Fusion protein analysis reveals the precise regulation between Hsp70 and Hsp100 during protein disaggregation

Abstract ClpB, a bacterial Hsp100, is a ring-shaped AAA+ chaperone that can reactivate aggregated proteins in cooperation with DnaK, a bacterial Hsp70, and its co-factors. ClpB subunits comprise two AAA+ modules with an interstitial rod-shaped M-domain. The M-domain regulates ClpB ATPase activity an...

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Autores principales:	Sayaka Hayashi, Yosuke Nakazaki, Kei Kagii, Hiromi Imamura, Yo-hei Watanabe
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2017
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/f9c750ace9384970858dcb9e1fcf681f
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spelling	oai:doaj.org-article:f9c750ace9384970858dcb9e1fcf681f2021-12-02T16:06:44ZFusion protein analysis reveals the precise regulation between Hsp70 and Hsp100 during protein disaggregation10.1038/s41598-017-08917-82045-2322https://doaj.org/article/f9c750ace9384970858dcb9e1fcf681f2017-08-01T00:00:00Zhttps://doi.org/10.1038/s41598-017-08917-8https://doaj.org/toc/2045-2322Abstract ClpB, a bacterial Hsp100, is a ring-shaped AAA+ chaperone that can reactivate aggregated proteins in cooperation with DnaK, a bacterial Hsp70, and its co-factors. ClpB subunits comprise two AAA+ modules with an interstitial rod-shaped M-domain. The M-domain regulates ClpB ATPase activity and interacts directly with the DnaK nucleotide-binding domain (NBD). Here, to clarify how these functions contribute to the disaggregation process, we constructed ClpB, DnaK, and aggregated YFP fusion proteins in various combinations. Notably, i) DnaK activates ClpB only when the DnaK substrate-binding domain (SBD) is in the closed conformation, affording high DnaK-peptide affinity; ii) although NBD alone can activate ClpB, SBD is required for disaggregation; and iii) tethering aggregated proteins to the activated ClpB obviates SBD requirements. These results indicate that DnaK activates ClpB only when the SBD tightly holds aggregated proteins adjacent to ClpB for effective disaggregation.Sayaka HayashiYosuke NakazakiKei KagiiHiromi ImamuraYo-hei WatanabeNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 7, Iss 1, Pp 1-14 (2017)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Sayaka Hayashi Yosuke Nakazaki Kei Kagii Hiromi Imamura Yo-hei Watanabe Fusion protein analysis reveals the precise regulation between Hsp70 and Hsp100 during protein disaggregation
description	Abstract ClpB, a bacterial Hsp100, is a ring-shaped AAA+ chaperone that can reactivate aggregated proteins in cooperation with DnaK, a bacterial Hsp70, and its co-factors. ClpB subunits comprise two AAA+ modules with an interstitial rod-shaped M-domain. The M-domain regulates ClpB ATPase activity and interacts directly with the DnaK nucleotide-binding domain (NBD). Here, to clarify how these functions contribute to the disaggregation process, we constructed ClpB, DnaK, and aggregated YFP fusion proteins in various combinations. Notably, i) DnaK activates ClpB only when the DnaK substrate-binding domain (SBD) is in the closed conformation, affording high DnaK-peptide affinity; ii) although NBD alone can activate ClpB, SBD is required for disaggregation; and iii) tethering aggregated proteins to the activated ClpB obviates SBD requirements. These results indicate that DnaK activates ClpB only when the SBD tightly holds aggregated proteins adjacent to ClpB for effective disaggregation.
format	article
author	Sayaka Hayashi Yosuke Nakazaki Kei Kagii Hiromi Imamura Yo-hei Watanabe
author_facet	Sayaka Hayashi Yosuke Nakazaki Kei Kagii Hiromi Imamura Yo-hei Watanabe
author_sort	Sayaka Hayashi
title	Fusion protein analysis reveals the precise regulation between Hsp70 and Hsp100 during protein disaggregation
title_short	Fusion protein analysis reveals the precise regulation between Hsp70 and Hsp100 during protein disaggregation
title_full	Fusion protein analysis reveals the precise regulation between Hsp70 and Hsp100 during protein disaggregation
title_fullStr	Fusion protein analysis reveals the precise regulation between Hsp70 and Hsp100 during protein disaggregation
title_full_unstemmed	Fusion protein analysis reveals the precise regulation between Hsp70 and Hsp100 during protein disaggregation
title_sort	fusion protein analysis reveals the precise regulation between hsp70 and hsp100 during protein disaggregation
publisher	Nature Portfolio
publishDate	2017
url	https://doaj.org/article/f9c750ace9384970858dcb9e1fcf681f
work_keys_str_mv	AT sayakahayashi fusionproteinanalysisrevealsthepreciseregulationbetweenhsp70andhsp100duringproteindisaggregation AT yosukenakazaki fusionproteinanalysisrevealsthepreciseregulationbetweenhsp70andhsp100duringproteindisaggregation AT keikagii fusionproteinanalysisrevealsthepreciseregulationbetweenhsp70andhsp100duringproteindisaggregation AT hiromiimamura fusionproteinanalysisrevealsthepreciseregulationbetweenhsp70andhsp100duringproteindisaggregation AT yoheiwatanabe fusionproteinanalysisrevealsthepreciseregulationbetweenhsp70andhsp100duringproteindisaggregation
_version_	1718384880452108288

Fusion protein analysis reveals the precise regulation between Hsp70 and Hsp100 during protein disaggregation

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