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

Research output: Contribution to journalArticlepeer-review

56 Citations (Scopus)
57 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 languageEnglish
Pages (from-to)456-470.e10
Number of pages26
JournalMolecular Cell
Volume68
Issue number2
DOIs
Publication statusPublished - 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

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