Stable overexpression of human beta 2-adrenergic receptors in mammalian
cells
Naunyn Schmiedebergs Arch Pharmacol, 345 (4):
444-51 (April 1992)Lohse, M J Research Support, Non-U.S. Gov't Germany Naunyn-Schmiedeberg's
archives of pharmacology Naunyn Schmiedebergs Arch Pharmacol. 1992
Apr;345(4):444-51..Abstract
Human beta 2-adrenergic receptors were overexpressed in chinese hamster
ovary (CHO) and human epitheloid carcinoma (HeLa) cells. Stable expression
in these cells was achieved by transfection of a vector containing
the cDNAs for the human beta 2-adrenergic receptor as well as for
dihydrofolate reductase. By stepwise increases of the concentration
of methotrexate - an inhibitor of dihydrofolate reductase - the expression
in CHO cells could be increased to levels of almost 200 pmol/mg membrane
protein, which is more than 1% of the total membrane protein. In
contrast, overexpression of the receptors in HeLa cells by the same
technique led to cell death. The receptors produced in overexpressing
CHO cells were correctly processed and were fully functional with
respect to their ligand binding and signalling properties. The adenylyl
cyclase activity of membranes from these cells responded with extremely
high sensitivity to the beta-adrenergic receptor agonist isoproterenol.
The receptors could be purified from these membranes to apparent
homogeneity by solubilization and chromatography on a single affinity
column. Thus, the expression system described here allows the preparation
of human beta 2-adrenergic receptors in quantities sufficient for
pharmacological and biochemical investigations.