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Activity‐Based Probes for HECT E3 Ubiquitin Ligases

Activity‐based probes (ABPs) have been used to dissect the biochemical/structural properties and cellular functions of deubiquitinases. However, their utility in studying cysteine‐based E3 ubiquitin ligases has been limited. In this study, we evaluate the use of ubiquitin‐ABPs (Ub‐VME and Ub‐PA) and...

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Autores principales: Byrne, Robert, Mund, Thomas, Licchesi, Julien D. F.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: John Wiley and Sons Inc. 2017
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5575557/
https://www.ncbi.nlm.nih.gov/pubmed/28425671
http://dx.doi.org/10.1002/cbic.201700006
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author Byrne, Robert
Mund, Thomas
Licchesi, Julien D. F.
author_facet Byrne, Robert
Mund, Thomas
Licchesi, Julien D. F.
author_sort Byrne, Robert
collection PubMed
description Activity‐based probes (ABPs) have been used to dissect the biochemical/structural properties and cellular functions of deubiquitinases. However, their utility in studying cysteine‐based E3 ubiquitin ligases has been limited. In this study, we evaluate the use of ubiquitin‐ABPs (Ub‐VME and Ub‐PA) and a novel set of E2–Ub‐ABPs on a panel of HECT E3 ubiquitin ligases. Our in vitro data show that ubiquitin‐ABPs can label HECT domains. We also provide the first evidence that, in addition to the RBR E3 ubiquitin ligase Parkin, E2–Ub‐ABPs can also label the catalytic HECT domains of NEDD4, UBE3C, and HECTD1. Importantly, the endogenous proteasomal E3 ligase UBE3C was also successfully labelled by Ub‐PA and His‐UBE2D2–Ub‐ABP in lysate of cells grown under basal conditions. Our findings provide novel insights into the use of ABPs for the study of HECT E3 ubiquitin ligases.
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spelling pubmed-55755572017-09-18 Activity‐Based Probes for HECT E3 Ubiquitin Ligases Byrne, Robert Mund, Thomas Licchesi, Julien D. F. Chembiochem Full Papers Activity‐based probes (ABPs) have been used to dissect the biochemical/structural properties and cellular functions of deubiquitinases. However, their utility in studying cysteine‐based E3 ubiquitin ligases has been limited. In this study, we evaluate the use of ubiquitin‐ABPs (Ub‐VME and Ub‐PA) and a novel set of E2–Ub‐ABPs on a panel of HECT E3 ubiquitin ligases. Our in vitro data show that ubiquitin‐ABPs can label HECT domains. We also provide the first evidence that, in addition to the RBR E3 ubiquitin ligase Parkin, E2–Ub‐ABPs can also label the catalytic HECT domains of NEDD4, UBE3C, and HECTD1. Importantly, the endogenous proteasomal E3 ligase UBE3C was also successfully labelled by Ub‐PA and His‐UBE2D2–Ub‐ABP in lysate of cells grown under basal conditions. Our findings provide novel insights into the use of ABPs for the study of HECT E3 ubiquitin ligases. John Wiley and Sons Inc. 2017-06-28 2017-07-18 /pmc/articles/PMC5575557/ /pubmed/28425671 http://dx.doi.org/10.1002/cbic.201700006 Text en © 2017 The Authors. Published by Wiley-VCH Verlag GmbH & Co. KGaA. This is an open access article under the terms of the Creative Commons Attribution (http://creativecommons.org/licenses/by/4.0/) License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.
spellingShingle Full Papers
Byrne, Robert
Mund, Thomas
Licchesi, Julien D. F.
Activity‐Based Probes for HECT E3 Ubiquitin Ligases
title Activity‐Based Probes for HECT E3 Ubiquitin Ligases
title_full Activity‐Based Probes for HECT E3 Ubiquitin Ligases
title_fullStr Activity‐Based Probes for HECT E3 Ubiquitin Ligases
title_full_unstemmed Activity‐Based Probes for HECT E3 Ubiquitin Ligases
title_short Activity‐Based Probes for HECT E3 Ubiquitin Ligases
title_sort activity‐based probes for hect e3 ubiquitin ligases
topic Full Papers
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5575557/
https://www.ncbi.nlm.nih.gov/pubmed/28425671
http://dx.doi.org/10.1002/cbic.201700006
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