Skip to main content
SpringerLink
Log in

Backbone 1H, 13C and 15N resonance assignment of the ubiquitin specific protease 7 catalytic domain (residues 208–554) in complex with a small molecule ligand

  • Article
  • Published:
Biomolecular NMR Assignments Aims and scope Submit manuscript

Abstract

The backbone 1H, 13C and 15N resonance assignment of Ubiquitin Specific Protease 7 catalytic domain (residues 208–554) was performed in its complex with a small molecule ligand and in its apo form as a reference. The amide 1H-15N signal intensities were boosted by an amide hydrogen exchange protocol, where expressed 2H, 13C, 15N-labeled protein was unfolded and re-folded to ensure exchange of amide deuterons to protons. The resonance assignments were used to determine chemical shift perturbations on ligand binding, which are consistent with the binding site observed by crystallography.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8

Similar content being viewed by others

Data availability

The backbone 1H, 13C and 15N resonance assignment of apo Ubiquitin Specific Protease 7 catalytic domain (residues 208–554) was deposited in Biological Magnetic Resonance Bank under accession code 51912. The assignment for the complex with compound 21 (O’Dowd et al. 2018a) was deposited under code 51913.

References

  • Ashton NW, Valles GJ, Jaiswal N, Bezsonova I, Woodgate R (2021) DNA polymerase iota interacts with both the TRAF-like and UBL1-2 domains of USP7. J Mol Biol 433(2):166733. https://doi.org/10.1016/j.jmb.2020.166733

    Article  Google Scholar 

  • Bezsonova I, Avvakumov G, Dhe-Paganon S, Arrowsmith CH (2010) Solution NMR structure of human ubiquitin specific protease Usp7 UBL domain (residues 537-664). NESG target hr4395c/ SGC-Toronto. Biological Magnetic Resonance Data Bank deposition code 16789. https://bmrb.io/data_library/summary/index.php?bmrbId=16789

  • Bezsonova I, Lemak A, Avvakumov G, Xue S, Dhe-Paganon S, Montelione GT, Arrowsmith CH (2010) Solution NMR structure of human ubiquitin specific protease Usp7 UBL domain (residues 537–664). NESG target hr4395c/ SGC-Toronto. Protein Data Bank deposition code 2KVR. https://www.rcsb.org/structure/2KVR

  • Ching W, Koyuncu E, Singh S, Arbelo-Roman C, Hartl B, Kremmer E, Speiseder T, Meier C, Dobner T (2013) A ubiquitin-specific protease possesses a decisive role for adenovirus replication and oncogene-mediated transformation. PLoS Pathog 9(3):e1003273. https://doi.org/10.1371/journal.ppat.1003273

    Article  Google Scholar 

  • Di Lello P, Rouge L, Pan B, Maurer T (2016) 1H, 13C and 15N backbone resonance assignment for the 40.5 kDa catalytic domain of ubiquitin specific protease 7 (USP7). Biomol NMR Assign 10(2):345–349. https://doi.org/10.1007/s12104-016-9698-3

    Article  Google Scholar 

  • Di Lello P, Pastor R, Murray JM, Blake RA, Cohen F, Crawford TD, Drobnick J, Drummond J, Kategaya L, Kleinheinz T, Maurer T, Rougé L, Zhao X, Wertz I, Ndubaku C, Tsui V (2017) Discovery of small-molecule inhibitors of ubiquitin specific protease 7 (USP7) using integrated NMR and in silico techniques. J Med Chem 60(24):10056–10070. https://doi.org/10.1021/acs.jmedchem.7b01293

    Article  Google Scholar 

  • Eldirany S, Pozhidaeva A, Bezsonova I (2018) NMR resonance assignment of ubiquitin specific protease 7 (USP7). Backbone resonance assignment of USP7 UBL45 domains (residues 884-1102). Biological Magnetic Resonance Data Bank deposition code 27529. https://bmrb.io/data_library/summary/index.php?bmrbId=27529

