De novo design of miniprotein agonists and antagonists targeting G protein-coupled receptors

Abstract G protein-coupled receptors (GPCRs) play key roles in physiology and are central targets for drug discovery and development, but the design of protein agonists and antagonists has been challenging as GPCRs are integral membrane proteins and conformationally dynamic. Here we describe computational de novo design methods and a high throughput “receptor diversion” microscopy-based screen for generating GPCR binding miniproteins with high affinity, potency and selectivity, and the use of these methods to generate agonists for MRGPRX1, NK1R and CCR5, as well as antagonists for CXCR4, CCR5, OXTR, GLP1R, GIPR, GCGR, PTH1R and CGRPR.. Cryo-electron microscopy data reveals atomic-level agreement between designed and experimentally determined structures for CGRPR- and CXCR4-bound antagonists and MRGPRX1-bound agonists. Our de novo design and screening approach opens new frontiers in GPCR drug discovery and development.