The effect of celastrol, a triterpene with antitumorigenic activity, on conformational and functional aspects of the human 90kDa heat shock protein Hsp90α, a chaperone implicated in the stabilization of the tumor phenotype

• Published in: Biochimica et biophysica acta Vol. 1840; no. 10; pp. 3145 - 3152
• Main Authors: Zanphorlin, Letícia M., Alves, Fernanda R., Ramos, Carlos H.I.
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.10.2014
• Subjects: Anticancer agents; antineoplastic activity; Antineoplastic Agents - chemistry; Antineoplastic Agents - pharmacology; antioxidants; Celastrol; cell viability; eukaryotic cells; heat shock proteins; Hsp90; HSP90 Heat-Shock Proteins - antagonists & inhibitors; HSP90 Heat-Shock Proteins - chemistry; HSP90 Heat-Shock Proteins - genetics; HSP90 Heat-Shock Proteins - metabolism; Humans; mechanism of action; Mitochondrial Membrane Transport Proteins - antagonists & inhibitors; Mitochondrial Membrane Transport Proteins - chemistry; Mitochondrial Membrane Transport Proteins - genetics; Mitochondrial Membrane Transport Proteins - metabolism; Molecular chaperones; mutants; Neoplasm Proteins - antagonists & inhibitors; Neoplasm Proteins - chemistry; Neoplasm Proteins - genetics; Neoplasm Proteins - metabolism; neoplasms; Neoplasms - drug therapy; Neoplasms - genetics; Neoplasms - metabolism; oligomerization; phenotype; Protein Binding; Protein Multimerization - drug effects; Protein Stability - drug effects; Protein Structure, Tertiary; Protein–ligand interaction; Recombinant Proteins - chemistry; Recombinant Proteins - genetics; Recombinant Proteins - metabolism; TPR co-chaperone; Triterpenes - chemistry; Triterpenes - pharmacology; triterpenoids; CD; TPR co-chaperone; SDS; Celastrol; DLS; PAGE; Anticancer agents; Hsp90α; Hsp90; Protein–ligand interaction; C-Hsp90α; Molecular chaperones; DSC; DSF; SEC–MALS
• ISSN: 0304-4165; 0006-3002; 1872-8006
• DOI: 10.1016/j.bbagen.2014.06.008

Hsp90 is a molecular chaperone essential for cell viability in eukaryotes that is associated with the maturation of proteins involved in important cell functions and implicated in the stabilization of the tumor phenotype of various cancers, making this chaperone a notably interesting therapeutic target. Celastrol is a plant-derived pentacyclic triterpenoid compound with potent antioxidant, anti-inflammatory and anticancer activities; however, celastrol's action mode is still elusive. In this work, we investigated the effect of celastrol on the conformational and functional aspects of Hsp90α. Interestingly, celastrol appeared to target Hsp90α directly as the compound induced the oligomerization of the chaperone via the C-terminal domain as demonstrated by experiments using a deletion mutant. The nature of the oligomers was investigated by biophysical tools demonstrating that a two-fold excess of celastrol induced the formation of a decameric Hsp90α bound throughout the C-terminal domain. When bound, celastrol destabilized the C-terminal domain. Surprisingly, standard chaperone functional investigations demonstrated that neither the in vitro chaperone activity of protecting against aggregation nor the ability to bind a TPR co-chaperone, which binds to the C-terminus of Hsp90α, were affected by celastrol. Celastrol interferes with specific biological functions of Hsp90α. Our results suggest a model in which celastrol binds directly to the C-terminal domain of Hsp90α causing oligomerization. However, the ability to protect against protein aggregation (supported by our results) and to bind to TPR co-chaperones are not affected by celastrol. Therefore celastrol may act primarily by inducing specific oligomerization that affects some, but not all, of the functions of Hsp90α. To the best of our knowledge, this study is the first work to use multiple probes to investigate the effect that celastrol has on the stability and oligomerization of Hsp90α and on the binding of this chaperone to Tom70. This work provides a novel mechanism by which celastrol binds Hsp90α. [Display omitted] •This work provides a novel mechanism by which celastrol binds Hsp90α.•Celastrol induced the oligomerization of Hsp90α via the C-terminal domain.•Hsp90α in vitro chaperone activity was not affected by celastrol.•Hsp90α ability to bind a TPR co-chaperone was not affected by celastrol.