An antibody specific for the propranolol binding site of cardiac muscle.

The stereospecific binding site for propranolol in cardiac membranes was solubilized with 0.25% sodium deoxycholate. Affinity chromatography on norepinephrine-Sepharose yielded a fraction that bound (-)-propranolol with an association constant 360 times that of the (+)-enantiomer. A Scatchard plot indicated a single binding site with a dissociation constant for propran0101 of 11 + 4 nM and a concentration of 1100 + 280 pmol/mg of protein. An estimated ‘7000to 8000-fold purification of the binding site was achieved. Ligand binding properties of the purified material were similar to those observed for the intact membrane with respect to affinity and specificity. An antiserum raised to the affinity purified binding site preparation effectively inhibited both propranolol binding and the stimulation of adenylate cyclase by catecholamines in intact cardiac membranes. Half-maximal inhibition of cyclase stimulation and propranolol binding occurred at approximately the same concentrations of an immune globulin fraction of this antiserum. While the immune globulin inhibited isoproterenolstimulated adenylate cyclase activity, it had no effect on basal, guanyl-B-y1 imidodiphosphate, or fluoridestimulated cyclase activity in cardiac membranes. The enzymatic activity of detergent solubilized adenylate cyclase was also unaffected by antibody. The immune globulin did not inhibit adenylate cyclase activity in hepatic membranes, indicating a structural difference between adrenergic receptors in two tissues that had previously been shown to differ functionally with respect to their spectrum of specificity for agonists and antagonists.