Mind the metal: a fragment library-derived zinc impurity binds the E2 ubiquitin-conjugating enzyme Ube2T and induces structural rearrangements

Francesca Morreale; Andrea Testa; Viduth Chaugule; Alessio Bortoluzzi; Alessio Ciulli; Helen Walden

doi:10.1021/acs.jmedchem.7b01071

Mind the metal: a fragment library-derived zinc impurity binds the E2 ubiquitin-conjugating enzyme Ube2T and induces structural rearrangements

Francesca Morreale, Andrea Testa, Viduth Chaugule, Alessio Bortoluzzi, Alessio Ciulli, Helen Walden (Lead / Corresponding author)

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Abstract

Efforts to develop inhibitors, activators and effectors of biological reactions using small molecule libraries are often hampered by interference compounds, artifacts and false positives that permeate the pool of initial hits. Here we report the discovery of a promising initial hit compound targeting the Fanconi anemia ubiquitin-conjugating enzyme Ube2T and describe its biophysical and biochemical characterization. Analysis of the co-crystal structure led to the identification of a contaminating zinc ion as solely responsible for the observed effects. Zinc binding to the active site cysteine induces a domain swap in Ube2T that leads to cyclic trimerization organized in an open ended linear assembly. Our study serves as a cautionary tale for screening small molecule libraries and provides insights into the structural plasticity of ubiquitin-conjugating enzymes

Original language	English
Pages (from-to)	8183-8191
Number of pages	9
Journal	Journal of Medicinal Chemistry
Volume	60
Issue number	19
Early online date	21 Sept 2017
DOIs	https://doi.org/10.1021/acs.jmedchem.7b01071
Publication status	Published - 12 Oct 2017

Keywords

Ubiquitin-conjugating enzymes
domain swap
false positives
metal-induced oligomerization
ubiquitination
X-ray crystallography

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10.1021/acs.jmedchem.7b01071Licence: CC BY

Final Published Version
© 2017 American Chemical Society. This is an open access article published under a Creative Commons Attribution (CC-BY) License, which permits unrestricted use, distribution and reproduction in any medium, provided the author and source are cited.
Final published version, 5.02 MBLicence: CC BY

Regulation of DNA Interstrand Crosslink Repaid by Ubiquitin (ICLUB) (ERC Consolidator)
Walden, H. (Investigator)
COMMISSION OF THE EUROPEAN COMMUNITIES
1/09/16 → 31/08/22
Project: Research
DrugE3CRL's: Probing Druggability of Multisubunit Complexes: E3 Cullin RING Ligases (ERC Starting Grant)
Ciulli, A. (Investigator)
COMMISSION OF THE EUROPEAN COMMUNITIES
1/05/13 → 30/04/18
Project: Research
Dissecting and Exploiting Molecular Recognition at Protein-Protein Interfaces (David Phillips Fellowship)
Ciulli, A. (Investigator)
Biotechnology and Biological Sciences Research Council
8/04/13 → 7/07/15
Project: Research

Ciulli, Alessio
- Centre for Targeted Protein Degradation - Professor of Chemical and Structural Biology
Person: Academic

Cite this

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title = "Mind the metal: a fragment library-derived zinc impurity binds the E2 ubiquitin-conjugating enzyme Ube2T and induces structural rearrangements",

abstract = "Efforts to develop inhibitors, activators and effectors of biological reactions using small molecule libraries are often hampered by interference compounds, artifacts and false positives that permeate the pool of initial hits. Here we report the discovery of a promising initial hit compound targeting the Fanconi anemia ubiquitin-conjugating enzyme Ube2T and describe its biophysical and biochemical characterization. Analysis of the co-crystal structure led to the identification of a contaminating zinc ion as solely responsible for the observed effects. Zinc binding to the active site cysteine induces a domain swap in Ube2T that leads to cyclic trimerization organized in an open ended linear assembly. Our study serves as a cautionary tale for screening small molecule libraries and provides insights into the structural plasticity of ubiquitin-conjugating enzymes",

keywords = "Ubiquitin-conjugating enzymes, domain swap, false positives, metal-induced oligomerization, ubiquitination, X-ray crystallography",

author = "Francesca Morreale and Andrea Testa and Viduth Chaugule and Alessio Bortoluzzi and Alessio Ciulli and Helen Walden",

