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Methods to assess helicase and translocation activities of human nuclear RNA exosome and RNA adaptor complexes
The nuclear RNA exosome collaborates with the MTR4 helicase and RNA adaptor complexes to process, surveil, and degrade RNA. Here we outline methods to characterize RNA translocation and strand displacement by exosome-associated helicases and adaptor complexes using fluorescence-based strand displace...
Autores principales: | , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
2022
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Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9382703/ https://www.ncbi.nlm.nih.gov/pubmed/35965016 http://dx.doi.org/10.1016/bs.mie.2022.03.060 |
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author | Puno, M. Rhyan Lima, Christopher D. |
author_facet | Puno, M. Rhyan Lima, Christopher D. |
author_sort | Puno, M. Rhyan |
collection | PubMed |
description | The nuclear RNA exosome collaborates with the MTR4 helicase and RNA adaptor complexes to process, surveil, and degrade RNA. Here we outline methods to characterize RNA translocation and strand displacement by exosome-associated helicases and adaptor complexes using fluorescence-based strand displacement assays. The design and preparation of substrates suitable for analysis of helicase and decay activities of reconstituted MTR4–exosome complexes are described. To aid structural and biophysical studies, we present strategies for engineering substrates that can stall helicases during translocation, providing a means to capture snapshots of interactions and molecular steps involved in substrate translocation and delivery to the exosome. |
format | Online Article Text |
id | pubmed-9382703 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2022 |
record_format | MEDLINE/PubMed |
spelling | pubmed-93827032022-08-17 Methods to assess helicase and translocation activities of human nuclear RNA exosome and RNA adaptor complexes Puno, M. Rhyan Lima, Christopher D. Methods Enzymol Article The nuclear RNA exosome collaborates with the MTR4 helicase and RNA adaptor complexes to process, surveil, and degrade RNA. Here we outline methods to characterize RNA translocation and strand displacement by exosome-associated helicases and adaptor complexes using fluorescence-based strand displacement assays. The design and preparation of substrates suitable for analysis of helicase and decay activities of reconstituted MTR4–exosome complexes are described. To aid structural and biophysical studies, we present strategies for engineering substrates that can stall helicases during translocation, providing a means to capture snapshots of interactions and molecular steps involved in substrate translocation and delivery to the exosome. 2022 2022-04-27 /pmc/articles/PMC9382703/ /pubmed/35965016 http://dx.doi.org/10.1016/bs.mie.2022.03.060 Text en https://creativecommons.org/licenses/by/4.0/This work is licensed under a Creative Commons Attribution 4.0 International License, which allows reusers to distribute, remix, adapt, and build upon the material in any medium or format, so long as attribution is given to the creator. The license allows for commercial use. |
spellingShingle | Article Puno, M. Rhyan Lima, Christopher D. Methods to assess helicase and translocation activities of human nuclear RNA exosome and RNA adaptor complexes |
title | Methods to assess helicase and translocation activities of human nuclear RNA exosome and RNA adaptor complexes |
title_full | Methods to assess helicase and translocation activities of human nuclear RNA exosome and RNA adaptor complexes |
title_fullStr | Methods to assess helicase and translocation activities of human nuclear RNA exosome and RNA adaptor complexes |
title_full_unstemmed | Methods to assess helicase and translocation activities of human nuclear RNA exosome and RNA adaptor complexes |
title_short | Methods to assess helicase and translocation activities of human nuclear RNA exosome and RNA adaptor complexes |
title_sort | methods to assess helicase and translocation activities of human nuclear rna exosome and rna adaptor complexes |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9382703/ https://www.ncbi.nlm.nih.gov/pubmed/35965016 http://dx.doi.org/10.1016/bs.mie.2022.03.060 |
work_keys_str_mv | AT punomrhyan methodstoassesshelicaseandtranslocationactivitiesofhumannuclearrnaexosomeandrnaadaptorcomplexes AT limachristopherd methodstoassesshelicaseandtranslocationactivitiesofhumannuclearrnaexosomeandrnaadaptorcomplexes |