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Hsp90/Cdc37 chaperone/co-chaperone complex, a novel junction anticancer target elucidated by the mode of action of herbal drug Withaferin A.

Grover A, Shandilya A, Agrawal V, Pratik P, Bhasme D, Bisaria VS, Sundar D - BMC Bioinformatics (2011)

Bottom Line: Arresting the chaperone cycle at these stages by targeting different co-chaperone/Hsp90 interactions seems to be quite a viable alternative and is likely to achieve similar consequences as that of Hsp90 direct inhibition with added favors of high specificity and reduced side effect profile.Formation of active Hsp90/Cdc37 complex is one of the essential steps for facilitation of chaperone client interaction, non-assembly of which can lead to prevention of the chaperone-client association resulting in apoptosis of tumor cells.Conclusively our results strongly suggest that withaferin A is a potent anticancer agent as ascertained by its potent Hsp90-client modulating capability.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biochemical Engineering and Biotechnology, Indian Institute of Technology Delhi, Hauz Khas, New Delhi 110016, India. abhinavgr@gmail.com

ABSTRACT

Background: HSPs (Heat shock proteins) are highly conserved ubiquitous proteins among species which are involved in maintaining appropriate folding and conformation of other proteins and are thus referred to as molecular chaperones. Hsp90 (Heat-shock protein 90 kDa) is one of a group of molecular chaperones responsible for managing protein folding and quality control in cell environment. However it is also involved in the maturation and stabilization of a wide range of oncogenic client proteins which are crucial for oncogenesis and malignant progression. Hsp90 requires a series of co-chaperones to assemble into a super-chaperone complex for its function. These co-chaperones bind and leave the complex at various stages to regulate the chaperoning process. Arresting the chaperone cycle at these stages by targeting different co-chaperone/Hsp90 interactions seems to be quite a viable alternative and is likely to achieve similar consequences as that of Hsp90 direct inhibition with added favors of high specificity and reduced side effect profile. The study conducted here is an attempt to explore the potential of Withania somnifera's major constituent WA (Withaferin A) in attenuating the Hsp90/Cdc37 chaperone/co-chaperone interactions for enhanced tumor arresting activity and to elucidate the underlying mode of action using computational approaches.

Results: Formation of active Hsp90/Cdc37 complex is one of the essential steps for facilitation of chaperone client interaction, non-assembly of which can lead to prevention of the chaperone-client association resulting in apoptosis of tumor cells. From our flexible docking analysis of WA into active Hsp90/Cdc37 complex in which key interfacing residues of the complex were kept flexible, disruption of the active association complex can be discerned. While docking of WA into segregated Hsp90 leaves the interface residues untouched. Thus the molecular docking analysis of WA into Hsp90 and active Hsp90/Cdc37 complex conducted in this study provides significant evidence in support of the proposed mechanism of chaperone assembly suppression by inhibition or disruption of active Hsp90/Cdc37 complex formation being accounted by non-assembly of the catalytically active Hsp90/Cdc37 complex. Results from the molecular dynamics simulations in water show that the trajectories of the protein complexed with ligand WA are stable over a considerably long time period of 4 ns, with the energies of the complex being lowered in comparison to the un-docked association complex, suggesting the thermodynamic stability of WA complexed Hsp90/Cdc37.

Conclusions: The molecular chaperone Hsp90 has been a promising target for cancer therapy. Cancer is a disease marked by genetic instability. Thus specific inhibition of individual proteins or signalling pathways holds a great potential for subversion of this genetic plasticity of cancers. This study is a step forward in this direction. Our computational analysis provided a rationalization to the ability of naturally occurring WA to alter the chaperone signalling pathway. The large value of binding energy involved in binding of WA to the active Hsp90/Cdc37 complex consolidates the thermodynamic stability of the binding. Our docking results obtained substantiate the hypothesis that WA has the potential to inhibit the association of chaperone (Hsp90) to its co-chaperone (Cdc37) by disrupting the stability of attachment of Hsp90 to Cdc37. Conclusively our results strongly suggest that withaferin A is a potent anticancer agent as ascertained by its potent Hsp90-client modulating capability.

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Interactions of docked withaferin A with Hsp90 receptor. (A) H-Bond interactions of the docked ligand with Hsp90 residues. (B) Docked withaferin A forming vdw interactions with the hydrophobic residues of Hsp90.
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Figure 3: Interactions of docked withaferin A with Hsp90 receptor. (A) H-Bond interactions of the docked ligand with Hsp90 residues. (B) Docked withaferin A forming vdw interactions with the hydrophobic residues of Hsp90.

