Header

UZH-Logo

Maintenance Infos

The kinetic parameters and energy cost of the Hsp70 chaperone as a polypeptide unfoldase


Sharma, S K; De los Rios, P; Christen, P; Lustig, A; Goloubinoff, P (2010). The kinetic parameters and energy cost of the Hsp70 chaperone as a polypeptide unfoldase. Nature chemical biology, 6(12):914-920.

Abstract

Hsp70-Hsp40-NEF and possibly Hsp100 are the only known molecular chaperones that can use the energy of ATP to convert stably pre-aggregated polypeptides into natively refolded proteins. However, the kinetic parameters and ATP costs have remained elusive because refolding reactions have only been successful with a molar excess of chaperones over their polypeptide substrates. Here we describe a stable, misfolded luciferase species that can be efficiently renatured by substoichiometric amounts of bacterial Hsp70-Hsp40-NEF. The reactivation rates increased with substrate concentration and followed saturation kinetics, thus allowing the determination of apparent V(max)' and K(m)' values for a chaperone-mediated renaturation reaction for the first time. Under the in vitro conditions used, one Hsp70 molecule consumed five ATPs to effectively unfold a single misfolded protein into an intermediate that, upon chaperone dissociation, spontaneously refolded to the native state, a process with an ATP cost a thousand times lower than expected for protein degradation and resynthesis.

Abstract

Hsp70-Hsp40-NEF and possibly Hsp100 are the only known molecular chaperones that can use the energy of ATP to convert stably pre-aggregated polypeptides into natively refolded proteins. However, the kinetic parameters and ATP costs have remained elusive because refolding reactions have only been successful with a molar excess of chaperones over their polypeptide substrates. Here we describe a stable, misfolded luciferase species that can be efficiently renatured by substoichiometric amounts of bacterial Hsp70-Hsp40-NEF. The reactivation rates increased with substrate concentration and followed saturation kinetics, thus allowing the determination of apparent V(max)' and K(m)' values for a chaperone-mediated renaturation reaction for the first time. Under the in vitro conditions used, one Hsp70 molecule consumed five ATPs to effectively unfold a single misfolded protein into an intermediate that, upon chaperone dissociation, spontaneously refolded to the native state, a process with an ATP cost a thousand times lower than expected for protein degradation and resynthesis.

Statistics

Citations

78 citations in Web of Science®
77 citations in Scopus®
Google Scholar™

Altmetrics

Downloads

0 downloads since deposited on 26 Jan 2011
0 downloads since 12 months

Additional indexing

Item Type:Journal Article, refereed, original work
Communities & Collections:04 Faculty of Medicine > Department of Biochemistry
07 Faculty of Science > Department of Biochemistry
Dewey Decimal Classification:570 Life sciences; biology
Language:English
Date:2010
Deposited On:26 Jan 2011 17:21
Last Modified:05 Apr 2016 14:28
Publisher:UNSPECIFIED
ISSN:1552-4450
Additional Information:Comment in: Nat Chem Biol. 2010 Dec;6(12):880-1.
Publisher DOI:https://doi.org/10.1038/nchembio.455
PubMed ID:20953191

Download

Preview Icon on Download
Filetype: PDF (Verlags-PDF) - Registered users only
Size: 1MB
View at publisher