A General Strategy for Discovery of Inhibitors and Activators of RING and U-box E3 Ligases with Ubiquitin Variants

Mads Gabrielsen; Lori Buetow; Mark A Nakasone; Syed Feroj Ahmed; Gary J Sibbet; Brian O Smith; Wei Zhang; Sachdev S Sidhu; Danny T Huang

doi:10.1016/j.molcel.2017.09.027

A General Strategy for Discovery of Inhibitors and Activators of RING and U-box E3 Ligases with Ubiquitin Variants

Mads Gabrielsen, Lori Buetow, Mark A Nakasone, Syed Feroj Ahmed, Gary J Sibbet, Brian O Smith, Wei Zhang, Sachdev S Sidhu, Danny T Huang

Biological Chemistry and Drug Discovery

Research output: Contribution to journal › Article › peer-review

49 Citations (Scopus)

13 Downloads (Pure)

Abstract

RING and U-box E3 ubiquitin ligases regulate diverse eukaryotic processes and have been implicated in numerous diseases, but targeting these enzymes remains a major challenge. We report the development of three ubiquitin variants (UbVs), each binding selectively to the RING or U-box domain of a distinct E3 ligase: monomeric UBE4B, phosphorylated active CBL, or dimeric XIAP. Structural and biochemical analyses revealed that UbVs specifically inhibited the activity of UBE4B or phosphorylated CBL by blocking the E2∼Ub binding site. Surprisingly, the UbV selective for dimeric XIAP formed a dimer to stimulate E3 activity by stabilizing the closed E2∼Ub conformation. We further verified the inhibitory and stimulatory functions of UbVs in cells. Our work provides a general strategy to inhibit or activate RING/U-box E3 ligases and provides a resource for the research community to modulate these enzymes.

Original language	English
Pages (from-to)	456-470.e10
Number of pages	26
Journal	Molecular Cell
Volume	68
Issue number	2
DOIs	https://doi.org/10.1016/j.molcel.2017.09.027
Publication status	Published - 19 Oct 2017

Keywords

Drug Discovery/methods
Enzyme Activators/chemistry
Enzyme Inhibitors/chemistry
HEK293 Cells
HeLa Cells
Humans
Protein Multimerization/drug effects
Tumor Suppressor Proteins/agonists
Ubiquitin-Protein Ligase Complexes/antagonists & inhibitors
Ubiquitin-Protein Ligases
X-Linked Inhibitor of Apoptosis Protein/agonists

Access to Document

10.1016/j.molcel.2017.09.027Licence: CC BY

Publisher final version
Gabrielsen et al., 2017, Molecular Cell 68, 456–470 October 19, 2017 ª 2017 The Author(s). Published by Elsevier Inc. https://doi.org/10.1016/j.molcel.2017.09.027
Final published version, 5.44 MBLicence: CC BY

Cite this

@article{b2fb8a6a4fe546d1aaf57b50d4f5e663,

title = "A General Strategy for Discovery of Inhibitors and Activators of RING and U-box E3 Ligases with Ubiquitin Variants",

abstract = "RING and U-box E3 ubiquitin ligases regulate diverse eukaryotic processes and have been implicated in numerous diseases, but targeting these enzymes remains a major challenge. We report the development of three ubiquitin variants (UbVs), each binding selectively to the RING or U-box domain of a distinct E3 ligase: monomeric UBE4B, phosphorylated active CBL, or dimeric XIAP. Structural and biochemical analyses revealed that UbVs specifically inhibited the activity of UBE4B or phosphorylated CBL by blocking the E2∼Ub binding site. Surprisingly, the UbV selective for dimeric XIAP formed a dimer to stimulate E3 activity by stabilizing the closed E2∼Ub conformation. We further verified the inhibitory and stimulatory functions of UbVs in cells. Our work provides a general strategy to inhibit or activate RING/U-box E3 ligases and provides a resource for the research community to modulate these enzymes.",

keywords = "Drug Discovery/methods, Enzyme Activators/chemistry, Enzyme Inhibitors/chemistry, HEK293 Cells, HeLa Cells, Humans, Protein Multimerization/drug effects, Tumor Suppressor Proteins/agonists, Ubiquitin-Protein Ligase Complexes/antagonists & inhibitors, Ubiquitin-Protein Ligases, X-Linked Inhibitor of Apoptosis Protein/agonists",

author = "Mads Gabrielsen and Lori Buetow and Nakasone, {Mark A} and Ahmed, {Syed Feroj} and Sibbet, {Gary J} and Smith, {Brian O} and Wei Zhang and Sidhu, {Sachdev S} and Huang, {Danny T}",

