structure of the deubiquitinase USP15 reveals a misaligned catalytic triad and an open ubiquitin-binding channel. Journal of Biological Chemistry, 293(45), 17362-17374. https://doi.org/10.1074/jbc.ra118.003857
DOI https://doi.org/10.1074/jbc.ra118.003857
Keywords Ubiquitin specific protease; Crystal structure; Cysteine protease; Deubiquitylation
Publisher URL http://www.jbc.org/content/early/2018/09/18/jbc.RA118.003857

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