Mentions: One possible mode of action which is proposed here for WA to act as a chaperone function inhibitor is by restriction/disintegration of the complex between Hsp90 and Cdc37. In order to explore the possibility of non-formation of the complex, we first carried out molecular docking studies with only the Hsp90 protein crystal structure. Before docking to be carried out, the structures of receptor macromolecules were minimized using Sander module of Amber. Macromolecular receptors minimized using Steepest Descent and Conjugate Gradient methods had comparatively lower potential energy values than those of the initial ones and are thus utilized further for carrying out the docking studies. Figure 2A shows the docked ligand WA into the selective HSP90 receptor. WA gets buried inside the pocket of HSP90 as depicted by mesh representation in Figure 2B. For this particular configuration the binding energy of WA with Hsp90 is -9.10 Kcal/mol with an Inhibition constant of 214.73 nM (Table 1). The binding of WA to Hsp90 is characterized by H-bonding between a terminal hydroxyl group of the ligand and the side chain carboxyl group of Asp 102. The other terminal hydroxyl group of WA is also involved in a weak H-bond with Asp 54 of Hsp90 (Figure 3A). The lengths of the two H-bonds formed are 3.78 and 5.71 Å. In the present docked structure, WA is also forming van der waals interactions with Leu 48, Asn 51, Asp 54, Ala 55, Leu 107, Ala 111, Val 136 and Phe 138 of Hsp90 (Figure 3B). These non-covalent interactions help stabilize the binding of the ligand with the macromolecule by lowering the energy. However, no interaction of WA with the proposed Cdc37 interacting residues could be observed, as is evident from Figure 4. These finding suggests that binding of WA to Hsp90 could only provide a naive hinderance to the attachment of Cdc37 to Hsp90, owing to the presence of yet available Cdc37 interacting residues of Hsp 90 upon WA binding. However, WA binding to Hsp90 may result in the formation of a deformed complex as the Hsp90/Cdc37 complex would not get assembled to its catalytically active form.


Hsp90/Cdc37 chaperone/co-chaperone complex, a novel junction anticancer target elucidated by the mode of action of herbal drug Withaferin A.

Grover A, Shandilya A, Agrawal V, Pratik P, Bhasme D, Bisaria VS, Sundar D - BMC Bioinformatics (2011)

Interactions of docked withaferin A with Hsp90 receptor. (A) H-Bond interactions of the docked ligand with Hsp90 residues. (B) Docked withaferin A forming vdw interactions with the hydrophobic residues of Hsp90.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3044286&req=5

Figure 3: Interactions of docked withaferin A with Hsp90 receptor. (A) H-Bond interactions of the docked ligand with Hsp90 residues. (B) Docked withaferin A forming vdw interactions with the hydrophobic residues of Hsp90.
Mentions: One possible mode of action which is proposed here for WA to act as a chaperone function inhibitor is by restriction/disintegration of the complex between Hsp90 and Cdc37. In order to explore the possibility of non-formation of the complex, we first carried out molecular docking studies with only the Hsp90 protein crystal structure. Before docking to be carried out, the structures of receptor macromolecules were minimized using Sander module of Amber. Macromolecular receptors minimized using Steepest Descent and Conjugate Gradient methods had comparatively lower potential energy values than those of the initial ones and are thus utilized further for carrying out the docking studies. Figure 2A shows the docked ligand WA into the selective HSP90 receptor. WA gets buried inside the pocket of HSP90 as depicted by mesh representation in Figure 2B. For this particular configuration the binding energy of WA with Hsp90 is -9.10 Kcal/mol with an Inhibition constant of 214.73 nM (Table 1). The binding of WA to Hsp90 is characterized by H-bonding between a terminal hydroxyl group of the ligand and the side chain carboxyl group of Asp 102. The other terminal hydroxyl group of WA is also involved in a weak H-bond with Asp 54 of Hsp90 (Figure 3A). The lengths of the two H-bonds formed are 3.78 and 5.71 Å. In the present docked structure, WA is also forming van der waals interactions with Leu 48, Asn 51, Asp 54, Ala 55, Leu 107, Ala 111, Val 136 and Phe 138 of Hsp90 (Figure 3B). These non-covalent interactions help stabilize the binding of the ligand with the macromolecule by lowering the energy. However, no interaction of WA with the proposed Cdc37 interacting residues could be observed, as is evident from Figure 4. These finding suggests that binding of WA to Hsp90 could only provide a naive hinderance to the attachment of Cdc37 to Hsp90, owing to the presence of yet available Cdc37 interacting residues of Hsp 90 upon WA binding. However, WA binding to Hsp90 may result in the formation of a deformed complex as the Hsp90/Cdc37 complex would not get assembled to its catalytically active form.