note = "This work was supported by Cancer Research UK (C596/A23278) and European Research Council (ERC) under the European Union{\textquoteright}s Horizon 2020 research and innovation programme (grant agreement n 647849) awarded to D.T.H. and a Genome Canada Disruptive Innovation in Genomics grant (OGI-119) and a CQDM-OCE EXPLORE Project (#23927) awarded to S.S.SCopyright {\textcopyright} 2017 The Author(s). Published by Elsevier Inc. All rights reserved.",

year = "2017",

month = oct,

day = "19",

doi = "10.1016/j.molcel.2017.09.027",

language = "English",

volume = "68",

pages = "456--470.e10",

journal = "Molecular Cell",

issn = "1097-2765",

publisher = "Elsevier",

number = "2",

}

TY - JOUR

T1 - A General Strategy for Discovery of Inhibitors and Activators of RING and U-box E3 Ligases with Ubiquitin Variants

AU - Gabrielsen, Mads

AU - Buetow, Lori

AU - Nakasone, Mark A

AU - Ahmed, Syed Feroj

AU - Sibbet, Gary J

AU - Smith, Brian O

AU - Zhang, Wei

AU - Sidhu, Sachdev S

AU - Huang, Danny T

N1 - This work was supported by Cancer Research UK (C596/A23278) and European Research Council (ERC) under the European Union’s Horizon 2020 research and innovation programme (grant agreement n 647849) awarded to D.T.H. and a Genome Canada Disruptive Innovation in Genomics grant (OGI-119) and a CQDM-OCE EXPLORE Project (#23927) awarded to S.S.SCopyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

PY - 2017/10/19

Y1 - 2017/10/19

N2 - RING and U-box E3 ubiquitin ligases regulate diverse eukaryotic processes and have been implicated in numerous diseases, but targeting these enzymes remains a major challenge. We report the development of three ubiquitin variants (UbVs), each binding selectively to the RING or U-box domain of a distinct E3 ligase: monomeric UBE4B, phosphorylated active CBL, or dimeric XIAP. Structural and biochemical analyses revealed that UbVs specifically inhibited the activity of UBE4B or phosphorylated CBL by blocking the E2∼Ub binding site. Surprisingly, the UbV selective for dimeric XIAP formed a dimer to stimulate E3 activity by stabilizing the closed E2∼Ub conformation. We further verified the inhibitory and stimulatory functions of UbVs in cells. Our work provides a general strategy to inhibit or activate RING/U-box E3 ligases and provides a resource for the research community to modulate these enzymes.

AB - RING and U-box E3 ubiquitin ligases regulate diverse eukaryotic processes and have been implicated in numerous diseases, but targeting these enzymes remains a major challenge. We report the development of three ubiquitin variants (UbVs), each binding selectively to the RING or U-box domain of a distinct E3 ligase: monomeric UBE4B, phosphorylated active CBL, or dimeric XIAP. Structural and biochemical analyses revealed that UbVs specifically inhibited the activity of UBE4B or phosphorylated CBL by blocking the E2∼Ub binding site. Surprisingly, the UbV selective for dimeric XIAP formed a dimer to stimulate E3 activity by stabilizing the closed E2∼Ub conformation. We further verified the inhibitory and stimulatory functions of UbVs in cells. Our work provides a general strategy to inhibit or activate RING/U-box E3 ligases and provides a resource for the research community to modulate these enzymes.

KW - Drug Discovery/methods

KW - Enzyme Activators/chemistry

KW - Enzyme Inhibitors/chemistry

KW - HEK293 Cells

KW - HeLa Cells

KW - Humans

KW - Protein Multimerization/drug effects

KW - Tumor Suppressor Proteins/agonists

KW - Ubiquitin-Protein Ligase Complexes/antagonists & inhibitors

KW - Ubiquitin-Protein Ligases

KW - X-Linked Inhibitor of Apoptosis Protein/agonists

U2 - 10.1016/j.molcel.2017.09.027

DO - 10.1016/j.molcel.2017.09.027

M3 - Article

C2 - 29053960

SN - 1097-2765

VL - 68

SP - 456-470.e10

JO - Molecular Cell

JF - Molecular Cell

IS - 2

ER -

A General Strategy for Discovery of Inhibitors and Activators of RING and U-box E3 Ligases with Ubiquitin Variants

Abstract

Keywords

Access to Document

Fingerprint

Cite this