title: A strategy for modulation of enzymes in the ubiquitin system.
abstract: The ubiquitin system regulates virtually all aspects of cellular function. We report a method to target the myriad enzymes that govern ubiquitination of protein substrates. We used massively diverse combinatorial libraries of ubiquitin variants to develop inhibitors of four deubiquitinases (DUBs) and analyzed the DUB-inhibitor complexes with crystallography. We extended the selection strategy to the ubiquitin conjugating (E2) and ubiquitin ligase (E3) enzymes and found that ubiquitin variants can also enhance enzyme activity. Last, we showed that ubiquitin variants can bind selectively to ubiquitin-binding domains. Ubiquitin variants exhibit selective function in cells and thus enable orthogonal modulation of specific enzymatic steps in the ubiquitin system.
authors.affiliation: Terrence Donnelly Center for Cellular and Biomolecular Research, University of Toronto, 160 College Street, Toronto, Ontario, Canada.
authors.forename: Andreas
authors.initials: A
authors.lastname: Ernst
authors.forename: George
authors.initials: G
authors.lastname: Avvakumov
authors.forename: Jiefei
authors.initials: J
authors.lastname: Tong
authors.forename: Yihui
authors.initials: Y
authors.lastname: Fan
authors.forename: Yanling
authors.initials: Y
authors.lastname: Zhao
authors.forename: Philipp
authors.initials: P
authors.lastname: Alberts
authors.forename: Avinash
authors.initials: A
authors.lastname: Persaud
authors.forename: John R
authors.initials: JR
authors.lastname: Walker
authors.forename: Ana-Mirela
authors.initials: AM
authors.lastname: Neculai
authors.forename: Dante
authors.initials: D
authors.lastname: Neculai
authors.forename: Andrew
authors.initials: A
authors.lastname: Vorobyov
authors.forename: Pankaj
authors.initials: P
authors.lastname: Garg
authors.forename: Linda
authors.initials: L
authors.lastname: Beatty
authors.forename: Pak-Kei
authors.initials: PK
authors.lastname: Chan
authors.forename: Yu-Chi
authors.initials: YC
authors.lastname: Juang
authors.forename: Marie-Claude
authors.initials: MC
authors.lastname: Landry
authors.forename: Christina
authors.initials: C
authors.lastname: Yeh
authors.forename: Elton
authors.initials: E
authors.lastname: Zeqiraj
authors.forename: Konstantina
authors.initials: K
authors.lastname: Karamboulas
authors.forename: Abdellah
authors.initials: A
authors.lastname: Allali-Hassani
authors.forename: Masoud
authors.initials: M
authors.lastname: Vedadi
authors.forename: Mike
authors.initials: M
authors.lastname: Tyers
authors.forename: Jason
authors.initials: J
authors.lastname: Moffat
authors.forename: Frank
authors.initials: F
authors.lastname: Sicheri
authors.forename: Laurence
authors.initials: L
authors.lastname: Pelletier
authors.forename: Daniel
authors.initials: D
authors.lastname: Durocher
authors.forename: Brian
authors.initials: B
authors.lastname: Raught
authors.forename: Daniela
authors.initials: D
authors.lastname: Rotin
authors.forename: Jianhua
authors.initials: J
authors.lastname: Yang
authors.forename: Michael F
authors.initials: MF
authors.lastname: Moran
authors.forename: Sirano
authors.initials: S
authors.lastname: Dhe-Paganon
authors.forename: Sachdev S
authors.initials: SS
authors.lastname: Sidhu
mesh_terms: D000595:Amino Acid Sequence; D020650:Combinatorial Chemistry Techniques; D017124:Conserved Sequence; D015195:Drug Design; D010450:Endopeptidases / Q000737:chemistry / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D008969:Molecular Sequence Data; D011480:Protease Inhibitors / Q000737:chemistry / Q000302:isolation & purification* / Q000494:pharmacology; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D054852:Small Molecule Libraries; D025801:Ubiquitin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D043222:Ubiquitin Thiolesterase / Q000737:chemistry / Q000378:metabolism*; D044763:Ubiquitin-Conjugating Enzymes / Q000737:chemistry / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases / Q000737:chemistry / Q000378:metabolism; D054875:Ubiquitination / Q000187:drug effects*
chemical_list: D011480:Protease Inhibitors; D054852:Small Molecule Libraries; C408875:USP21 protein, human; D025801:Ubiquitin; D044763:Ubiquitin-Conjugating Enzymes; D044767:Ubiquitin-Protein Ligases; D010450:Endopeptidases; D043222:Ubiquitin Thiolesterase