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Inhibition of SCF ubiquitin ligases by engineered ubiquitin variants that target the Cul1 binding site on the Skp1–F-box interface

Journal Article · · Proceedings of the National Academy of Sciences of the United States of America
 [1];  [2];  [1];  [2];  [1];  [3];  [1];  [4];  [1];  [5];  [1]
  1. Univ. of Toronto, ON (Canada). Terrence Donnelly Center for Cellular and Biomolecular Research. Banting and Best Dept. of Medical Research. Dept. of Molecular Genetics
  2. Mount Sinai Hospital, Toronto, ON (Canada). Lunenfeld-Tanenbaum Research Inst.
  3. Cornell Univ., Argonne, IL (United States). Dept. of Chemistry and Chemical Biology
  4. Mount Sinai Hospital, Toronto, ON (Canada). Lunenfeld-Tanenbaum Research Inst.; Univ. of Montreal, QC (Canada). Inst. of Immunology and Cancer Research
  5. Mount Sinai Hospital, Toronto, ON (Canada). Lunenfeld-Tanenbaum Research Inst.; Univ. of Toronto, ON (Canada). Dept. of Molecular Genetics
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Skp1–Cul1–F-box (SCF) E3 ligases play key roles in multiple cellular processes through ubiquitination and subsequent degradation of substrate proteins. Although Skp1 and Cul1 are invariant components of all SCF complexes, the 69 different human F-box proteins are variable substrate binding modules that determine specificity. SCF E3 ligases are activated in many cancers and inhibitors could have therapeutic potential. Here, we used phage display to develop specific ubiquitin-based inhibitors against two F-box proteins, Fbw7 and Fbw11. Unexpectedly, the ubiquitin variants bind at the interface of Skp1 and F-box proteins and inhibit ligase activity by preventing Cul1 binding to the same surface. Using structure-based design and phage display, we modified the initial inhibitors to generate broad-spectrum inhibitors that targeted many SCF ligases, or conversely, a highly specific inhibitor that discriminated between even the close homologs Fbw11 and Fbw1. We propose that most F-box proteins can be targeted by this approach for basic research and for potential cancer therapies.
Research Organization:
Univ. of Toronto, ON (Canada)
Sponsoring Organization:
Canadian Cancer Society Research Inst. (Canada); Canadian Inst. of Health Research (CIHR) (Canada); National Inst. of Health (NIH) (United States); USDOE Office of Science (SC), Basic Energy Sciences (BES) (SC-22)
Contributing Organization:
Cornell Univ., Argonne, IL (United States); Mount Sinai Hospital, Toronto, ON (Canada); Univ. of Montreal, QC (Canada)
Grant/Contract Number:
AC02-06CH11357
OSTI ID:
1247362
Journal Information:
Proceedings of the National Academy of Sciences of the United States of America, Journal Name: Proceedings of the National Academy of Sciences of the United States of America Journal Issue: 13 Vol. 113; ISSN 0027-8424
Publisher:
National Academy of Sciences, Washington, DC (United States)Copyright Statement
Country of Publication:
United States
Language:
ENGLISH

References (38)

