In Situ Click Chemistry Screen Facilitated by an On‐Nanoparticle DNA‐Encoded Library Identifies Highly Selective and Potent Peptoid Ligands for a Phosphatase

Capture agents that selectively bind to biological targets are indispensable tools in diagnostics, therapeutics, and biomedical research. However, discovering such capture agents, particularly for structurally conserved or challenging targets, remains a challenge. Here, we describe a protein-templated in situ click strategy enabled by a nanoparticle-based DNA-encoded library (nanoDEL) platform. The nanoDEL enables the construction and screening of vastly large, chemically diverse combinatorial libraries with high redundancy, far exceeding the scale and throughput of conventional approaches, such as one-bead-one-compound and solution-phase in situ click methods. Thus, this enables the rapid and efficient identification of high-affinity, high-selectivity ligands in a single selection round, eliminating the need for iterative screening. To demonstrate the utility of this strategy, we performed an in situ click screening of a 27-million-member nanoDEL of azido-functionalized peptoids, in the presence of a weak and promiscuous alkyne-bearing anchor ligand. This yielded bidentate inhibitors of protein tyrosine phosphatase 1B, a challenging target due to its highly conserved active site. These inhibitors exhibited nanomolar potency and exceptional selectivity over closely related phosphatases. This work represents a broadly applicable strategy for discovering high-performance capture agents, particularly for selectively targeting closely related protein families or isoforms where achieving selectivity remains a critical challenge.