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Covalent Inhibition of Ubc13 Affects Ubiquitin Signaling and Reveals Active Site Elements Important for Targeting

Curtis D Hodge1, Ross A Edwards1, Craig J Markin1

  • 1†Department of Biochemistry, University of Alberta, Edmonton, Alberta, Canada T6G 2H7.

ACS Chemical Biology
|April 25, 2015
PubMed

Insights

Two inhibitors, NSC697923 and BAY 11-7082, target the Ubc13 enzyme by forming covalent adducts. This research reveals unique structural features of Ubc13, crucial for developing specific inhibitors.

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Structural Biology

Background:

  • Ubc13 is an E2 ubiquitin conjugating enzyme vital for DNA damage and NF-κB signaling pathways.
  • Dysregulation of these pathways is implicated in various diseases, making Ubc13 a potential therapeutic target.

Purpose of the Study:

  • To elucidate the structural basis of Ubc13 inhibition by NSC697923 and BAY 11-7082.
  • To validate Ubc13 as the primary target for NSC697923's cellular effects.

Main Methods:

  • X-ray crystallography to determine the structures of Ubc13-inhibitor complexes.
  • Site-directed mutagenesis to create a Ubc13-resistant mutant.
  • Cellular assays to assess the impact of Ubc13 inhibition on DNA damage and NF-κB signaling.

Main Results:

  • The structures reveal that both NSC697923 and BAY 11-7082 form covalent adducts with Ubc13's active site cysteine via Michael addition.
  • These inhibitors bind within a unique groove specific to Ubc13.
  • A Ubc13 mutant resistant to NSC697923 confirmed that Ubc13 is the main target responsible for inhibiting DNA damage and NF-κB signaling.

Conclusions:

  • The unique structural features of Ubc13's active site provide a foundation for designing highly specific Ubc13 inhibitors.
  • Targeting Ubc13 offers a promising strategy for modulating DNA damage and NF-κB signaling pathways.

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