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Updated: Nov 3, 2025

Author Spotlight: Evaluating Biophysical Assays for Characterizing PROTACS Ternary Complexes
Published on: January 12, 2024
An Exploration of Chemical Properties Required for Cooperative Stabilization of the 14-3-3 Interaction with
Madita Wolter1, Dario Valenti1,2, Peter J Cossar1
1Department of Biomedical Engineering, Laboratory of Chemical Biology and Institute for Complex Molecular Systems, Eindhoven University of Technology, P.O. Box 513, 5600 MB Eindhoven, The Netherlands.
None:
Protein-protein modulation has emerged as a proven approach to drug discovery. While significant progress has been gained in developing protein-protein interaction (PPI) inhibitors, the orthogonal approach of PPI stabilization lacks established methodologies for drug design. Here, we report the systematic ″bottom-up″ development of a reversible covalent PPI stabilizer. An imine bond was employed to anchor the stabilizer at the interface of the 14-3-3/p65 complex, leading to a molecular glue that elicited an 81-fold increase in complex stabilization. Utilizing protein crystallography and biophysical assays, we deconvoluted how chemical properties of a stabilizer translate to structural changes in the ternary 14-3-3/p65/molecular glue complex. Furthermore, we explore how this leads to high cooperativity and increased stability of the complex.
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