RXFP3 is a class A rhodospsin-like G-protein coupled receptor (GPCR) expressed within the regions of the rodent and primate brain. It is the cognate receptor for the neuropeptide relaxin-3 which together, controls neuronal modalities such as sleep, attention, feeding, cognition and the response to stress hence represents an attractive novel target for the therapeutic treatment of a range of mental illnesses.

Modern GPCR drug development is encumbered by a lack of information about the protein structure and dynamics underlying GPCR function. Methods that improve the stability of GPCRs are needed to facilitate the application of these important proteins to biochemical and biophysical techniques such as crystallography and nuclear magnetic resonance spectroscopy (NMR), aiding in the identification and design of new drugs.  One such method is CHESS, or Cellular High throughput Encapsulation, Solubilisation and Screening. CHESS is directed evolution approach which works by encapsulating mutant libraries of cells in detergent-stable capsules, keeping the cells intact when solubilized in detergent and probing for stable mutants using fluorescence ligands. The mutants in the capsules that are detergent-stable remain active and able to bind fluorescence ligands and so can be sorted using Fluorescence Activated Cell Sorting (FACS).

In this project, we first established the bacterial expression of RXFP3 protein which provides functional protein in a scalable way. This will allow the application of RXFP3 to directed evolution methods to identify high expressing and stabilised receptor variants from large mutant libraries including CHESS. Encapsulation of the RXFP3-expressing cells and binding of ligands to RXFP3 within encapsulated cells were analysed. With these stabilised receptors, greater biochemical and biophysical studies can be carried out to provide structural and functional insights to RXFP3 and other GPCRs.