De Novo Modular Development of a Foldameric Protein–Protein Interaction Inhibitor for Separate Hot Spots: A Dynamic Covalent Assembly Approach

Protein–protein interactions stabilized by multiple separate hot spots are highly challenging targets for synthetic scaffolds. Surface‐mimetic foldamers bearing multiple recognition segments are promising candidate inhibitors. In this work, a modular bottom‐up approach is implemented by identifying short foldameric recognition segments that interact with the independent hot spots, and connecting them through dynamic covalent library (DCL) optimization. The independent hot spots of a model target (calmodulin) are mapped with hexameric β‐peptide helices using a pull‐down assay. Recognition segment hits are subjected to a target‐templated DCL ligation through thiol–disulfide exchange. The most potent derivative displays low nanomolar affinity towards calmodulin and effectively inhibits the calmodulin–TRPV1 interaction. The DCL assembly of the folded segments offers an efficient approach towards the de novo development of a high‐affinity inhibitor of protein–protein interactions.