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Expression of proteins in insect cells using recombinant baculoviruses has gained wide use in the G protein-coupled receptor
(GPCR) community. This expression system produces high yields of functional receptor, is able to perform post-translational
modifications, and is readily adaptable to large-scale culture. Here, we describe the generic methods for expressing a GPCR
using baculovirus-infected insect cells, including the maintenance of insect cell culture. Data are presented for polyhedrin promoter-driven expression of a C-terminal 6Ă—histidine-tagged mammalian M2 muscarinic receptor in Sf9 cells. Results demonstrate that expressed receptor could be detected and quantified using radiolabeled ligand binding, that
expression was maximal at approximately 72 h post-infection, and that expression levels could be altered by addition of various
ligands to cultures of infected insect cells.
Affiliation(s): (2) School of Biology, Flinders University, Bedford Park, SA, Australia
(3) SARDI Entomology, Adelaide, SA, Australia
(4) CSIRO Molecular and Health Technologies, Adelaide, SA, Australia
(5) CSIRO Human Nutrition, Adelaide, SA, Australia
(3) SARDI Entomology, Adelaide, SA, Australia
(4) CSIRO Molecular and Health Technologies, Adelaide, SA, Australia
(5) CSIRO Human Nutrition, Adelaide, SA, Australia
Book Title: G Protein-Coupled Receptors in Drug Discovery
Series: Methods in Molecular Biology | Volume: 552 | Pub. Date: Aug-01-2009 | Page Range: 115-129 | DOI: 10.1007/978-1-60327-317-6_8
Subject: Pharmacology/Toxicology
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