Cargando…
Application of fluorescence correlation spectroscopy to investigate the dynamics of a ribosome-associated trigger factor in Escherichia coli
Co-translational protein folding is one of the central topics in molecular biology. In Escherichia coli, trigger factor (TF) is a primary chaperone that facilitates co-translational folding by directly interacting with nascent polypeptide chains on translating ribosomes. In this study, we applied fl...
Autores principales: | , , , , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Frontiers Media S.A.
2022
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9452904/ https://www.ncbi.nlm.nih.gov/pubmed/36090041 http://dx.doi.org/10.3389/fmolb.2022.891128 |
_version_ | 1784785020544614400 |
---|---|
author | Niwa, Tatsuya Nakazawa, Koki Hoshi, Kensuke Tadakuma, Hisashi Ito, Koichi Taguchi, Hideki |
author_facet | Niwa, Tatsuya Nakazawa, Koki Hoshi, Kensuke Tadakuma, Hisashi Ito, Koichi Taguchi, Hideki |
author_sort | Niwa, Tatsuya |
collection | PubMed |
description | Co-translational protein folding is one of the central topics in molecular biology. In Escherichia coli, trigger factor (TF) is a primary chaperone that facilitates co-translational folding by directly interacting with nascent polypeptide chains on translating ribosomes. In this study, we applied fluorescence correlation spectroscopy (FCS), which can analyze the diffusion properties of fluorescent molecules by measuring the fluctuations of the fluorescent intensity, to investigate the interaction between TF and a nascent chain on translating ribosomes both in vitro and in vivo. The FCS analysis with a reconstituted cell-free translation system revealed that the interaction of fluorescently labeled TF with a nascent chain depended on the emergence of the nascent chain from the ribosome exit tunnel, and this interaction was not inhibited by excess amounts of other chaperones. Furthermore, the translation-dependent interaction between GFP-fused TFs and nascent chains was also observed in living E. coli cells. The FCS-based approach established here could be an effective method to investigate the dynamics of other ribosome-associated chaperones besides TF. |
format | Online Article Text |
id | pubmed-9452904 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2022 |
publisher | Frontiers Media S.A. |
record_format | MEDLINE/PubMed |
spelling | pubmed-94529042022-09-09 Application of fluorescence correlation spectroscopy to investigate the dynamics of a ribosome-associated trigger factor in Escherichia coli Niwa, Tatsuya Nakazawa, Koki Hoshi, Kensuke Tadakuma, Hisashi Ito, Koichi Taguchi, Hideki Front Mol Biosci Molecular Biosciences Co-translational protein folding is one of the central topics in molecular biology. In Escherichia coli, trigger factor (TF) is a primary chaperone that facilitates co-translational folding by directly interacting with nascent polypeptide chains on translating ribosomes. In this study, we applied fluorescence correlation spectroscopy (FCS), which can analyze the diffusion properties of fluorescent molecules by measuring the fluctuations of the fluorescent intensity, to investigate the interaction between TF and a nascent chain on translating ribosomes both in vitro and in vivo. The FCS analysis with a reconstituted cell-free translation system revealed that the interaction of fluorescently labeled TF with a nascent chain depended on the emergence of the nascent chain from the ribosome exit tunnel, and this interaction was not inhibited by excess amounts of other chaperones. Furthermore, the translation-dependent interaction between GFP-fused TFs and nascent chains was also observed in living E. coli cells. The FCS-based approach established here could be an effective method to investigate the dynamics of other ribosome-associated chaperones besides TF. Frontiers Media S.A. 2022-08-25 /pmc/articles/PMC9452904/ /pubmed/36090041 http://dx.doi.org/10.3389/fmolb.2022.891128 Text en Copyright © 2022 Niwa, Nakazawa, Hoshi, Tadakuma, Ito and Taguchi. https://creativecommons.org/licenses/by/4.0/This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. |
spellingShingle | Molecular Biosciences Niwa, Tatsuya Nakazawa, Koki Hoshi, Kensuke Tadakuma, Hisashi Ito, Koichi Taguchi, Hideki Application of fluorescence correlation spectroscopy to investigate the dynamics of a ribosome-associated trigger factor in Escherichia coli |
title | Application of fluorescence correlation spectroscopy to investigate the dynamics of a ribosome-associated trigger factor in Escherichia coli
|
title_full | Application of fluorescence correlation spectroscopy to investigate the dynamics of a ribosome-associated trigger factor in Escherichia coli
|
title_fullStr | Application of fluorescence correlation spectroscopy to investigate the dynamics of a ribosome-associated trigger factor in Escherichia coli
|
title_full_unstemmed | Application of fluorescence correlation spectroscopy to investigate the dynamics of a ribosome-associated trigger factor in Escherichia coli
|
title_short | Application of fluorescence correlation spectroscopy to investigate the dynamics of a ribosome-associated trigger factor in Escherichia coli
|
title_sort | application of fluorescence correlation spectroscopy to investigate the dynamics of a ribosome-associated trigger factor in escherichia coli |
topic | Molecular Biosciences |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9452904/ https://www.ncbi.nlm.nih.gov/pubmed/36090041 http://dx.doi.org/10.3389/fmolb.2022.891128 |
work_keys_str_mv | AT niwatatsuya applicationoffluorescencecorrelationspectroscopytoinvestigatethedynamicsofaribosomeassociatedtriggerfactorinescherichiacoli AT nakazawakoki applicationoffluorescencecorrelationspectroscopytoinvestigatethedynamicsofaribosomeassociatedtriggerfactorinescherichiacoli AT hoshikensuke applicationoffluorescencecorrelationspectroscopytoinvestigatethedynamicsofaribosomeassociatedtriggerfactorinescherichiacoli AT tadakumahisashi applicationoffluorescencecorrelationspectroscopytoinvestigatethedynamicsofaribosomeassociatedtriggerfactorinescherichiacoli AT itokoichi applicationoffluorescencecorrelationspectroscopytoinvestigatethedynamicsofaribosomeassociatedtriggerfactorinescherichiacoli AT taguchihideki applicationoffluorescencecorrelationspectroscopytoinvestigatethedynamicsofaribosomeassociatedtriggerfactorinescherichiacoli |