Characterisation of the Trichinella spiralis deubiquitinating enzyme, TsUCH37, an evolutionarily conserved proteasome interaction partner.
Trichinella spiralis is a zoonotic parasitic nematode that causes trichinellosis, a disease that has been identified on all continents except Antarctica. During chronic infection, T. spiralis larvae infect skeletal myofibres, severely disrupting their differentiation state.An activity-based probe, H...
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ftdoajarticles:oai:doaj.org/article:cb97651065db4f33a35a148683a1edc4 2023-05-15T13:51:44+02:00 Characterisation of the Trichinella spiralis deubiquitinating enzyme, TsUCH37, an evolutionarily conserved proteasome interaction partner. Rhiannon R White Sachiko Miyata Eliseo Papa Eric Spooner Kleoniki Gounaris Murray E Selkirk Katerina Artavanis-Tsakonas 2011-10-01T00:00:00Z https://doi.org/10.1371/journal.pntd.0001340 https://doaj.org/article/cb97651065db4f33a35a148683a1edc4 EN eng Public Library of Science (PLoS) http://europepmc.org/articles/PMC3186758?pdf=render https://doaj.org/toc/1935-2727 https://doaj.org/toc/1935-2735 1935-2727 1935-2735 doi:10.1371/journal.pntd.0001340 https://doaj.org/article/cb97651065db4f33a35a148683a1edc4 PLoS Neglected Tropical Diseases, Vol 5, Iss 10, p e1340 (2011) Arctic medicine. Tropical medicine RC955-962 Public aspects of medicine RA1-1270 article 2011 ftdoajarticles https://doi.org/10.1371/journal.pntd.0001340 2022-12-31T03:22:24Z Trichinella spiralis is a zoonotic parasitic nematode that causes trichinellosis, a disease that has been identified on all continents except Antarctica. During chronic infection, T. spiralis larvae infect skeletal myofibres, severely disrupting their differentiation state.An activity-based probe, HA-Ub-VME, was used to identify deubiquitinating enzyme (DUB) activity in lysate of T. spiralis L1 larvae. Results were analysed by immuno-blot and immuno-precipitation, identifying a number of potential DUBs. Immuno-precipitated proteins were subjected to LC/MS/MS, yielding peptides with sequence homology to 5 conserved human DUBs: UCH-L5, UCH-L3, HAUSP, OTU 6B and Ataxin-3. The predicted gene encoding the putative UCH-L5 homologue, TsUCH37, was cloned and recombinant protein was expressed and purified. The deubiquitinating activity of this enzyme was verified by Ub-AMC assay. Co-precipitation of recombinant TsUCH37 showed that the protein associates with putative T. spiralis proteasome components, including the yeast Rpn13 homologue ADRM1. In addition, the UCH inhibitor LDN-57444 exhibited specific inhibition of recombinant TsUCH37 and reduced the viability of cultured L1 larvae.This study reports the identification of the first T. spiralis DUB, a cysteine protease that is putatively orthologous to the human protein, hUCH-L5. Results suggest that the interaction of this protein with the proteasome has been conserved throughout evolution. We show potential for the use of inhibitor compounds to elucidate the role of UCH enzymes in T. spiralis infection and their investigation as therapeutic targets for trichinellosis. Article in Journal/Newspaper Antarc* Antarctica Arctic Directory of Open Access Journals: DOAJ Articles Arctic PLoS Neglected Tropical Diseases 5 10 e1340 |
institution |
Open Polar |
collection |
Directory of Open Access Journals: DOAJ Articles |
op_collection_id |
ftdoajarticles |
language |
English |
topic |
Arctic medicine. Tropical medicine RC955-962 Public aspects of medicine RA1-1270 |
spellingShingle |
Arctic medicine. Tropical medicine RC955-962 Public aspects of medicine RA1-1270 Rhiannon R White Sachiko Miyata Eliseo Papa Eric Spooner Kleoniki Gounaris Murray E Selkirk Katerina Artavanis-Tsakonas Characterisation of the Trichinella spiralis deubiquitinating enzyme, TsUCH37, an evolutionarily conserved proteasome interaction partner. |
