RESUMEN
Destabilizing mutations in small heat shock proteins (sHsps) are linked to multiple diseases; however, sHsps are conformationally dynamic, lack enzymatic function and have no endogenous chemical ligands. These factors render sHsps as classically "undruggable" targets and make it particularly challenging to identify molecules that might bind and stabilize them. To explore potential solutions, we designed a multi-pronged screening workflow involving a combination of computational and biophysical ligand-discovery platforms. Using the core domain of the sHsp family member Hsp27/HSPB1 (Hsp27c) as a target, we applied mixed solvent molecular dynamics (MixMD) to predict three possible binding sites, which we confirmed using NMR-based solvent mapping. Using this knowledge, we then used NMR spectroscopy to carry out a fragment-based drug discovery (FBDD) screen, ultimately identifying two fragments that bind to one of these sites. A medicinal chemistry effort improved the affinity of one fragment by ~50-fold (16 µM), while maintaining good ligand efficiency (~0.32 kcal/mol/non-hydrogen atom). Finally, we found that binding to this site partially restored the stability of disease-associated Hsp27 variants, in a redox-dependent manner. Together, these experiments suggest a new and unexpected binding site on Hsp27, which might be exploited to build chemical probes.