Premium
Heterologous expression of functional G‐protein‐coupled receptors in Caenorhabditis elegans
Author(s) -
Salom David,
Cao Pengxiu,
Sun Wenyu,
Kramp Kristopher,
Jastrzebska Beata,
Jin Hui,
Feng Zhaoyang,
Palczewski Krzysztof
Publication year - 2012
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fj.11-197780
Subject(s) - g protein coupled receptor , rhodopsin , caenorhabditis elegans , heterologous expression , opsin , heterologous , biology , transducin , receptor , g protein , microbiology and biotechnology , biochemistry , membrane protein , recombinant dna , chemistry , membrane , retinal , gene
New strategies for expression, purification, functional characterization, and structural determination of membrane‐spanning G‐protein‐coupled receptors (GPCRs) are constantly being developed because of their importance to human health. Here, we report a Caenorhabditis elegans heterologous expression system able to produce milligram amounts of functional native and engineered GPCRs. Both bovine opsin [(b)opsin] and human adenosine A 2A subtype receptor [(h)A 2A R] expressed in neurons or muscles of C. elegans were localized to cell membranes. Worms expressing these GPCRs manifested changes in motor behavior in response to light and ligands, respectively. With a newly devised protocol, 0.6‐1 mg of purified homogenous 9‐ cis ‐retinal‐bound bovine isorhodopsin [(b)isoRho] and ligand‐bound (h)A 2A R were obtained from C. elegans from one 10‐L fermentation at low cost. Purified recombinant (b)isoRho exhibited its signature absor‐bance spectrum and activated its cognate G‐protein transducin in vitro at a rate similar to native rhodopsin (Rho) obtained from bovine retina. Generally high expression levels of 11 native and mutant GPCRs demonstrated the potential of this C. elegans system to produce milligram quantities of high‐quality GPCRs and possibly other membrane proteins suitable for detailed characterization.—Salom, D., Cao, P., Sun, W., Kramp, K., Jastrzebska, B., Jin, H., Feng, Z., Palczewski, K. Heterologous expression of functional G‐protein‐coupled receptors in Caenorhabditis elegans . FASEB J. 26, 492–502 (2012). www.fasebj.org