N‐Bromoacetyl‐amino‐cyanopindolol: a highly potent beta‐adrenergic affinity label blocks irreversibly a non‐protein component tightly associated with the receptor

A new chemical affinity label for the β‐adrenergic receptor, based on the structure of pindolol, has been synthesized and iodinated with ¹²⁵I. The compound, N‐bromoacetylamino‐cyanopindolol (BAM‐CYP), has an apparent dissociation constant of 44 ± 7 pM towards the turkey erythrocyte membranes. This compound blocks irreversibly both the ability of β‐adrenergic receptors to bind ¹²⁵I‐cyanopindolol and the ability of β‐receptors to activate adenylate cyclase in the presence of β‐agonists. Furthermore, the irreversible binding of BAM‐CYP to half of the β‐receptor sites abolishes the ligand binding activity of all the sites. These findings suggest that the β‐receptor is oligomeric in its native state. Although ¹²⁵I‐BAM‐CYP blocks irreversibly and specifically the β‐adrenergic receptor, it does so by labeling a non‐protein component, most probably a water‐soluble lipid. The labeling is stereospecific since it is prevented by l‐propranolol and not by d‐propranolol. It is suggested that this lipid is tightly associated with the receptor in close proximity to the binding site. It is also suggested that this water‐soluble lipid fraction may prove crucial for the optimal interaction between the β‐adrenergic receptor and the components of adenylate cyclase.