IMPROVED MEMBRANE ISOLATION IN THE PURIFICATION OF BETA(2)-ADRENOCEPTORS FROM TRANSGENIC ESCHERICHIA-COLI

beta(2)-Adrenoceptors (beta(2)-AR) have been purified from many mammal ian tissues. Unfortunately, other beta-AR subtypes expressed in the sa me cells are usually copurified, contaminating the preparation and int erfering with subsequent investigations such as receptor characterizat ion, ligand binding studies, immunoprecipitation, or development of an ti-receptor antibodies. The advent of molecular biology techniques has facilitated the expression of beta(2)-AR in cells in which no other s imilar molecules are present; thus, receptor purification has been sim plified, beta(2)-AR expressed in Escherichia coli provides a convenien t source of receptor without the need for specialized culture faciliti es required for eukaryotic cells. The greater complexity of the gram-n egative cell wall structure, however, complicates the purification of membrane-bound receptor from this source, in this report, we describe a reliable method for the partial purification of membrane-bound beta( 2)-AR from transgenic E. coli. Spheroplast formation followed by cell disruption and a carbonate wash procedure provided beta(2)-AR bound to bacterial inner membrane in high yield. (C) 1997 Academic Press.