  • Fernández-Montalván A, Bouwmeester T, Joberty G, Mader R, Mahnke M, Pierrat B, Schlaeppi J-M, Worpenberg S, Gerhartz B (2007) Biochemical characterization of USP7 reveals post-translational modification sites and structural requirements for substrate processing and subcellular localization. FEBS J 274:4256–4270. https://doi.org/10.1111/j.1742-4658.2007.05952.x

    Article  Google Scholar 

  • Gavory G, O’Dowd CR, Helm MD, Flasz J, Arkoudis E, Dossang A, Hughes C, Cassidy E, McClelland K, Odrzywol E, Page N, Barker O, Miel H, Harrison T (2018) Discovery and characterization of highly potent and selective allosteric USP7 inhibitors. Nat Chem Biol 14:118–125. https://doi.org/10.1038/nchembio.2528

    Article  Google Scholar 

  • Holowaty MN, Zeghouf M, Hong Wu, Tellam J, Athanasopoulos V, Greenblatt J, Frappier L (2003) Interaction with the herpesvirus-associated ubiquitin-specific protease HAUSP/USP7. J Biol Chem 278(32):29987–29994. https://doi.org/10.1074/jbc.M303977200

    Article  Google Scholar 

  • Hyberts SG, Takeuchi K, Wagner G (2010) Poisson-gap sampling and FM reconstruction for enhancing resolution and sensitivity of protein NMR data. J Am Chem Soc 132(7):2145–2147. https://doi.org/10.1021/ja908004w

    Article  Google Scholar 

  • Hu M, Li P, Li M, Wei Gu, Cohen RE, Shi Y (2002) Crystal structure of a UBP-family deubiquitinating enzyme in isolation and in complex with ubiquitin aldehyde. Cell 111(7):1041–1054. https://doi.org/10.1016/S0092-8674(02)01199-6

    Article  Google Scholar 

  • Jaravine V, Ibraghimov I, Orekhov VY (2006) Removal of a time barrier for high-resolution multidimensional NMR spectroscopy. Nat Methods 3:605–607. https://doi.org/10.1038/nmeth900

    Article  Google Scholar 

  • Kategaya L, Di Lello P, Rougé L, Pastor R, Clark KR, Drummond J, Kleinheinz T, Lin E, Upton J-P, Prakash S, Heideker J, McCleland M, Ritorto MS, Alessi DR, Trost M, Bainbridge TW, Kwok MCM, Ma TP, Stiffler Z, Brasher B, Tang Y, Jaishankar P, Hearn BR, Renslo AR, Arkin MR, Cohen F, Kebing Yu, Peale F, Gnad F, Chang MT, Klijn C, Blackwood E, Martin SE, Forrest WF, Ernst JA, Ndubaku C, Wang X, Beresini MH, Tsui V, Schwerdtfeger C, Blake RA, Murray J, Maurer T, Wertz IE (2017) USP7 small-molecule inhibitors interfere with ubiquitin binding. Nature 550:534–538. https://doi.org/10.1038/nature24006

    Article  ADS  Google Scholar 

  • Keller RLJ (2004) The computer aided resonance assignment tutorial. Cantina Verlag, Goldau

    Google Scholar 

  • Kim RQ, Willem J.van Dijk, Titia K. Sixma, (2016) Structure of USP7 catalytic domain and three Ubl-domains reveals a connector α-helix with regulatory role. J Struct Biol 195(1):11–18. https://doi.org/10.1016/j.jsb.2016.05.005

    Article  Google Scholar 

  • Kim RQ, Geurink PP, Mulder MPC, Fish A, Ekkebus R, El Oualid F, van Dijk WJ, van Dalen D, Ovaa H, van Ingen H, Sixma TK (2019) Kinetic analysis of multistep USP7 mechanism shows critical role for target protein in activity. Nat Commun 10:231. https://doi.org/10.1038/s41467-018-08231-5

