Enforced N-domain Proximity Stimulates Hsp90 ATPase Activity and Is Compatible with Function in Vivo

Pullen, Lester; Bolon, Daniel N.

doi:10.1074/jbc.m111.223131

Cited by 33 publications

(33 citation statements)

References 35 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The CL connects the N and M domains, which both contribute catalytic amino acids for ATP hydrolysis. The CL has been implicated in the ATP-and cochaperone-driven conformational cycle of Hsp90 (10,22,34,41). Our results indicate an additional critical function of CL and CX: imparting high solubility on Hsp90 in complex with aggregation-prone clients.…”

Section: Discussionmentioning

confidence: 69%

“…Deletions that extend into regions ordered within the Hsp90 crystal structure resulted in reduced growth at temperatures of above 37°C (10). While flexibility is clearly inherent to Hsp90 (17,41), it can be dramatically reduced (10,34,46) without impairing yeast growth. In addition, flexibility alone is insufficient to explain the evolutionary conservation of negative charge in the CL.…”

mentioning

confidence: 99%

See 1 more Smart Citation

Solubility-Promoting Function of Hsp90 Contributes to Client Maturation and Robust Cell Growth

2012

View full text Add to dashboard Cite

The Hsp90 chaperone is required for the maturation of signal transduction clients, including many kinases and nuclear steroid hormone receptors. The binding and hydrolysis of ATP by Hsp90 drive conformational rearrangements in three structure domains. Two intrinsically disordered regions of Hsp90 located between these domains and at the C terminus have traditionally been considered to impart flexibility. We discovered that the charged nature of these acid-rich disordered regions imparts a solubility-promoting function to Hsp90 that is important for its cellular activity in yeast. Both the solubility-promoting function and ATPase activity must occur in the same Hsp90 molecule in order to support robust growth, suggesting that the solubility-promoting function is required during the ATP-driven client maturation process. Expression of model clients together with Hsp90 variants indicated interdependent solubilities mediated by the aggregation propensities of both the client and Hsp90. We propose a model whereby the charge-rich disordered regions of Hsp90 serve a solubility-promoting function important for complexes with aggregation-prone clients. These findings demonstrate a novel biological function of the intrinsically disordered regions in Hsp90 and provide a compelling rationale for why their charged properties are conserved throughout eukaryotic evolution.

show abstract

Section: Discussionmentioning

confidence: 69%

mentioning

confidence: 99%

Solubility-Promoting Function of Hsp90 Contributes to Client Maturation and Robust Cell Growth

2012

View full text Add to dashboard Cite

show abstract

“…In spite of recent works, the involvement of Hsp90's domains in oligomerization process remains unclear as MC-HSP90 exists in three major states (dimer, tetramer and hexamer), and as NTDs could also participate in this process as they are able to self associate too [48]. Moreover, the NTDs are involved in Hsp90 dimer transient dimerization [49][50][51]: such interactions between NTDs in the dimer could also exist within oligomers. Even if the role of oligomers in the Hsp90 chaperone cycle is not yet understood, oligomers are known to exhibit a chaperone activity preventing protein aggregation and maintaining them in a folding-competent state [52].…”

Section: Introductionmentioning

confidence: 89%

Hsp90 oligomerization process: How can p23 drive the chaperone machineries?

Lepvrier

Nigen

Moullintraffort

et al. 2015

Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics

View full text Add to dashboard Cite

“…This construct has similar ATPase, p23 binding, and kinetics as characterized before (10). In addition, such fused Hsp90 are viable in yeast (27). Cysteine point mutations were created with the QuickChange Multi Site-Directed Mutagenesis Kit (Stratagene).…”

Section: Methodsmentioning

confidence: 99%

“…Because the hydrolysis rate is very slow compared to all other rates, Hsp90 in the absence of cochaperones and substrate proteins operates close to thermal equilibrium; occasional ATP hydrolysis only slightly shifts it out of equilibrium. The hydrolysis step itself happens in the closed state because N-terminal dimerization is necessary for ATP hydrolysis (19,27).…”

Section: The N Terminus Is Likely More Extended Than Given By the Crymentioning

confidence: 99%

Heat shock protein 90’s mechanochemical cycle is dominated by thermal fluctuations

Ratzke

Berkemeier

Hugel

2011

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

The molecular chaperone and heat shock protein 90 (Hsp90) exists mainly as a homodimer in the cytoplasm. Each monomer has an ATPase in its N-terminal domain and undergoes large conformational changes during Hsp90's mechanochemical cycle. The threecolor single-molecule assay and data analysis presented in the following allows one to observe at the same time nucleotide binding and the conformational changes in Hsp90. Surprisingly, and completely unlike the prior investigated systems, nucleotides can bind to the N-terminally open and closed state without strictly forcing the protein into a specific conformation. Both the transitions between the conformational states and the nucleotide binding/ unbinding are mainly thermally driven. Furthermore, the two ATP binding sites show negative cooperativity; i.e., nucleotides do not bind independently to the two monomers. We thus reveal a picture of how nucleotide binding and conformational changes are connected in the molecular chaperone Hsp90, which has far-ranging consequences for its function and is distinct from previously investigated motor proteins.conformational dynamics | three-color FRET | single molecule | intramolecular communication | protein structure M olecular chaperones are proteins that assist the folding or unfolding and the assembly or disassembly of other proteins (1). A very abundant molecular chaperone in eukaryotes is the heat shock protein 90 (Hsp90) comprising around 1% of cytosolic protein (2, 3). The protein consists of two elongated monomers that have two dimerization interfaces in the N domain and in the C domain, respectively. Like motor proteins, it is an ATPase, but the mechanistic function of ATP binding and hydrolysis is still unclear. Current models assume that one ATP each binds to the N-terminal domains in the open state, followed by N-terminal closing and ATP hydrolysis in a strict succession (4)-similar to all to-date investigated motor proteins (5-9). Because of experimental limitations, it was not possible to directly observe nucleotide binding and conformational changes at the same time and thus detect the mechanochemical cycle of Hsp90. To overcome this limitation we designed a three color single-molecule Förster resonance energy transfer (smFRET) assay with alternating laser excitation (ALEX). This allows one to detect when ATP binds to which monomer and at the same time to read out the conformational state of Hsp90. Surprisingly, we find that ATP binds to the N-terminal open and close conformations, but not independently. Therefore, our data show that the picture of a successive conformational cycle has to be replaced by stochastic transitions in-between conformations of Hsp90. Nevertheless, there is an anticorrelated communication between the N termini throughout the whole protein.

show abstract

Enforced N-domain Proximity Stimulates Hsp90 ATPase Activity and Is Compatible with Function in Vivo

Cited by 33 publications

References 35 publications

Solubility-Promoting Function of Hsp90 Contributes to Client Maturation and Robust Cell Growth

Solubility-Promoting Function of Hsp90 Contributes to Client Maturation and Robust Cell Growth

Hsp90 oligomerization process: How can p23 drive the chaperone machineries?

Heat shock protein 90’s mechanochemical cycle is dominated by thermal fluctuations

Contact Info

Product

Resources

About