Resolving Hot Spots in the C-Terminal Dimerization Domain that Determine the Stability of the Molecular Chaperone Hsp90 |
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| Authors: | Emanuele Ciglia Janina Vergin Sven Reimann Sander H. J. Smits Lutz Schmitt Georg Groth Holger Gohlke |
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| Affiliation: | 1. Institute for Pharmaceutical and Medicinal Chemistry, Heinrich-Heine-University, Düsseldorf, Germany.; 2. Institute for Biochemical Plant Physiology, Heinrich-Heine-University, Düsseldorf, Germany.; 3. Institute of Biochemistry, Heinrich-Heine-University, Düsseldorf, Germany.; Fred Hutchinson Cancer Research Center, United States of America, |
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| Abstract: | Human heat shock protein of 90 kDa (hHsp90) is a homodimer that has an essential role in facilitating malignant transformation at the molecular level. Inhibiting hHsp90 function is a validated approach for treating different types of tumors. Inhibiting the dimerization of hHsp90 via its C-terminal domain (CTD) should provide a novel way to therapeutically interfere with hHsp90 function. Here, we predicted hot spot residues that cluster in the CTD dimerization interface by a structural decomposition of the effective energy of binding computed by the MM-GBSA approach and confirmed these predictions using in silico alanine scanning with DrugScorePPI. Mutation of these residues to alanine caused a significant decrease in the melting temperature according to differential scanning fluorimetry experiments, indicating a reduced stability of the mutant hHsp90 complexes. Size exclusion chromatography and multi-angle light scattering studies demonstrate that the reduced stability of the mutant hHsp90 correlates with a lower complex stoichiometry due to the disruption of the dimerization interface. These results suggest that the identified hot spot residues can be used as a pharmacophoric template for identifying and designing small-molecule inhibitors of hHsp90 dimerization. |
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