    Article  ADS  Google Scholar 

  • Lamberto I, Liu X, Seo H-S, Schauer NJ, Iacob RE, Wanyi Hu, Das D, Mikhailova T, Weisberg EL, Engen JR, Anderson KC, Chauhan D, Dhe-Paganon S, Buhrlage SJ (2017) Structure-guided development of a potent and selective non-covalent active-site inhibitor of USP7. Cell Chem Biol 24:1490–1500. https://doi.org/10.1016/j.chembiol.2017.09.003

    Article  Google Scholar 

  • Leger PR, Hu DX, Biannic B, Bui M, Han X, Karbarz E, Maung J, Okano A, Osipov M, Shibuya GM, Young K, Higgs C, Abraham B, Bradford D, Cho C, Colas C, Jacobson S, Ohol YM, Pookot D, Rana P, Sanchez J, Shah N, Sun M, Wong S, Brockstedt DG, Kassner PD, Schwarz JB, Wustrow DJ (2020) Discovery of potent, selective, and orally bioavailable inhibitors of USP7 with in vivo antitumor activity. J Med Chem 63:5398–5420. https://doi.org/10.1021/acs.jmedchem.0c00245

    Article  Google Scholar 

  • Li R, Woodward C (1999) The hydrogen exchange core and protein folding. Protein Sci 8:1571–1591. https://doi.org/10.1110/ps.8.8.1571

    Article  Google Scholar 

  • Li M, Chen D, Shiloh A, Luo J, Nikolaev AY, Qin J, Wei Gu (2002) Deubiquitination of p53 by HAUSP is an important pathway for p53 stabilization. Nature 416:648–653. https://doi.org/10.1038/nature737

    Article  ADS  Google Scholar 

  • Li M, Liu S, Chen H, Zhou X, Zhou J, Zhou S, Yuan H, Qing-Long Xu, Liu J, Cheng K, Sun H, Wang Y, Chen C, Wen X (2020) N-benzylpiperidinol derivatives as novel USP7 inhibitors: Structure–activity relationships and X-ray crystallographic studies. Eur J Med Chem 199:112279. https://doi.org/10.1016/j.ejmech.2020.112279

    Article  Google Scholar 

  • Molland K, Zhou Q, Mesecar AD (2014) A 2.2 Å resolution structure of the USP7 catalytic domain in a new space group elaborates upon structural rearrangements resulting from ubiquitin binding. Acta Cryst F70:283–287. https://doi.org/10.1107/S2053230X14002519

    Article  Google Scholar 

  • Morotti A, Panuzzo C, Crivellaro S, Pergolizzi B, Familiari U, Berger AH, Saglio G, Pandolfi PP (2014) BCR-ABL disrupts PTEN nuclear-cytoplasmic shuttling through phosphorylation-dependent activation of HAUSP. Leukemia 28:1326–1333. https://doi.org/10.1038/leu.2013.370

    Article  Google Scholar 

  • O’Dowd CR, Helm MD, Shane Rountree JS, Flasz JT, Arkoudis E, Miel H, Hewitt PR, Jordan L, Barker O, Hughes C, Rozycka E, Cassidy E, McClelland K, Odrzywol E, Page N, Feutren-Burton S, Dvorkin S, Gavory G, Harrison T (2018a) Identification and structure-guided development of pyrimidinone based USP7 inhibitors. ACS Med Chem Lett 9(3):238–243. https://doi.org/10.1021/acsmedchemlett.7b00512

    Article  Google Scholar 

  • O’Dowd CR, Harrison T, Hewitt PR, Shane Rountree JS, Miel H, Burkamp F, Jordan L, Helm MD, Brocatelli F, Crawford JJ, Gazzard L, Wertz I, Lee W (2018b) Piperidine derivatives as inhibitors of ubiquitin specific protease 7. Patent WO 2018/073602 A1. https://patentscope.wipo.int/search/en/detail.jsf?docId=WO2018073602

  • Ozen A, Rouge L, Bashore C, Hearn BR, Skelton NJ, Dueber EC (2018) Selectively modulating conformational states of USP7 catalytic domain for activation. Structure 26:72–84. https://doi.org/10.1016/j.str.2017.11.010

