Mechanism of ubiquitin ligation and lysine prioritization by a HECT E3
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Ubiquitination by HECT E3 enzymes regulates myriad processes, including tumor suppression, transcription, protein trafficking, and degradation. HECT E3s use a two-step mechanism to ligate ubiquitin to target proteins. The first step is guided by interactions between the catalytic HECT domain and the E2∼ubiquitin intermediate, which promote formation of a transient, thioester-bonded HECT∼ubiquitin intermediate. Here we report that the second step of ligation is mediated by a distinct catalytic architecture established by both the HECT E3 and its covalently linked ubiquitin. The structure of a chemically trapped proxy for an E3∼ubiquitin-substrate intermediate reveals three-way interactions between ubiquitin and the bilobal HECT domain orienting the E3∼ubiquitin thioester bond for ligation, and restricting the location of the substrate-binding domain to prioritize target lysines for ubiquitination. The data allow visualization of an E2-to-E3-to-substrate ubiquitin transfer cascade, and show how HECT-specific ubiquitin interactions driving multiple reactions are repurposed by a major E3 conformational change to promote ligation.
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HECT ligases bind the target then transfer ubiquitin onto themselves before transferring ubiquitin to the target.
“The second class, HECT ligases, bind the target then use an intermediate strategy of transferring ubiquitin onto themselves before transferring ubiquitin to the target112,113.”
From PROTAC targeted protein degraders: the past is prologue (2022) · cites this paper as [113]
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- Authors
- Kamadurai, H. B., Qiu, Y., Deng, A., Harrison, J. S., MacDonald, C., Actis, M., Rodrigues, P., Miller, D. J., Souphron, J., Lewis, S. M., Kurinov, I., Fujii, N., Hammel, M., Piper, R., Kuhlman, B., & Schulman, B. A.
- Affiliations
- St. Jude Children's Research Hospital (US)
- University of North Carolina at Chapel Hill (US)
- University of Iowa (US)
- Centre National de la Recherche Scientifique (FR)
- Cornell University (US)
- Lawrence Berkeley National Laboratory (US)
- Corresponding author
- Kamadurai, Hari B
- Schulman, Brenda A
- ORCID
- Venue
- eLife
- Publisher
- eLife Sciences Publications, Ltd
- Published
- 2013-08-08
- Volume
- 2
- Pages
- e00828-e00828
- ISSN
- 2050-084X (ISSN-L)
- PMC
- PMC3738095
- OpenAlex
- W2037607907
- Cited by
- 165
- References
- 60
- Citations by year
- 2026: 10 · 2025: 10 · 2024: 19 · 2023: 8 · 2022: 8 · 2021: 12 · 2020: 15 · 2019: 7 · 2018: 8 · 2017: 19 · 2016: 17 · 2015: 14 · 2014: 14 · 2013: 4
- FWCI
- 7.31
- Type
- journal article
- Language
- en
- Open access
- gold
- doi.org
- Licence
- Locations
- eLife (publishedVersion, cc-by)
- Europe PMC (PubMed Central) (publishedVersion, cc-by)
- PubMed Central (publishedVersion, cc-by)
- PubMed (publishedVersion)
- Carolina Digital Repository (University of North Carolina at Chapel Hill) (submittedVersion)
- Scholarly Commons (University of the Pacific) (submittedVersion, cc-by)
- DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
- OSTI OAI (U.S. Department of Energy Office of Scientific and Technical Information) (submittedVersion)
- UNC Libraries
- Funders
- Howard Hughes Medical Institute (045102)
- National Institutes of Health (R01GM069530, 5P30CA021765, R01GM073960, 5P41RR015301-10, 8P41 GM103403-10, 5R01 GM058202)
- ALSAC
- DOE (DE-AC02-06CH11357)
- American Heart Association
- U.S. Department of Energy (DE-AC02-06CH11357)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Ubiquitin and proteasome pathways
- Physical Sciences › Materials Science › Materials Chemistry › Enzyme Structure and Function
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Protein Degradation and Inhibitors
- Keywords
- HECT E3 ubiquitin ligase, ubiquitination, HECT domain, thioester bond, substrate-binding domain, protein degradation, protein structure
- MeSH terms
- Amino Acid Sequence, Lysine, Molecular Sequence Data, Protein Conformation, Mutagenesis, Sequence Homology, Amino Acid, Catalytic Domain, Ubiquitin, Ubiquitin-Protein Ligases
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