Semliki Forest virus vectors for overexpression of 101 G protein-coupled receptors in mammalian host cells.
Semliki Forest virus vectors were applied for the evaluation of 101 G protein-coupled receptors in three mammalian cell lines. Western blotting demonstrated that 95 of the 101 tested GPCRs showed positive signals. A large number of the GPCRs were expressed at high levels suggesting receptor yields in the range of 1 mg/L or higher, suitable for structural biology applications. Specific binding assays on a selected number of GPCRs were carried out to compare the correlation between total and functional protein expression. Ligands and additives supplemented to the cell culture medium were evaluated for expression enhancement. Selected GPCRs were also expressed from mutant SFV vectors providing enhanced protein expression and reduced host cell toxicity in attempts to further improve receptor yields.
Hassaine
Gherici
G
BioXtal, Chemin des Croisettes 22, CH-1066 Epalinges, Switzerland.
Wagner
Renaud
R
Kempf
Juliette
J
Cherouati
Nadia
N
Hassaine
Nouzha
N
Prual
Cecile
C
André
Nicolas
N
Reinhart
Christoph
C
Pattus
Franc
F
Lundstrom
Kenneth
K
D000818:Animals; D002460:Cell Line; D006224:Cricetinae; D003470:Culture Media / Q000737:chemistry; D020869:Gene Expression Profiling; D005822:Genetic Vectors / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D011869:Radioligand Assay; D043562:Receptors, G-Protein-Coupled / Q000235:genetics / Q000378:metabolism*; D012672:Semliki forest virus / Q000235:genetics / Q000378:metabolism
D003470:Culture Media; D043562:Receptors, G-Protein-Coupled