Differences in conformational dynamics within the Hsp90 chaperone family reveal mechanistic insights

The molecular chaperones of the Hsp90 family are essential in all eukaryotic cells. They assist late folding steps and maturation of many different proteins, called clients, that are not related in sequence or structure. Hsp90 interaction with its clients appears to be coupled to a series of conform...

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Main Authors:	Graf, Christian, Lee, Chung-Tien, Eva Meier-Andrejszki, L., Nguyen, Minh T. N., Mayer, Matthias P.
Format:	Online
Language:	English
Published:	Frontiers Media S.A. 2014
Online Access:	https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4428384/

id	pubmed-4428384
recordtype	oai_dc
spelling	pubmed-44283842015-05-18 Differences in conformational dynamics within the Hsp90 chaperone family reveal mechanistic insights Graf, Christian Lee, Chung-Tien Eva Meier-Andrejszki, L. Nguyen, Minh T. N. Mayer, Matthias P. Molecular Biosciences The molecular chaperones of the Hsp90 family are essential in all eukaryotic cells. They assist late folding steps and maturation of many different proteins, called clients, that are not related in sequence or structure. Hsp90 interaction with its clients appears to be coupled to a series of conformational changes. Using hydrogen exchange mass spectrometry (HX-MS) we investigated the structural dynamics of human Hsp90β (hHsp90) and yeast Hsp82 (yHsp82). We found that eukaryotic Hsp90s are much more flexible than the previously studied Escherichia coli homolog (EcHtpG) and that nucleotides induce much smaller changes. More stable conformations in yHsp82 are obtained in presence of co-chaperones. The tetratricopeptide repeat (TPR) domain protein Cpr6 causes a different amide proton protection pattern in yHsp82 than the previously studied TPR-domain protein Sti1. In the simultaneous presence of Sti1 and Cpr6, protection levels are observed that are intermediate between the Sti1 and the Cpr6 induced changes. Surprisingly, no bimodal distributions of the isotope peaks are detected, suggesting that both co-chaperones affect both protomers of the Hsp90 dimer in a similar way. The cochaperones Sba1 was found previously in the crystal structure bound to the ATP hydrolysis-competent conformation of Hsp90, which did not allow to distinguish the mode of Sba1-mediated inhibition of Hsp90's ATPase activity by stabilizing the pre- or post-hydrolysis step. Our HX-MS experiments now show that Sba1 binding leads to a protection of the ATP binding lid, suggesting that it inhibits Hsp90's ATPase activity by slowing down product release. This hypothesis was verified by a single-turnover ATPase assay. Together, our data suggest that there are much smaller energy barriers between conformational states in eukaryotic Hsp90s than in EcHtpG and that co-chaperones are necessary in addition to nucleotides to stabilize defined conformational states. Frontiers Media S.A. 2014-06-10 /pmc/articles/PMC4428384/ /pubmed/25988145 http://dx.doi.org/10.3389/fmolb.2014.00004 Text en Copyright © 2014 Graf, Lee, Eva Meier-Andrejszki, Nguyen and Mayer. http://creativecommons.org/licenses/by/3.0/ 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) or licensor 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.
repository_type	Open Access Journal
institution_category	Foreign Institution
institution	US National Center for Biotechnology Information
building	NCBI PubMed
collection	Online Access
language	English
format	Online
author	Graf, Christian Lee, Chung-Tien Eva Meier-Andrejszki, L. Nguyen, Minh T. N. Mayer, Matthias P.
spellingShingle	Graf, Christian Lee, Chung-Tien Eva Meier-Andrejszki, L. Nguyen, Minh T. N. Mayer, Matthias P. Differences in conformational dynamics within the Hsp90 chaperone family reveal mechanistic insights
author_facet	Graf, Christian Lee, Chung-Tien Eva Meier-Andrejszki, L. Nguyen, Minh T. N. Mayer, Matthias P.
author_sort	Graf, Christian
title	Differences in conformational dynamics within the Hsp90 chaperone family reveal mechanistic insights
title_short	Differences in conformational dynamics within the Hsp90 chaperone family reveal mechanistic insights
title_full	Differences in conformational dynamics within the Hsp90 chaperone family reveal mechanistic insights
title_fullStr	Differences in conformational dynamics within the Hsp90 chaperone family reveal mechanistic insights
title_full_unstemmed	Differences in conformational dynamics within the Hsp90 chaperone family reveal mechanistic insights
title_sort	differences in conformational dynamics within the hsp90 chaperone family reveal mechanistic insights
description	The molecular chaperones of the Hsp90 family are essential in all eukaryotic cells. They assist late folding steps and maturation of many different proteins, called clients, that are not related in sequence or structure. Hsp90 interaction with its clients appears to be coupled to a series of conformational changes. Using hydrogen exchange mass spectrometry (HX-MS) we investigated the structural dynamics of human Hsp90β (hHsp90) and yeast Hsp82 (yHsp82). We found that eukaryotic Hsp90s are much more flexible than the previously studied Escherichia coli homolog (EcHtpG) and that nucleotides induce much smaller changes. More stable conformations in yHsp82 are obtained in presence of co-chaperones. The tetratricopeptide repeat (TPR) domain protein Cpr6 causes a different amide proton protection pattern in yHsp82 than the previously studied TPR-domain protein Sti1. In the simultaneous presence of Sti1 and Cpr6, protection levels are observed that are intermediate between the Sti1 and the Cpr6 induced changes. Surprisingly, no bimodal distributions of the isotope peaks are detected, suggesting that both co-chaperones affect both protomers of the Hsp90 dimer in a similar way. The cochaperones Sba1 was found previously in the crystal structure bound to the ATP hydrolysis-competent conformation of Hsp90, which did not allow to distinguish the mode of Sba1-mediated inhibition of Hsp90's ATPase activity by stabilizing the pre- or post-hydrolysis step. Our HX-MS experiments now show that Sba1 binding leads to a protection of the ATP binding lid, suggesting that it inhibits Hsp90's ATPase activity by slowing down product release. This hypothesis was verified by a single-turnover ATPase assay. Together, our data suggest that there are much smaller energy barriers between conformational states in eukaryotic Hsp90s than in EcHtpG and that co-chaperones are necessary in addition to nucleotides to stabilize defined conformational states.
publisher	Frontiers Media S.A.
publishDate	2014
url	https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4428384/
_version_	1613222277123407872

Differences in conformational dynamics within the Hsp90 chaperone family reveal mechanistic insights

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