note = "This work was supported by the Medical Research Council (MRC grant number MC_UU_12016/12); the EMBO Young Investigator Programme to H.W.; the European Research Council (ERC-2015-CoG-681582 ICLUb consolidator grant to H.W.; ERC-2012-StG- 311460 DrugE3CRLs starting grant to A.C.); the UK Biotechnology and Biological Sciences Research Council (BBSRC BB/G023123/2 David Phillips Fellowship to A.C.); and the Wellcome Trust (strategic awards 100476/Z/12/Z for biophysics and drug discovery and 094090/Z/10/Z for structural biology and X-ray crystallography to BCDD). We thank the Diamond Light Source for beamtime (BAG proposal MX10071) and beamline support at beamline I04-1.",

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doi = "10.1021/acs.jmedchem.7b01071",

language = "English",

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journal = "Journal of Medicinal Chemistry",

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T2 - a fragment library-derived zinc impurity binds the E2 ubiquitin-conjugating enzyme Ube2T and induces structural rearrangements

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AU - Testa, Andrea

AU - Chaugule, Viduth

AU - Bortoluzzi, Alessio

AU - Ciulli, Alessio

AU - Walden, Helen

N1 - This work was supported by the Medical Research Council (MRC grant number MC_UU_12016/12); the EMBO Young Investigator Programme to H.W.; the European Research Council (ERC-2015-CoG-681582 ICLUb consolidator grant to H.W.; ERC-2012-StG- 311460 DrugE3CRLs starting grant to A.C.); the UK Biotechnology and Biological Sciences Research Council (BBSRC BB/G023123/2 David Phillips Fellowship to A.C.); and the Wellcome Trust (strategic awards 100476/Z/12/Z for biophysics and drug discovery and 094090/Z/10/Z for structural biology and X-ray crystallography to BCDD). We thank the Diamond Light Source for beamtime (BAG proposal MX10071) and beamline support at beamline I04-1.

PY - 2017/10/12

Y1 - 2017/10/12

N2 - Efforts to develop inhibitors, activators and effectors of biological reactions using small molecule libraries are often hampered by interference compounds, artifacts and false positives that permeate the pool of initial hits. Here we report the discovery of a promising initial hit compound targeting the Fanconi anemia ubiquitin-conjugating enzyme Ube2T and describe its biophysical and biochemical characterization. Analysis of the co-crystal structure led to the identification of a contaminating zinc ion as solely responsible for the observed effects. Zinc binding to the active site cysteine induces a domain swap in Ube2T that leads to cyclic trimerization organized in an open ended linear assembly. Our study serves as a cautionary tale for screening small molecule libraries and provides insights into the structural plasticity of ubiquitin-conjugating enzymes

AB - Efforts to develop inhibitors, activators and effectors of biological reactions using small molecule libraries are often hampered by interference compounds, artifacts and false positives that permeate the pool of initial hits. Here we report the discovery of a promising initial hit compound targeting the Fanconi anemia ubiquitin-conjugating enzyme Ube2T and describe its biophysical and biochemical characterization. Analysis of the co-crystal structure led to the identification of a contaminating zinc ion as solely responsible for the observed effects. Zinc binding to the active site cysteine induces a domain swap in Ube2T that leads to cyclic trimerization organized in an open ended linear assembly. Our study serves as a cautionary tale for screening small molecule libraries and provides insights into the structural plasticity of ubiquitin-conjugating enzymes

KW - Ubiquitin-conjugating enzymes

KW - domain swap

KW - false positives

KW - metal-induced oligomerization

KW - ubiquitination

KW - X-ray crystallography

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M3 - Article

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JO - Journal of Medicinal Chemistry

JF - Journal of Medicinal Chemistry

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ER -

Mind the metal: a fragment library-derived zinc impurity binds the E2 ubiquitin-conjugating enzyme Ube2T and induces structural rearrangements

Abstract

Keywords

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Projects

Regulation of DNA Interstrand Crosslink Repaid by Ubiquitin (ICLUB) (ERC Consolidator)

DrugE3CRL's: Probing Druggability of Multisubunit Complexes: E3 Cullin RING Ligases (ERC Starting Grant)

Dissecting and Exploiting Molecular Recognition at Protein-Protein Interfaces (David Phillips Fellowship)

Profiles

Ciulli, Alessio

Cite this