A Human DUB Protein Array for Clarification of Linkage Specificity of Polyubiquitin Chain and Application to Evaluation of Its Inhibitors

Biomedicines
Hirotaka TakahashiTatsuya Sawasaki

Abstract

Protein ubiquitinations play pivotal roles in many cellular processes, including homeostasis, responses to various stimulations, and progression of diseases. Deubiquitinating enzymes (DUBs) remove ubiquitin molecules from ubiquitinated proteins and cleave the polyubiquitin chain, thus negatively regulating numerous ubiquitin-dependent processes. Dysfunctions of many DUBs reportedly cause various diseases; therefore, DUBs are considered as important drug targets, although the biochemical characteristics and cellular functions of many DUBs are still unclear. Here, we established a human DUB protein array to detect the activity and linkage specificity of almost all human DUBs. Using a wheat cell-free protein synthesis system, 88 full-length recombinant human DUB proteins were prepared and termed the DUB array. In vitro DUB assays were performed with all of these recombinant DUBs, using eight linkage types of diubiquitins as substrates. As a result, 80 DUBs in the array showed DUB activities, and their linkage specificities were determined. These 80 DUBs included many biochemically uncharacterized DUBs in the past. In addition, taking advantage of these active DUB proteins, we applied the DUB array to evaluate the selectivities of ...Continue Reading

References

Oct 6, 1998·Annual Review of Biochemistry·A Hershko, A Ciechanover
Dec 2, 2004·Biochimica Et Biophysica Acta·Alexander Y Amerik, Mark Hochstrasser
Dec 6, 2005·Cell·Sebastian M B NijmanRené Bernards
Aug 15, 2006·Nature Cell Biology·Tingting YaoRobert E Cohen
Dec 18, 2007·Molecular Cell·Martin A CohnAlan D D'Andrea
Sep 13, 2008·ACS Chemical Biology·Fang Yi, Lynne Regan
Nov 26, 2008·BMC Biochemistry·Shweta SinghalRohan T Baker
Aug 13, 2009·Molecular Cancer Therapeutics·Frédéric CollandLaurent Daviet
Sep 11, 2010·Nature·Byung-Hoon LeeDaniel Finley
Dec 2, 2010·The Journal of Biological Chemistry·Stephen D WeeksPatrick J Loll
Apr 25, 2012·Annual Review of Biochemistry·David Komander, Michael Rape
Jul 13, 2013·Biochimica Et Biophysica Acta·Ziad M Eletr, Keith D Wilkinson
Oct 17, 2013·Molecular Cancer Therapeutics·Helena MistryKalindi Parmar
Aug 28, 2014·Nature Communications·Maria Stella RitortoMatthias Trost
Nov 5, 2014·Journal of Medicinal Chemistry·Chudi Ndubaku, Vickie Tsui
Feb 17, 2015·Nature Structural & Molecular Biology·Yusuke SatoShuya Fukai
May 15, 2015·Oncotarget·Rongbin WeiShangfeng Liu
Oct 21, 2016·Nature·Tycho E T MevissenDavid Komander
Oct 19, 2017·Nature·Lorna KategayaIngrid E Wertz
Oct 19, 2017·Nature·Andrew P TurnbullDavid Komander
Dec 29, 2017·Biochemical and Biophysical Research Communications·Atsushi UematsuTatsuya Sawasaki
Feb 25, 2018·Nature Communications·Thomas HermannsKay Hofmann
Mar 23, 2018·Nature Communications·David S HewingsIngrid E Wertz
Dec 18, 2018·Cell Chemical Biology·Ingrid E Wertz, Xiaojing Wang
Jan 4, 2020·Biochemical and Biophysical Research Communications·Satoshi YamanakaTatsuya Sawasaki

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Citations

Apr 13, 2021·Frontiers in Molecular Biosciences·Marie-Laure FogeronAnja Böckmann

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Methods Mentioned

BETA
ubiquitination
protein array
in vitro transcription
Deubiquitination
Protein Gel
Assay
AlphaScreen
cleavage assay
interaction assay

Software Mentioned

ImageJ
AlphaScreen

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