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Proc Natl Acad Sci U S A. 2007 Mar 20;104(12):5163-8. Epub 2007 Mar 2.

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

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1
Department of Biochemistry, Case Western Reserve University School of Medicine, Cleveland, OH 44106, USA.

Abstract

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 G(i)-coupled designer receptor in hippocampal neurons (hM(4)D) 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.

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PMID:
17360345
PMCID:
PMC1829280
DOI:
10.1073/pnas.0700293104
[Indexed for MEDLINE]
Free PMC Article
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