topic_facet |
Arctic medicine. Tropical medicine RC955-962 Public aspects of medicine RA1-1270 |
description |
Trichinella spiralis is a zoonotic parasitic nematode that causes trichinellosis, a disease that has been identified on all continents except Antarctica. During chronic infection, T. spiralis larvae infect skeletal myofibres, severely disrupting their differentiation state.An activity-based probe, HA-Ub-VME, was used to identify deubiquitinating enzyme (DUB) activity in lysate of T. spiralis L1 larvae. Results were analysed by immuno-blot and immuno-precipitation, identifying a number of potential DUBs. Immuno-precipitated proteins were subjected to LC/MS/MS, yielding peptides with sequence homology to 5 conserved human DUBs: UCH-L5, UCH-L3, HAUSP, OTU 6B and Ataxin-3. The predicted gene encoding the putative UCH-L5 homologue, TsUCH37, was cloned and recombinant protein was expressed and purified. The deubiquitinating activity of this enzyme was verified by Ub-AMC assay. Co-precipitation of recombinant TsUCH37 showed that the protein associates with putative T. spiralis proteasome components, including the yeast Rpn13 homologue ADRM1. In addition, the UCH inhibitor LDN-57444 exhibited specific inhibition of recombinant TsUCH37 and reduced the viability of cultured L1 larvae.This study reports the identification of the first T. spiralis DUB, a cysteine protease that is putatively orthologous to the human protein, hUCH-L5. Results suggest that the interaction of this protein with the proteasome has been conserved throughout evolution. We show potential for the use of inhibitor compounds to elucidate the role of UCH enzymes in T. spiralis infection and their investigation as therapeutic targets for trichinellosis. |
format |
Article in Journal/Newspaper |
author |
Rhiannon R White Sachiko Miyata Eliseo Papa Eric Spooner Kleoniki Gounaris Murray E Selkirk Katerina Artavanis-Tsakonas |
author_facet |
Rhiannon R White Sachiko Miyata Eliseo Papa Eric Spooner Kleoniki Gounaris Murray E Selkirk Katerina Artavanis-Tsakonas |
author_sort |
Rhiannon R White |
title |
Characterisation of the Trichinella spiralis deubiquitinating enzyme, TsUCH37, an evolutionarily conserved proteasome interaction partner. |
title_short |
Characterisation of the Trichinella spiralis deubiquitinating enzyme, TsUCH37, an evolutionarily conserved proteasome interaction partner. |
title_full |
Characterisation of the Trichinella spiralis deubiquitinating enzyme, TsUCH37, an evolutionarily conserved proteasome interaction partner. |
title_fullStr |
Characterisation of the Trichinella spiralis deubiquitinating enzyme, TsUCH37, an evolutionarily conserved proteasome interaction partner. |
title_full_unstemmed |
Characterisation of the Trichinella spiralis deubiquitinating enzyme, TsUCH37, an evolutionarily conserved proteasome interaction partner. |
title_sort |
characterisation of the trichinella spiralis deubiquitinating enzyme, tsuch37, an evolutionarily conserved proteasome interaction partner. |
publisher |
Public Library of Science (PLoS) |
publishDate |
2011 |
url |
https://doi.org/10.1371/journal.pntd.0001340 https://doaj.org/article/cb97651065db4f33a35a148683a1edc4 |
geographic |
Arctic |
geographic_facet |
Arctic |
genre |
Antarc* Antarctica Arctic |
genre_facet |
Antarc* Antarctica Arctic |
op_source |
PLoS Neglected Tropical Diseases, Vol 5, Iss 10, p e1340 (2011) |
op_relation |
http://europepmc.org/articles/PMC3186758?pdf=render https://doaj.org/toc/1935-2727 https://doaj.org/toc/1935-2735 1935-2727 1935-2735 doi:10.1371/journal.pntd.0001340 https://doaj.org/article/cb97651065db4f33a35a148683a1edc4 |
op_doi |
https://doi.org/10.1371/journal.pntd.0001340 |
container_title |
PLoS Neglected Tropical Diseases |
container_volume |
5 |
container_issue |
10 |
container_start_page |
e1340 |
_version_ |
1766255760977690624 |