A Generic Method for the Production of Cell Lines Expressing High Levels of 7-Transmembrane Receptors

Authors: Koller K.J.; Whitehorn E.A.; Tate E.; Ries T.; Aguilar B.; Chernov-Rogan T.; Davis A.M.; Dobbs A.; Yen M.; Barrett R.W.

Source: Analytical Biochemistry, Volume 250, Number 1, July 1997 , pp. 51-60(10)

Publisher: Academic Press

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Abstract:

G-protein-coupled or 7-transmembrane receptors (7TMRs) are often studied after heterologous expression in mammalian cells such as COS-7, CHO-K1, or HEK-293s. In this paper, we describe the development of a rapid and generic method for producing stable Chinese hamster ovary cell lines expressing high levels of recombinant 7TMRs by N-terminal tagging these proteins with the hemagglutinin (HA) sequence. To illustrate the broad applicability of this technique, we have presented data from cell lines expressing a glycoprotein hormone receptor for follicle-stimulating hormone (FSHR), CXC- (CXCR-2), and CC-chemokine (CCR-1) receptors and peptide receptors from the somatostatin (SSTR1, 2, 5) and neuropeptide Y (NPY-Y2, -Y4 Rs) families. Typically, cell lines with a receptor density of 1 to 15 pmol/mg protein are produced with this method. The presence of the HA tag does not adversely affect the binding or functional activity of the receptors.

Language: English

Document Type: Research article

Affiliations: Affymax Research Institute, 4001 Miranda Avenue, Palo Alto, California, 94304:

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