Evolving the lock to fit the key to create a family of G protein-coupled receptors potently activated by an inert ligand

BN Armbruster, X Li, MH Pausch… - Proceedings of the …, 2007 - National Acad Sciences
BN Armbruster, X Li, MH Pausch, S Herlitze, BL Roth
Proceedings of the National Academy of Sciences, 2007National Acad Sciences
We evolved muscarinic receptors in yeast to generate a family of G protein-coupled
receptors (GPCRs) that are activated solely by a pharmacologically inert drug-like and
bioavailable compound (clozapine-N-oxide). Subsequent screening in human cell lines
facilitated the creation of a family of muscarinic acetylcholine GPCRs suitable for in vitro and
in situ studies. We subsequently created lines of telomerase-immortalized human pulmonary
artery smooth muscle cells stably expressing all five family members and found that each …
We evolved muscarinic receptors in yeast to generate a family of G protein-coupled receptors (GPCRs) that are activated solely by a pharmacologically inert drug-like and bioavailable compound (clozapine-N-oxide). Subsequent screening in human cell lines facilitated the creation of a family of muscarinic acetylcholine GPCRs suitable for in vitro and in situ studies. We subsequently created lines of telomerase-immortalized human pulmonary artery smooth muscle cells stably expressing all five family members and found that each one faithfully recapitulated the signaling phenotype of the parent receptor. We also expressed a Gi-coupled designer receptor in hippocampal neurons (hM4D) and demonstrated its ability to induce membrane hyperpolarization and neuronal silencing. We have thus devised a facile approach for designing families of GPCRs with engineered ligand specificities. Such reverse-engineered GPCRs will prove to be powerful tools for selectively modulating signal-transduction pathways in vitro and in vivo.
National Acad Sciences