A principle tenet in pharmacology is that neurotransmitters, drugs, and hormones interact with specific receptor sites to produce or inhibit a characteristic physiological response. In the case of synaptic transmission and many hormone responsive systems, these specific receptor sites are presumed to be integral membrane proteins. The purification folds obtained by the methods represent a 6500-fold purification from initial detergent extracts and a greater than 80000-fold purification from membrane preparations. As the techniques were found to be extremely successful for the purification of the β2-adrenergic receptor, we sought to purify the β1-adrenergic receptor from the turkey red blood cell. Purification and characterization of hormone and drug receptors has been a difficult task primarily due to the extremely small quantities of these macromolecules in most cell types. β-Adrenergic receptors are closely coupled to the enzyme adenylate cyclase and have been the focus of considerable attention.

Turkey red blood cell (RBC), Beta/sub 1/-adrenergic receptors (Bar) were prepared to electrophoretic homogeneity and denatured protein used to prepare rabbit anti-Bar antibodies. Anti-Bar activity was confirmed by immuno-adsorption of (/sup 125/I) cyanopindolol (CYP) labeled Bar. The catecholamine biosynthetic enzyme dopamine beta hydroxylase (DBH) was purified from bovine adrenal medullae chromaffin vesicles by ion exchange, size exclusion and concanavalin-A-Sepharose chromatography. Final DBH specific activities were 42 +/- 4 U/mg protein. Homogeneity was confirmed by non-denaturing PAGE. Bar was compared to DBH by anti-Bar antibody cross-reactivity. DBH and Bar were recognized by anti-Bar antibodies on immunoblotting. No interactions were observed with preimmune controls. Similar results were obtained with glycosylated and deglycosylated DBH suggesting that the antibodies recognize DBH amino acid sequence and not associated carbohydrate. Cross-reactive antibodies were purified by affinity chromatography using immobilized DBH and shown to immuno-adsorb (/sup 125/I)CYP labeled Bar. These results suggest that the catecholamine biosynthetic enzyme DBH and Bar may be related in structure.