Functional expression of rat alpha 2B-adrenoceptor in Escherichia coli.

Rat alpha 2B-adrenoceptor was expressed in Escherichia coli using 'ATG vector' containing cDNA encoding the 'non-glycosylated rat alpha 2-adrenoceptor' (RNG alpha 2). The highest receptor binding activity (using the alpha 2-adrenoceptor ligand [3H]MK 912) was found when transfected bacteria cultures were grown at 30 degrees C for about 4 h after induction with isopropyl beta-D-thiogalactopyranoside (IPTG). Saturation experiments showed that the radioligand bound to a single saturable site with a Kd of 1.42 +/- 0.09 nM and capacity of 281 +/- 6 fmol/mg protein. Binding constants of 14 compounds for the rat alpha 2B-adrenoceptor expressed in E. coli were determined and compared to the values previously obtained for the rat alpha 2B-adrenoceptor when expressed in COS cells as well as for native neonatal rat lung alpha 2B-adrenoceptors. The results indicate that when the rat alpha 2B-adrenoceptor is expressed in E. coli it retains identical ligand binding properties to those found when the receptor is present in the eukaryotic system. Expressing alpha 2B-adrenoceptors in E. coli would, therefore, seem to constitute a valid alternative in, e.g., drug screening and structure analysis of the alpha 2B-adrenoceptors.