NMRPipe: A multidimensional spectral processing system based on UNIX pipes journal November 1995
NMRPipe: A multidimensional spectral processing system based on UNIX pipes journal November 1995
[21] Phage display for selection of novel binding peptides book January 2000
[20] Processing of X-ray diffraction data collected in oscillation mode book January 1997
Control of Meiotic and Mitotic Progression by the F Box Protein β-Trcp1 In Vivo journal June 2003
Tumor Suppression by the Fbw7 Ubiquitin Ligase: Mechanisms and Opportunities journal October 2014
Ablation of Fbxw7 Eliminates Leukemia-Initiating Cells by Preventing Quiescence journal March 2013
Cand1 Promotes Assembly of New SCF Complexes through Dynamic Exchange of F Box Proteins journal March 2013
Human-Chromatin-Related Protein Interactions Identify a Demethylase Complex Required for Chromosome Segregation journal July 2014
High-affinity Human Antibodies from Phage-displayed Synthetic Fab Libraries with a Single Framework Scaffold journal July 2004
Structure of a Fbw7-Skp1-Cyclin E Complex: Multisite-Phosphorylated Substrate Recognition by SCF Ubiquitin Ligases journal April 2007
WD40 Repeat Propellers Define a Ubiquitin-Binding Domain that Regulates Turnover of F Box Proteins journal November 2010
[21] Phage display for selection of novel binding peptides book January 2000
Control of Meiotic and Mitotic Progression by the F Box Protein β-Trcp1 In Vivo journal June 2003
Insights into SCF ubiquitin ligases from the structure of the Skp1–Skp2 complex journal November 2000
Structure of the Cul1–Rbx1–Skp1–F boxSkp2 SCF ubiquitin ligase complex journal April 2002
An allosteric inhibitor of substrate recognition by the SCFCdc4 ubiquitin ligase journal June 2010
Fbxw7α- and GSK3-mediated degradation of p100 is a pro-survival mechanism in multiple myeloma journal March 2012
Roles of F-box proteins in cancer journal March 2014
The ubiquitin-specific protease USP47 is a novel β-TRCP interactor regulating cell survival journal December 2009
Cavin1 intrinsically disordered domains are essential for fuzzy electrostatic interactions and caveola formation journal February 2021
Multi-functionality of a tryptophan residue conserved in substrate-binding groove of GH19 chitinases journal January 2021
Homodimer of Two F-box Proteins βTrCP1 or βTrCP2 Binds to IκBα for Signal-dependent Ubiquitination journal January 2000
Systematic analysis and nomenclature of mammalian F-box proteins journal November 2004
Fbw7 dimerization determines the specificity and robustness of substrate degradation journal December 2013
Phaser crystallographic software journal July 2007
Coot model-building tools for molecular graphics journal November 2004
PHENIX: a comprehensive Python-based system for macromolecular structure solution journal January 2010
Phaser crystallographic software journal July 2007
Coot model-building tools for molecular graphics journal November 2004
PHENIX: a comprehensive Python-based system for macromolecular structure solution journal January 2010
A Strategy for Modulation of Enzymes in the Ubiquitin System journal January 2013
A switch between two-, three-, and four-stranded coiled coils in GCN4 leucine zipper mutants journal November 1993
Emerging therapies targeting the ubiquitin proteasome system in cancer journal January 2014
Emerging therapies targeting the ubiquitin proteasome system in cancer journal January 2014
Functional Diversity and Structural Disorder in the Human Ubiquitination Pathway journal May 2013
PHENIX: a comprehensive Python-based system for macromolecular structure solution. text January 2010
The Fbw7 and BetaTRCP E3 ubiquitin ligases and their roles in tumorigenesis journal January 2012

Cited By (14)

Structure and Function of Viral Deubiquitinating Enzymes journal November 2017
A General Strategy for Discovery of Inhibitors and Activators of RING and U-box E3 Ligases with Ubiquitin Variants journal October 2017
Emerging drug development technologies targeting ubiquitination for cancer therapeutics journal July 2019
Identification and Characterization of Mutations in Ubiquitin Required for Non-covalent Dimer Formation journal September 2019
UBB pseudogene 4 encodes functional ubiquitin variants journal March 2020
Protein Engineering in the Ubiquitin System: Tools for Discovery and Beyond journal February 2020
Proteomic identification of the oncoprotein STAT3 as a target of a novel Skp1 inhibitor journal November 2016
Specific targeting of the deubiquitinase and E3 ligase families with engineered ubiquitin variants: Gorelik and Sidhu journal November 2016
Peptides meet ubiquitin: Simple interactions regulating complex cell signaling journal September 2018
Generating Intracellular Modulators of E3 Ligases and Deubiquitinases from Phage-Displayed Ubiquitin Variant Libraries book January 2018
Humanization and directed evolution of the selenium-containing scFv phage abzyme journal January 2018
Saturation scanning of ubiquitin variants reveals a common hot spot for binding to USP2 and USP21 journal July 2016
Peptide design by optimization on a data-parameterized protein interaction landscape journal October 2018
Site-specific inhibition of the small ubiquitin-like modifier (SUMO)-conjugating enzyme Ubc9 selectively impairs SUMO chain formation journal August 2017

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Related Subjects

59 BASIC BIOLOGICAL SCIENCES
60 APPLIED LIFE SCIENCES
Cul1 affinity
Fbxw11
Fbxw7
SCF inhibitors
β-Trcp