Bottom Line: Arresting the chaperone cycle at these stages by targeting different co-chaperone/Hsp90 interactions seems to be quite a viable alternative and is likely to achieve similar consequences as that of Hsp90 direct inhibition with added favors of high specificity and reduced side effect profile.Formation of active Hsp90/Cdc37 complex is one of the essential steps for facilitation of chaperone client interaction, non-assembly of which can lead to prevention of the chaperone-client association resulting in apoptosis of tumor cells.Conclusively our results strongly suggest that withaferin A is a potent anticancer agent as ascertained by its potent Hsp90-client modulating capability.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biochemical Engineering and Biotechnology, Indian Institute of Technology Delhi, Hauz Khas, New Delhi 110016, India. abhinavgr@gmail.com

ABSTRACT

Background: HSPs (Heat shock proteins) are highly conserved ubiquitous proteins among species which are involved in maintaining appropriate folding and conformation of other proteins and are thus referred to as molecular chaperones. Hsp90 (Heat-shock protein 90 kDa) is one of a group of molecular chaperones responsible for managing protein folding and quality control in cell environment. However it is also involved in the maturation and stabilization of a wide range of oncogenic client proteins which are crucial for oncogenesis and malignant progression. Hsp90 requires a series of co-chaperones to assemble into a super-chaperone complex for its function. These co-chaperones bind and leave the complex at various stages to regulate the chaperoning process. Arresting the chaperone cycle at these stages by targeting different co-chaperone/Hsp90 interactions seems to be quite a viable alternative and is likely to achieve similar consequences as that of Hsp90 direct inhibition with added favors of high specificity and reduced side effect profile. The study conducted here is an attempt to explore the potential of Withania somnifera's major constituent WA (Withaferin A) in attenuating the Hsp90/Cdc37 chaperone/co-chaperone interactions for enhanced tumor arresting activity and to elucidate the underlying mode of action using computational approaches.

Results: Formation of active Hsp90/Cdc37 complex is one of the essential steps for facilitation of chaperone client interaction, non-assembly of which can lead to prevention of the chaperone-client association resulting in apoptosis of tumor cells. From our flexible docking analysis of WA into active Hsp90/Cdc37 complex in which key interfacing residues of the complex were kept flexible, disruption of the active association complex can be discerned. While docking of WA into segregated Hsp90 leaves the interface residues untouched. Thus the molecular docking analysis of WA into Hsp90 and active Hsp90/Cdc37 complex conducted in this study provides significant evidence in support of the proposed mechanism of chaperone assembly suppression by inhibition or disruption of active Hsp90/Cdc37 complex formation being accounted by non-assembly of the catalytically active Hsp90/Cdc37 complex. Results from the molecular dynamics simulations in water show that the trajectories of the protein complexed with ligand WA are stable over a considerably long time period of 4 ns, with the energies of the complex being lowered in comparison to the un-docked association complex, suggesting the thermodynamic stability of WA complexed Hsp90/Cdc37.

Conclusions: The molecular chaperone Hsp90 has been a promising target for cancer therapy. Cancer is a disease marked by genetic instability. Thus specific inhibition of individual proteins or signalling pathways holds a great potential for subversion of this genetic plasticity of cancers. This study is a step forward in this direction. Our computational analysis provided a rationalization to the ability of naturally occurring WA to alter the chaperone signalling pathway. The large value of binding energy involved in binding of WA to the active Hsp90/Cdc37 complex consolidates the thermodynamic stability of the binding. Our docking results obtained substantiate the hypothesis that WA has the potential to inhibit the association of chaperone (Hsp90) to its co-chaperone (Cdc37) by disrupting the stability of attachment of Hsp90 to Cdc37. Conclusively our results strongly suggest that withaferin A is a potent anticancer agent as ascertained by its potent Hsp90-client modulating capability.

Show MeSH
Related in: MedlinePlus