Human β-adrenergic receptors. Simultaneous purification of β₁- and β₂-adrenergic-receptor peptides

β-Adrenergic receptors from basal membranes of human placenta were purified from digitonin extracts by sequential rounds of affinity chromatography, hydrophobic chromatography, ion-exchange chromatography and steric-exclusion h.p.l.c. Basal membranes display both β₁- and β₂-adrenergic receptors, in the ratio 65:35. Affinity chromatography, hydrophobic chromatography on heptylamine-Sepharose and ion-exchange chromatography on DEAE-Sephacel removed most of the contaminating proteins, and final purification of the receptor to apparent homogeneity was achieved by steric-exclusion h.p.l.c. The purified receptors showed M(r) 67,000 on SDS/polyacrylamide-gel electrophoresis under reducing conditions. Specific binding of radioligand to the purified β-adrenergic receptors displayed stereoselectivity, and the agonist competition profiles demonstrated the presence of both β₁- and β₂-receptors. By using the subtype-selective ligands CGP-20712A (β₁-selective) and ICI-118,551 (β₂-selective), the purified M(r)-67,000 species was shown to be composed of equivalent amounts of β₁- and β₂-adrenergic receptors. Affinity chromatography on Sepharose-alprenolol and sequential elution with 1 μM-CGP-20712A followed by 100 μM-(-)-alprenolol permitted β₁-adrenergic receptors to be resolved from the mixture of β₁-/β₂-adrenergic receptors. The pharmacologically distinct human β₁- and β₂-adrenergic receptors are shown to be structurally very similar peptides.