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

Abstract

© 2018 Ward et al. 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 ∼10 Å 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.

Citation

Ward, S. J., Gratton, H. E., Indrayudha, P., Michavila, C., Mukhopadhyay, R., Maurer, S. K., Caulton, S. G., Emsley, J., & 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

• Journal Article Type: Article
• Acceptance Date: Sep 18, 2018
• Online Publication Date: Sep 18, 2018
• Publication Date: Nov 9, 2018
• Deposit Date: Oct 11, 2018
• Publicly Available Date: Dec 5, 2018
• Journal: The 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
• DOI: https://doi.org/10.1074/jbc.RA118.003857
• Keywords: Ubiquitin specific protease; Crystal structure; Cysteine protease; Deubiquitylation
• Public URL: https://nottingham-repository.worktribe.com/output/1159420
• Publisher URL: https://www.sciencedirect.com/science/article/pii/S0021925820312643?via%3Dihub
• Contract Date: Dec 5, 2018