    Article  Google Scholar 

  • Palazón-Riquelme P, Worboys JD, Green J, Valera A, Martín-Sánchez F, Pellegrini C, Brough D, López-Castejón G (2018) USP7 and USP47 deubiquitinases regulate NLRP3 inflammasome activation. EMBO Rep 19:e44766. https://doi.org/10.15252/embr.201744766

    Article  Google Scholar 

  • Pervushin K, Riek R, Wider G, Wüthrich K (2017) Attenuated T2 relaxation by mutual cancellation of dipole–dipole coupling and chemical shift anisotropy indicates an avenue to NMR structures of very large biological macromolecules in solution. Proc Natl Acad Sci USA 94(23):12366–12371. https://doi.org/10.1073/pnas.94.23.12366

    Article  ADS  Google Scholar 

  • Pfoh R, Lacdao IK, Georges AA, Capar A, Zheng H, Frappier L, Saridakis V (2015) Crystal structure of USP7 ubiquitin-like domains with an ICP0 peptide reveals a novel mechanism used by viral and cellular proteins to target USP7. PLoS Pathog 11(6):e1004950. https://doi.org/10.1371/journal.ppat.1004950

    Article  Google Scholar 

  • Pozhidaeva AK, Mohni KN, Dhe-Paganon S, Arrowsmith CH, Weller SK, Korzhnev DM, Bezsonova I (2015) Structural characterization of interaction between human ubiquitin-specific protease 7 and immediate-early protein ICP0 of herpes simplex virus-1. J Biol Chem 290(38):22907–22918. https://doi.org/10.1074/jbc.M115.664805

    Article  Google Scholar 

  • Pozhidaeva A, Valles G, Wang F, Jian Wu, Sterner DE, Nguyen P, Joseph Weinstock KG, Kumar S, Kanyo J, Wright D, Bezsonova I (2017) USP7-specific inhibitors target and modify the enzyme’s active site via distinct chemical mechanisms. Cell Chem Biol 24(12):1501–1512. https://doi.org/10.1016/j.chembiol.2017.09.004

    Article  Google Scholar 

  • Rouge L, Bainbridge TW, Kwok M, Tong R, Di Lello P, Wertz IE, Maurer T, Ernst JA, Murray J (2016) Molecular understanding of USP7 substrate recognition and C-terminal activation. Structure 24:1335–1345. https://doi.org/10.1016/j.str.2016.05.020

    Article  Google Scholar 

  • Saridakis V, Sheng Yi, Sarkari F, Holowaty MN, Shire K, Nguyen T, Zhang RG, Liao J, Lee W, Edwards AM, Arrowsmith CH, Frappier L (2005) Structure of the p53 binding domain of HAUSP/USP7 bound to epstein-barr nuclear antigen 1: implications for EBV-mediated immortalization. Mol Cell 18(1):25–36. https://doi.org/10.1016/j.molcel.2005.02.029

    Article  Google Scholar 

  • Turnbull AP, Ioannidis S, Krajewski WW, Pinto-Fernandez A, Heride C, Martin ACL, Tonkin LM, Townsend EC, Buker SM, Lancia DR, Caravella JA, Toms AV, Charlton TM, Lahdenranta J, Wilker E, Follows BC, Evans NJ, Stead L, Alli C, Zarayskiy VV, Talbot AC, Buckmelter AJ, Wang M, McKinnon CL, Saab F, McGouran JF, Century H, Gersch M, Pittman MS, Gary Marshall C, Raynham TM, Simcox M, Stewart LMD, McLoughlin SB, Escobedo JA, Bair KW, Dinsmore CJ, Hammonds TR, Kim S, Urbé S, Clague MJ, Kessler BM, Komander D (2017) Molecular basis of USP7 inhibition by selective small-molecule inhibitors. Nature 550:481–486. https://doi.org/10.1038/nature24451

    Article  ADS  Google Scholar 

  • Valles G, Pozhidaeva A, Korzhnev D, Bezsonova I (2022) Backbone and ILV side-chain NMR resonance assignments of the catalytic domain of human deubiquitinating enzyme USP7. Biomol NMR Assign 16(2):197–203. https://doi.org/10.1007/s12104-022-10079-2

