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|Title:||GPCR expression using baculovirus-infected Sf9 cells|
|Citation:||G Protein-Coupled Receptors in Drug Discovery, 2009 / Leifert, W.R. (ed./s), Ch.8, pp.115-129|
|Series/Report no.:||Methods in Molecular Biology; 552|
|Amanda L. Aloia, Richard V. Glatz, Edward J. McMurchie, and Wayne R. Leifert|
|Abstract:||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 6 x histidine-tagged mammalian M(2) 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.|
|Keywords:||Baculovirus; Insect cell; GPCR expression; M2 muscarinic receptor|
|Rights:||© Humana Press, a part of Springer Science+Business Media, LLC 2009|
|Appears in Collections:||Molecular and Biomedical Science publications|
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