摘要：SummaryHsp90 is a target for anti-cancer drug development. Both the conformational events tuned by ATP/ADP and co-chaperones and the chaperoning cycle timing are required for Hsp90's fully functional display. Interfering with either one of the conformational events or the cycle timing will down-regulate Hsp90's function. In this manuscript, non-covalent allosteric modulators(SOMCL-16-171andSOMCL-16-175) targeting Hsp90α’s middle domain (Hsp90M) were developed for the first time. Multiple techniques were then applied to characterize the interactions between two active compounds and Hsp90α. Two loops and one α-helix (F349-N360, K443-E451, and D372-G387) in Hsp90M were identified responsible for the recognition ofSOMCL-16-171andSOMCL-16-175. Meanwhile, the binding ofSOMCL-16-171andSOMCL-16-175to Hsp90M was demonstrated to allosterically modulate the structure and function of Hsp90α’s N-terminal domain. Finally, cellular assays were conducted to evaluate the cellular activity ofSOMCL-16-175, and the results indicate thatSOMCL-16-175destabilizes Hsp90's client proteins and reduces cell viability.Graphical AbstractDisplay OmittedHighlights•Allosteric modulators targeting Hsp90α's middle domain were developed for the first time•Key elements in Hsp90M for the recognition of allosteric modulators were identified•Compound SOMCL-16-175 promotes Hsp90α’s ATPase activity and reduces cell viability•SOMCL-16-175 destabilizes Hsp90's clients without triggering heat shock responseMolecular Biology; Molecular Structure; Cancer
