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Updated: May 11, 2026
06:06
In Vitro Analysis of E3 Ubiquitin Ligase Function
Published on: May 14, 2021
A structurally unique E2-binding domain activates ubiquitination by the ERAD E2, Ubc7p, through multiple mechanisms.
Meredith B Metzger1, Yu-He Liang, Ranabir Das
1Laboratory of Protein Dynamics and Signaling, Center for Cancer Research, National Cancer Institute, Frederick, MD 21702-1201, USA.
Molecular Cell
|May 14, 2013
Summary
Cue1p
Area of Science:
- Biochemistry
- Molecular Biology
- Cell Biology
Background:
- Cue1p is crucial for yeast endoplasmic reticulum (ER)-associated degradation (ERAD) E3 ligase complexes.
- It binds the ERAD ubiquitin-conjugating enzyme (E2), Ubc7p, to the ER and activates it through unknown mechanisms.
Purpose of the Study:
- To determine the crystal structure of the Ubc7p-binding region (U7BR) of Cue1p with Ubc7p.
- To elucidate the mechanisms by which Cue1p activates Ubc7p and stimulates ERAD.
Main Methods:
- Crystal structure determination of the Cue1p U7BR-Ubc7p complex.
- Biochemical assays to assess ubiquitin transfer and E2 charging.
Main Results:
- The U7BR adopts a unique three-α-helix structure that binds to the backside of Ubc7p.
- Essential residues for Ubc7p binding are also critical for Ubc7p activation and ERAD.
- The U7BR stimulates both RING-dependent and RING-independent ubiquitin transfer.
- The U7BR enhances E1-mediated Ubc7p charging with ubiquitin.
Conclusions:
- Cue1p's U7BR directly binds and activates Ubc7p, enhancing ubiquitin transfer and E1 charging.
- This provides mechanistic insights into how E3 complex components stimulate ubiquitination for ERAD.
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