A Generic Method for the Production of Cell Lines Expressing High Levels of 7-Transmembrane Receptors
| Autor: | Emily Tate, Tania Chernov-Rogan, Ronald W. Barrett, Erik A. Whitehorn, Kerry J. Koller, Ann M. Davis, Mary Yen, Tracy Ries, Andrea Dobbs, Barbara Aguilar |
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| Rok vydání: | 1997 |
| Předmět: |
Genetic Vectors
Molecular Sequence Data Biophysics Gene Expression Receptors Cell Surface CHO Cells Protein Sorting Signals Biology Transfection Biochemistry GTP-Binding Proteins Cell surface receptor Cricetinae Animals Somatostatin receptor 1 Amino Acid Sequence HA-tag Receptor Molecular Biology Chinese hamster ovary cell Antibodies Monoclonal Cell Biology Neuropeptide Y receptor Molecular biology Recombinant Proteins Hemagglutinins Hormone receptor Heterologous expression Plasmids |
| Zdroj: | Analytical Biochemistry. 250:51-60 |
| ISSN: | 0003-2697 |
| DOI: | 10.1006/abio.1997.2190 |
| Popis: | 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. |
| Databáze: | OpenAIRE |
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