    Article  Google Scholar 

  • van Loosdregt J, Fleskens V, Juan Fu, Brenkman AB, Bekker CPJ, Pals CEGM, Meerding J, Berkers CR, Barbi J, Gröne A, Sijts AJAM, Maurice MM, Kalkhoven E, Prakken BJ, Ovaa H, Pan F, Zaiss DMW, Coffer PJ (2013) Stabilization of the transcription factor Foxp3 by the deubiquitinase USP7 increases treg-cell-suppressive capacity. Immunity 39(2):259–271. https://doi.org/10.1016/j.immuni.2013.05.018

    Article  Google Scholar 

  • Alonso-de Vega I, Martín Y, Smits VA (2014) USP7 controls Chk1 protein stability by direct deubiquitination. Cell Cycle 13(24):3921–3926. https://doi.org/10.4161/15384101.2014.973324

    Article  Google Scholar 

  • Vladislav Yu, Orekhov II, Billeter M (2003) Optimizing resolution in multidimensional NMR by three-way decomposition. J Biomol NMR 27(2):165–173. https://doi.org/10.1023/A:1024944720653

    Article  Google Scholar 

  • Wang Z, Kang W, You Y, Pang J, Ren H, Suo Z, Liu H, Zheng Y (2019) USP7: novel drug target in cancer therapy. Front Pharmacol 10:427. https://doi.org/10.3389/fphar.2019.00427

    Article  Google Scholar 

  • Xiang Q, Hyunwoo Ju, Li Q, Mei S-C, Chen D, Choi YB, Nicholas J (2018) Human herpesvirus 8 interferon regulatory factors 1 and 3 mediate replication and latency activities via interactions with USP7 deubiquitinase. J Virol 92(7):e2003–e2017. https://doi.org/10.1128/jvi.02003-17

    Article  Google Scholar 

Download references

Funding

This work was supported by Innovate UK (Knowledge Transfer Partnership 11447).

Author information

Authors and Affiliations

  1. Manchester Institute of Biotechnology, University of Manchester, Manchester, United Kingdom

    Maya J. Pandya, Matthew J. Cliff & Jonathan P. Waltho

  2. C4X Discovery Ltd, Manchester One, 53 Portland Street, Manchester, M1 3LD, United Kingdom

    Maya J. Pandya, Wojciech Augustyniak, Hugh R. W. Dannatt & Martin J. Watson

  3. Biophysics Department, NMR and Protein Production, Evotec SE, Hamburg, Germany

    Ilka Lindner, Anne Stinn, Jan Kahmann & Koen Temmerman

Authors
  1. Maya J. Pandya
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Wojciech Augustyniak
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Matthew J. Cliff
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Ilka Lindner
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Anne Stinn
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Jan Kahmann
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Koen Temmerman
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Hugh R. W. Dannatt
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Jonathan P. Waltho
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Martin J. Watson
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

M.J.P., M.J.C., J.P.W. and M.J.W. initiated the project and designed the experimental protocol. I.L, A.S., J.K. and K.T. expressed and purified protein. M.J.P. performed hydrogen exchange/protein re-folding. M.J.P. M.J.C and H.R.W.D. recorded spectra. W.A. performed resonance assignments, analysed the results, wrote the manuscript and prepared the figures. All authors reviewed the manuscript.

Corresponding author

Correspondence to Wojciech Augustyniak.

Ethics declarations

Competing interests

The authors declare no competing interests.

Ethical approval

Not applicable.

Additional information

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Pandya, M.J., Augustyniak, W., Cliff, M.J. et al. Backbone 1H, 13C and 15N resonance assignment of the ubiquitin specific protease 7 catalytic domain (residues 208–554) in complex with a small molecule ligand. Biomol NMR Assign 18, 33–44 (2024). https://doi.org/10.1007/s12104-024-10165-7

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12104-024-10165-7

Keywords

Search

Navigation