Thermodynamics of radicicol binding to human Hsp90 alpha and beta isoforms

Asta Zubrienė (1) , Malgorzata Gutkowska (2, 3) , Jurgita Matulienė (1) , Romanas Chaleckis (1) , Vilma Michailovienė (1) , Aliona Voroncova (1) , Česlovas Venclovas (1) , Alicja Zylicz (2) , Maciej Zylicz (2) , Daumantas Matulis (1)

1 Institute of Biotechnology
2 International Institute of Molecular and Cell Biology in Warsaw
3 The Nencki Institute of Experimental Biology

Radicicol is a natural antibiotic that specifically inhibits chaperone Hsp90 activity and binds to its active site with nanomolar affinity. Radicicol has been widely used as a lead compound to generate synthetic analogs with reduced toxicity and increased stability that could be employed clinically. Here we present a detailed thermodynamic description of radicicol binding to human Hsp90 and yeast Hsc82 studied by isothermal titration calorimetry and thermal shift assay. Titrations as a function of pH showed a linked protonation event upon radicicol binding. The intrinsic binding constant and the thermodynamic parameters (including the enthalpy, entropy, and heat capacity) were determined for yeast Hsc82, and human alpha and beta Hsp90. Recent experimental evidence in literature shows that yeast Hsc82 has significant differences from human Hsp90 isozymes. Here we support this by demonstrating differences in radicicol binding thermodynamics to these proteins. The intrinsic enthalpy of radicicol binding to Hsc82 was -46.7kJ/mol, to Hsp90alpha -70.7kJ/mol, and to Hsp90beta was -66.8kJ/mol. The enthalpies of binding were significantly different, while the intrinsic dissociation constants were quite similar, equal to 0.25, 0.04, and 0.15 nM, respectively. The structural features responsible for such large difference in binding enthalpy but small difference in the intrinsic binding Gibbs free energy are discussed.

Mots clés: Isothermal titration calorimetry; Thermal shift assay; ThermoFluor; Heat shock protein 90; Enthalpy

Domaines: Chimie-Physique [physics.chem-ph]

Asta Zubrienė, Malgorzata Gutkowska, Jurgita Matulienė, Romanas Chaleckis, Vilma Michailovienė, et al.. Thermodynamics of radicicol binding to human Hsp90 alpha and beta isoforms. Biophysical Chemistry, 2010, 152 (1-3), pp.153. ⟨10.1016/j.bpc.2010.09.003⟩. ⟨hal-00694295⟩