Fig. 3. Applications of the monobody technology for facilitating drug discovery. (a) Exquisite selectivity of monobodies facilitates drug target validation and discovery of druggable sites. A panel of monobodies (teal) was developed for inhibiting aGPCR and dissecting the functional role of individual extracellular domains (left). A monobody selectively recognizes STAT3 over other STAT family members. Monobody 12VC1 selectively inhibits RAS mutants, G12C and G12V, over other WT RAS isoforms. (b) Monobody-assisted discovery of new targetable interfaces for inhibition (PDB ID: 5E95).⁵⁶ The NS1 monobody inhibited RAS by binding (yellow) to the α4–β6–α5 interface (enclosed with the black boundary) that is important for RAS: RAS association, which is away from the effector binding interface (red). Structure of monobody Mb33 bound to MLKL 4HB domain (PDB ID: 6UX8).⁶⁰ (c) Monobody MB2 (PDB ID: 6C83, teal)⁵⁹ allosterically inhibited AurA by targeting the pocket. The equivalent pocket of a homologous kinase, PDK1, is targeted by small molecules inhibitors (PDB ID: 3ORX, magenta, PDB ID: 4RQK, green)^146,147 (d) A peptide inhibitor (PDB ID: 5WGQ, red)¹⁴⁸ that binds to ER-α closely resembles the FG loop of a monobody (PDB ID: 2OCF, teal).¹⁴⁹ Small molecule (PDB ID: 6E23, green)¹⁵⁰ and peptide inhibitor (PDB ID: 5VFC magenta)¹⁵¹ that targets WDR5 closely resembles the FG loop of a WDR5-binding monobody (PDB ID: 6BYN, teal).⁵⁷.